Hepatopancreatoduodenectomy with delayed division of the pancreatic parenchyma when utilizing a right lateral approach to the superior mesenteric artery.

BACKGROUND: Hepatopancreatoduodenectomy (HPD) is a high-risk surgical procedure. Delayed division of the pancreatic parenchyma (DDPP) was reported as a novel technique in HPD for reducing postoperative pancreatic fistula. However, it is often difficult to dissect the pancreatic head nerve plexus while leaving the pancreatic parenchyma intact, particularly in patients with a bulky tumor with vascular invasion. Of the various reported approaches to the superior mesenteric artery, the right lateral approach can provide a useful surgical field to conduct DDPP in HPD. CASE PRESENTATION: A 78-year-old man visited a local clinic with itching and jaundice. Laboratory tests revealed elevated hepatobiliary enzyme, total bilirubin, and tumor markers. Enhanced computed tomography, endoscopic retrograde cholangiopancreatography, and intraductal ultrasonography of the bile duct were performed, and he was diagnosed with perihilar cholangiocarcinoma with invasion to the right hepatic artery (40 × 15 mm, Bismuth IIIa, cT3N0M0 cStage III). After neoadjuvant chemotherapy, he underwent left hepatectomy with caudate lobectomy, pancreatoduodenectomy, and combined resection of right hepatic artery using DDPP with a right lateral approach to the superior mesenteric artery. The pathological diagnosis was perihilar cholangiocarcinoma ypT3N1M0 ypStage IIIC, R0 resection. He was discharged on postoperative day 57 in good health and has been doing well for 6 months since the surgery. CONCLUSIONS: We present an effective application of the right lateral approach to the superior mesenteric artery in DDPP during HPD. This procedure can provide a clear surgical field to easily divide the pancreatic head nerve plexus before transection of the pancreatic parenchyma.

Proteomic analysis of serum extracellular vesicles reveals Fibulin-3 as a new marker predicting liver-related events in MASLD.

BACKGROUND: There is a need for novel noninvasive markers for metabolic dysfunction-associated steatotic liver disease (MASLD) to stratify patients at high risk for liver-related events including liver cancer and decompensation. In the present study, we used proteomic analysis of proteins in extracellular vesicles (EVs) to identify new biomarkers that change with fibrosis progression and can predict the development of liver-related events. METHODS: We analyzed serum EVs from 50 patients with MASLD assessed for liver fibrosis by biopsy and identified proteins that altered with advanced fibrosis. A further evaluation was conducted on another cohort of 463 patients with MASLD with biopsy. RESULTS: Eight candidate proteins were identified by proteomic analysis of serum EVs. Among them, serum levels of Fibulin-3, Fibulin-1, and Ficolin 1 correlated with their EV levels. In addition, serum Fibulin-3 and serum Fibulin-1 levels changed significantly with advanced fibrosis. Using another cohort with biopsy, we found that the serum Fibulin-3 concentration was significantly greater in those with advanced fibrosis but that the serum Fibulin-1 concentration was not significantly different. Multivariate Cox proportional hazards analysis revealed that a higher Fibrosis-4 (FIB-4) index and higher serum Fibulin-3 concentration were independent risk factors for liver-related events. When the cutoff value for the serum Fibulin-3 concentration was 6.0 µg/mL according to the Youden index of AUROCs, patients with high serum Fibulin-3 significantly more frequently developed liver-related events than did other patients. Validation using another cohort of 226 patients with clinically diagnosed MASLD confirmed that high serum Fibulin-3 levels are associated with a greater frequency of liver-related events. CONCLUSIONS: Serum Fibulin-3 was identified as a biomarker for predicting liver-related events in patients with MASLD.

Noninvasive transthoracic doppler flow velocity and invasive thermodilution to assess coronary flow reserve.

Background: Coronary flow reserve (CFR) provides prognostication and coronary physiological information, including epicardial coronary stenosis and microvascular function. The relationship between stress transthoracic Doppler echocardiography (TDE)-derived coronary flow velocity reserve (CFRS-TDE) and thermodilution-derived coronary flow reserve (CFRthermo) before and after elective percutaneous coronary intervention (PCI) remains unclear. Methods: This single-center prospective registry study evaluated patients who underwent fractional flow reserve (FFR)-guided elective PCI for left anterior descending artery (LAD) lesions with wire-based invasive physiological measurements and pre- and post-PCI stress TDE examinations. Results: A total of 174 LAD lesions from 174 patients were included in the final analysis. A modest correlation was detected between the pre-PCI CFRS-TDE and the pre-PCI CFRthermo (r=0.383, P<0.001). The frequently used CFRS-TDE threshold of 2.0 corresponded to a pre-PCI CFRthermo of 2.18. Pre-PCI CFRS-TDE underestimated pre-PCI CFRthermo [1.89 (1.44-2.31) vs. 2.05 (1.38-2.93), P<0.001]. Both CFRS-TDE and CFRthermo increased significantly post-PCI [pre-PCI CFRS-TDE 1.89 vs. post-PCI CFRS-TDE 2.33, P<0.001; pre-PCI CFRthermo 2.05 (1.38-2.93) vs. post-PCI CFRthermo 2.59 (1.63-3.55), P<0.001]. In contrast, there was no significant relationship between changes in CFRS-TDE and changes in CFRthermo after PCI (r=0.008, P=0.915) or between post-PCI CFRS-TDE and post-PCI CFRthermo (r=0.054, P=0.482). Conclusions: Pre-PCI CFRS-TDE and CFRthermo are modestly correlated, but post-PCI CFRS-TDE and CFRthermo have no correlation. CFRS-TDE and CFRthermo are not interchangeable, particularly post-PCI, suggesting that the two metrics represent different coronary physiologies after PCI.

Impact of radiological and pathological splenic vein involvement in patients with resectable pancreatic body or tail cancer.

PURPOSE: Several studies have reported a negative impact on survival associated with splenic vessel involvement, especially splenic artery (SpA) involvement, in patients diagnosed with pancreatic body or tail cancer. However, there is limited research on splenic vein (SpV) involvement. Therefore, we aimed to elucidate the significance of splenic vessel involvement, especially SpV involvement, in patients with resectable pancreatic body or tail cancer. METHODS: Between January 2007 and December 2021, 116 consecutive patients underwent distal pancreatectomies for pancreatic body or tail cancer. Among them, this study specifically examined 88 patients with resectable pancreatic body or tail cancer to elucidate prognostic factors using a multivariable Cox proportional analysis. The Kaplan-Meier method evaluated the impact of SpV involvement in terms of both radiological and pathological aspects and the efficacy of neoadjuvant therapy. RESULTS: Higher pre-operative carcinoembryonic antigen levels, larger tumour size, pathological SpV invasion, and non-completion of adjuvant therapy were identified as independent poor prognostic factors for overall survival (OS) and recurrence-free survival (RFS). Additionally, patients with radiological SpV encasement had significantly worse prognoses in terms of OS (p = 0.039) and RFS (p < 0.001). The sensitivity and specificity of multidetector-row computed tomography for detecting pathological SpV invasion were 81.0% and 61.2%, respectively. However, the prognostic impact of neoadjuvant therapy could not be determined, regardless of radiological SpV involvement. CONCLUSION: Radiological and pathological SpV involvement is a poor prognostic factor for patients with resectable pancreatic body or tail cancer. New innovative treatments and effective neoadjuvant therapy regimens are required for patients with SpV involvement.

Diagnostic value of computed tomography myocardial perfusion imaging to detect coexisting microvascular dysfunction in patients with obstructive epicardial coronary artery disease.

Background: Computed tomography myocardial perfusion (CT-MP) has reported usefulness in assessing hemodynamically significant epicardial coronary artery lesions. However, the diagnostic ability of the absolute coronary flow using CT-MP to detect coronary microvascular dysfunction (CMD) remains elusive. This prospective cohort study aimed to assess the diagnostic value of CT-MP in evaluating coexisting CMD in patients with functionally significant epicardial coronary stenosis and to analyze the predictive factors of lesions with CMD. Methods: Sixty-eight patients with chronic coronary syndrome (CCS) and de novo single functionally significant stenosis [fractional flow reserve (FFR) ≤0.80] were studied. CMD was defined as an index of microcirculatory resistance ≥25. We compare clinical background and CT-MP findings between patients with and without CMD (CMD, n=29; non-CMD, n=39). CT-MP, and quantitative and qualitative plaque assessments were included in computed tomography angiography assessment. Logistic regression analysis was performed to predict CMD. Results: FFR, invasive wire-derived coronary flow reserve (CFRwire) and index of microcirculatory resistance were 0.68 [interquartile range (IQR), 0.59-0.74], 1.71 (IQR, 1.24-2.88), and 22.6 (IQR, 15.1-34.5), respectively. The rest and hyperemic-myocardial blood flow (MBF) and CT-MP-derived CFR (CFRCT-MP) were 0.83 (0.64-1.03) mL/min/g, 2.14 (1.30-2.92) mL/min/g, and 2.19 (1.44-3.37), respectively. In the territories with CMD, hyperemic-MBF was significantly lower than in those without [1.68 (IQR, 0.84-2.44) vs. 2.31 (IQR, 1.67-3.34) mL/min/g, P=0.015] and the prevalence of CFRCT-MP <2.0 was higher in the lesions with CMD than in those without (62.1% vs. 28.2%, P=0.011), while FFR values were similar. Fibrofatty and necrotic core component volume was greater in the vessels with CMD than in those without [31.8 (IQR, 19.0-48.9) vs. 25.1 (IQR, 17.2-32.1) mm3, P=0.045]. Multivariable logistic regression analysis showed that hyperemic-MBF and fibrofatty and necrotic core component volume were independent predictors of CMD territories [odds ratio (OR) =0.583; 95% confidence interval (CI): 0.355-0.958; P=0.033 and OR =1.040; 95% CI: 1.010-1.070; P=0.011]. Conclusions: Quantitative assessment of absolute coronary flow using pre-percutaneous coronary intervention (PCI) CT-MP, and comprehensive plaque analysis using computed tomography angiography may help detect coexisting subtended microvascular dysfunction in territories with functionally significant epicardial coronary lesions. Further studies are required to elucidate the clinical significance of coexisting CMD in patients with CCS undergoing PCI.

Substituted kynurenic acid derivatives as fluorophore-based probes for D- and L-amino acid oxidase assays and their in vitro application in eels.

High levels of D-amino acid oxidase (DAO) are associated with neurological and psychiatric disorders, while L-amino acid oxidase (LAO) exhibits antimicrobial and antitumor properties. The enzymatic conversion of the non-fluorescent kynurenine (KYN) into the endogenous weak fluorescent kynurenic acid (KYNA) by the action of DAO has previously been reported. However, the fluorescence of KYNA can be improved by changing the substituents on the aromatic rings. In this study, we prepared different 6-phenyl-substituted KYNA derivatives and investigated their fluorescence properties. Among them, 2-MePh-KYNA showed the maximum fluorescence quantum yield of 0.881 at 340 nm excitation and 418 nm emission wavelengths. The effects of solvent properties (dielectric constant, pKa, viscosity, and proticity) on the fluorescence intensity (FLI) of the KYNA derivatives were explored. The FLI of 2-MePh-KYNA was significantly large in protic solvents. Subsequently, 2-MePh-D-KYN and 2-MePh-L-KYN were prepared with high enantiopurity (>99.25%) for the enzymatic conversion. 2-MePh-D-KYN exhibited high sensitivity (∼19 times that of a commercial DAO substrate and ∼60 times that of the previously reported MeS-D-KYN) and high selectivity, as it was not cross-reactive towards LAO, while 2-MePh-L-KYN was also converted into 2-MePh-KYNA by LAO. Furthermore, the 2-MePh-D-KYN probe successfully detected DAO in eel liver, kidney, and heparin-anticoagulated plasma in the in vitro study.

Antithrombotic drug use does not affect the fecal immunochemical test PPV for colorectal cancer, but warfarin may have an impact in a Japanese cohort.

BACKGROUND: The fecal immunochemical test (FIT) is used for colorectal cancer (CRC) screening. Patients on antithrombotic drugs (ATs) are often screened for CRC, but the effect of ATs on FIT results is controversial. METHODS: We divided individuals with FIT-positive results into two groups, patients treated with and without ATs, and retrospectively compared invasive CRC rates, advanced neoplasia detection rates (ANDRs), adenoma detection rates (ADRs), and polyp detection rates (PDRs) between the two groups. We evaluated the factors influencing the FIT positive predictive value (PPV) using propensity matching, adjusting for age, sex, and bowel preparation. RESULTS: We enrolled 2327 individuals (54.9% male; mean age, 66.7  ± â€Š12.7 years). We grouped 463 individuals into the AT user group and 1864 into the nonuser group. Patients in the AT user group were significantly older and more likely to be male. After propensity score matching for age, sex, and Boston bowel preparation scale, the ADR and PDR in the AT user group were significantly lower than those in the nonuser group. Univariate logistic analysis revealed that multiple AT use (odds ratio [OR]: .39, p < 0.001) had the lowest OR for FIT PPV, followed by age- and sex-adjusted factors for the ADR and any AT use (OR: .67, p = 0.0007). No significant factors related to AT use were observed among age-adjusted predictive factors for invasive CRC, but warfarin use was a borderline significant positive predictive factor (OR: 2.23, p = 0.059). CONCLUSION: AT use may not affect the PPV for detecting invasive CRC in patients with positive FIT results, but warfarin may have an impact.

STAT3 is Activated by CTGF-mediated Tumor-stroma Cross Talk to Promote HCC Progression.

BACKGROUND & AIMS: Signal transducer and activator of transcription 3 (STAT3) is known as a pro-oncogenic transcription factor. Regarding liver carcinogenesis, however, it remains controversial whether activated STAT3 is pro- or anti-tumorigenic. This study aimed to clarify the significance and mechanism of STAT3 activation in hepatocellular carcinoma (HCC). METHODS: Hepatocyte-specific Kras-mutant mice (Alb-Cre KrasLSL-G12D/+; KrasG12D mice) were used as a liver cancer model. Cell lines of hepatoma and stromal cells including stellate cells, macrophages, T cells, and endothelial cells were used for culture. Surgically resected 12 HCCs were used for human analysis. RESULTS: Tumors in KrasG12D mice showed up-regulation of phosphorylated STAT3 (p-STAT3), together with interleukin (IL)-6 family cytokines, STAT3 target genes, and connective tissue growth factor (CTGF). Hepatocyte-specific STAT3 knockout (Alb-Cre KrasLSL-G12D/+ STAT3fl/fl) downregulated p-STAT3 and CTGF and suppressed tumor progression. In coculture with stromal cells, proliferation, and expression of p-STAT3 and CTGF, were enhanced in hepatoma cells via gp130/STAT3 signaling. Meanwhile, hepatoma cells produced CTGF to stimulate integrin/nuclear factor kappa B signaling and up-regulate IL-6 family cytokines from stromal cells, which could in turn activate gp130/STAT3 signaling in hepatoma cells. In KrasG12D mice, hepatocyte-specific CTGF knockout (Alb-Cre KrasLSL-G12D/+ CTGFfl/fl) downregulated p-STAT3, CTGF, and IL-6 family cytokines, and suppressed tumor progression. In human HCC, single cell RNA sequence showed CTGF and IL-6 family cytokine expression in tumor cells and stromal cells, respectively. CTGF expression was positively correlated with that of IL-6 family cytokines and STAT3 target genes in The Cancer Genome Atlas. CONCLUSIONS: STAT3 is activated by CTGF-mediated tumor-stroma crosstalk to promote HCC progression. STAT3-CTGF positive feedback loop could be a therapeutic target.

Efficacy and Safety of a Strategy for Reviewing Intravenous Antibiotics for Hospitalized Japanese Patients with Uncomplicated Diverticulitis: A Single-center Observational Study.

Objective Treatment for uncomplicated diverticulitis (UD) is not well established. We evaluated the strategy of reviewing intravenous antibiotics for hospitalized Japanese patients with UD. Methods Treatment was based on the physician's choice until August 2018; the indications for hospitalization and treatment have been standardized since September 2018. In this study, we monitored the use of intravenous antibiotics administered to patients hospitalized for UD and then reviewed the need for them on hospital day 3. We compared patients' length of antibiotic use, hospital stay, health care cost, and complications via the review strategy from September 2018 to December 2020 and via the previous physicians' choice strategy from January 2016 to August 2018. Results Two hundred and forty-seven patients were admitted to our hospital because of acute colonic diverticulitis from January 2016 to December 2020. After excluding complicated cases, 106 individuals were enrolled during the period of physician's choice; 87 were enrolled when treatment review was employed. There were no significant differences in age, sex, inflammation site, or severity during the first hospital visit. The median duration of antibiotic use was significantly reduced from 5 to 4 days (p=0.0075), with no marked increase in rates of transfer to surgery, mortality, or readmission due to recurrence. A more significant proportion of patients completed 3-day antibiotic treatment with the review strategy than with the physician's choice strategy (6.6% vs. 25.3%, p=0.0004). However, the length of hospital stay and total medical costs did not decrease. Conclusion The strategy of reviewing treatment on day 3 after hospitalization for UD safety reduced the duration of antibiotic use, but the hospital stay and health care costs did not decrease.

Asteltoxin inhibits extracellular vesicle production through AMPK/mTOR-mediated activation of lysosome function.

Cancer cells secrete aberrantly large amounts of extracellular vesicles (EVs) including exosomes, which originate from multivesicular bodies (MVBs). Because EVs potentially contribute to tumor progression, EV inhibitors are of interest as novel therapeutics. We screened a fungal natural product library. Using cancer cells engineered to secrete luciferase-labeled EVs, we identified asteltoxin, which inhibits mitochondrial ATP synthase, as an EV inhibitor. Low concentrations of asteltoxin inhibited EV secretion without inducing mitochondrial damage. Asteltoxin attenuated cellular ATP levels and induced AMPK-mediated mTORC1 inactivation. Consequently, MiT/TFE transcription factors are translocated into the nucleus, promoting transcription of lysosomal genes and lysosome activation. Electron microscopy analysis revealed that the number of lysosomes increased relative to that of MVBs and the level of EVs decreased after treatment with asteltoxin or rapamycin, an mTORC1 inhibitor. These findings suggest that asteltoxin represents a new type of EV inhibitor that controls MVB fate.

Immunoelectron Microscopic Characterization of Vasopressin-Producing Neurons in the Hypothalamo-Pituitary Axis of Non-Human Primates by Use of Formaldehyde-Fixed Tissues Stored at -25 °C for Several Years.

Translational research often requires the testing of experimental therapies in primates, but research in non-human primates is now stringently controlled by law around the world. Tissues fixed in formaldehyde without glutaraldehyde have been thought to be inappropriate for use in electron microscopic analysis, particularly those of the brain. Here we report the immunoelectron microscopic characterization of arginine vasopressin (AVP)-producing neurons in macaque hypothalamo-pituitary axis tissues fixed by perfusion with 4% formaldehyde and stored at -25 °C for several years (4-6 years). The size difference of dense-cored vesicles between magnocellular and parvocellular AVP neurons was detectable in their cell bodies and perivascular nerve endings located, respectively, in the posterior pituitary and median eminence. Furthermore, glutamate and the vesicular glutamate transporter 2 could be colocalized with AVP in perivascular nerve endings of both the posterior pituitary and the external layer of the median eminence, suggesting that both magnocellular and parvocellular AVP neurons are glutamatergic in primates. Both ultrastructure and immunoreactivity can therefore be sufficiently preserved in macaque brain tissues stored long-term, initially for light microscopy. Taken together, these results suggest that this methodology could be applied to the human post-mortem brain and be very useful in translational research.

Estrogens influence female itch sensitivity via the spinal gastrin-releasing peptide receptor neurons.

There are sex differences in somatosensory sensitivity. Circulating estrogens appear to have a pronociceptive effect that explains why females are reported to be more sensitive to pain than males. Although itch symptoms develop during pregnancy in many women, the underlying mechanism of female-specific pruritus is unknown. Here, we demonstrate that estradiol, but not progesterone, enhances histamine-evoked scratching behavior indicative of itch in female rats. Estradiol increased the expression of the spinal itch mediator, gastrin-releasing peptide (GRP), and increased the histamine-evoked activity of itch-processing neurons that express the GRP receptor (GRPR) in the spinal dorsal horn. The enhancement of itch behavior by estradiol was suppressed by intrathecal administration of a GRPR blocker. In vivo electrophysiological analysis showed that estradiol increased the histamine-evoked firing frequency and prolonged the response of spinal GRP-sensitive neurons in female rats. On the other hand, estradiol did not affect the threshold of noxious thermal pain and decreased touch sensitivity, indicating that estradiol separately affects itch, pain, and touch modalities. Thus, estrogens selectively enhance histamine-evoked itch in females via the spinal GRP/GRPR system. This may explain why itch sensation varies with estrogen levels and provides a basis for treating itch in females by targeting GRPR.

Comparison of colonoscopy after colonic diverticulitis and positive fecal immunochemical tests for the detection of colorectal neoplasia.

Background and study aims The relationship between acute colonic diverticulitis and colorectal cancer (CRC) is unclear, but colonoscopy is recommended to exclude malignancy. We compared the detection rates for colorectal neoplasia in patients with colonic diverticulitis and asymptomatic patients who had positive fecal immunochemical tests (FITs). Patients and methods In total, 282 patients with acute colonic diverticulitis were hospitalized in our hospital from February 2011 to December 2019. Of them, 143 patients with diverticulitis and 1819 with positive FITs patients during the same period underwent colonoscopy without a prior colonoscopy within 5 years. We retrospectively compared these patients in terms of the invasive CRC rate, advanced neoplasia detection rate (ANDR), adenoma detection rate (ADR), and polyp detection rate (PDR). Results Compared to the diverticulitis group, the FIT-positive group had a significantly higher CRC rate (0 vs 2.7 %, P  = 0.0061), ANDR (5.6 vs. 14.0 %, P  = 0.0017), ADR (19.6 vs. 53.2 %, P  < .0001), and PDR (44.1 vs. 91.0 %, P  < .0001). Using 1:1 propensity score matching based on age and sex, we obtained 276 matched patients in both groups. After matching, no difference was found in the CRC rate (0 vs 0.7 %) or ANDR (5.8 vs 7.3 %) between groups, but the ADR and PDR were significantly higher in the FIT-positive group (20.3 vs 43.5 %, P  < .0001; 45.7 % vs 86.2 %, P  < .0001). Conclusion Patients with acute diverticulitis had lower ADRs and PDRs than patients with positive FITs.

Radiation exposure dose of fluoroscopy-guided gastrointestinal procedures: A single-center retrospective study.

Background and study aims Fluoroscopy-guided gastrointestinal procedures (FGPs) are increasingly common. However, the radiation exposure (RE) to patients undergoing FGPs is still unclear. We examined the actual RE of FGPs. Patients and methods This retrospective, single-center cohort study included consecutive FGPs, including endoscopic retrograde cholangiopancreatography (ERCP), interventional endoscopic ultrasound (EUS), enteral stenting, balloon-assisted enteroscopy, tube placement, endoscopic injection sclerotherapy (EIS), esophageal balloon dilatation and repositioning for sigmoid volvulus, from September 2012 to June 2019. We measured the air kerma (AK, mGy), dose area product (DAP, Gycm 2 ), and fluoroscopy time (FT, min) for each procedure. Results In total, 3831 patients were enrolled. Overall, 2778 ERCPs were performed. The median AK, DAP, and FT were as follows: ERCP: 109 mGy, 13.3 Gycm 2 and 10.0 min; self-expandable enteral stenting (SEMS): 62 mGy, 12.4 Gycm 2 and 10.4 min; tube placement: 40 mGy, 13.8 Gycm 2 and 11.1 min; balloon-assisted enteroscopy: 43 mGy, 22.4 Gycm 2 and 18.2 min; EUS cyst drainage (EUS-CD): 96 mGy, 18.3 Gycm 2 and 10.4 min; EIS: 36 mGy, 8.1 Gycm 2 and 4.4 min; esophageal balloon dilatation: 9 mGy, 2.2 Gycm 2 and 1.8 min; and repositioning for sigmoid volvulus: 7 mGy, 4.7 Gycm 2 and 1.6 min. Conclusion This large series reporting actual RE doses of various FGPs could serve as a reference for future prospective studies.

LC-MS/MS Analysis of Thiol-Containing Amino Acids in Exosomal Fraction of Serum.

It has been suggested that thiol-containing amino acids could be used as biomarkers for diseases associated with oxidative stress. We investigated the thiol-containing amino acids, homocysteine (Hcy), cysteine (Cys), glutathione (GSH) and γ-glutamylcysteine (γ-GluCys), in commercial human serum by using liquid chromatography-tandem mass spectrometry (LC-MS/MS) after precolumn derivatization with 4-fluoro-7-sulfobenzofurazan. This method was applied to determine the composition of thiol-containing amino acids in exosomes prepared from the serum. Hcy, Cys, GSH and γ-GluCys could be detected in the exosomal fraction, and the ratio of each thiol-containing amino acid was similar to those in the corresponding native serum. Cys (94.76%) was most enriched in the exosomal fraction, followed by GSH (2.97%), γ-GluCys (1.59%) and Hcy (0.68%). These findings suggest that thiol-containing amino acids, Hcy, Cys, GSH and γ-GluCys, are included in exosomes in human serum.

Detection and Characterization of Estrogen Receptor Beta Expression in the Brain with Newly Developed Transgenic Mice.

Two types of nuclear estrogen receptors, ERα and ERß, have been shown to be differentially involved in the regulation of various types of behaviors. Due to a lack of tools for identifying ERß expression, detailed anatomical distribution and neurochemical characteristics of ERß expressing cells and cellular co-expression with ERα remain unclear. We have generated transgenic mice ERß-RFPtg, in which RFP was inserted downstream of ERß BAC promotor. We verified RFP signals as ERß by confirming: (1) high ERß mRNA levels in RFP-expressing cells collected by fluorescence-activated cell sorting; and (2) co-localization of ERß mRNA and RFP proteins in the paraventricular nucleus (PVN). Strong ERß-RFP signals were found in the PVN, medial preoptic area (MPOA), bed nucleus of the stria terminalis, medial amygdala (MeA), and dorsal raphe nucleus (DRN). In the MPOA and MeA, three types of cell populations were identified; those expressing both ERα and ERß, and those expressing exclusively either ERα or ERß. The majority of PVN and DRN cells expressed only ERß-RFP. Further, ERß-RFP positive cells co-expressed oxytocin in the PVN, and tryptophan hydroxylase 2 and progesterone receptors in the DRN. In the MeA, some ERß-RFP positive cells co-expressed oxytocin receptors. These findings collectively suggest that ERß-RFPtg mice can be a powerful tool for future studies on ERß function in the estrogenic regulation of social behaviors.

Cholecystohepatic duct detected during laparoscopic cholecystectomy: a case report.

BACKGROUND: The cholecystohepatic duct is a rare form of an aberrant hepatic duct that connects to the gallbladder. Although cholecystohepatic duct is reported to be a very rare anomaly, injury of cholecystohepatic duct during cholecystectomy may result in serious complications. Herein, we present a case of cholecystohepatic duct in the ventral branch of the right posterior inferior segmental bile duct detected during laparoscopic cholecystectomy. CASE PRESENTATION: A 77-year-old woman with cholecystolithiasis had been referred to our hospital for surgery. Drip infusion cholecystocholangiography-computed tomography revealed a bile duct branch without communication between the intra- and extrabiliary systems, although the existence of this aberrant hepatic duct was not suspected preoperatively. A 4-port laparoscopic cholecystectomy was performed. After critical view of safety was confirmed, the cystic artery and duct were divided after double clipping. During antegrade mobilization of the gallbladder from the gallbladder bed, a thin, white cord-like material connecting the gallbladder neck and bed was detected. After clipping and dividing it, a cholecystohepatic duct injury was recognized through rechecking the results of the preoperative examinations. Biliary reconstruction was considered unnecessary because of the lesion's small drainage area. The postoperative course was uneventful, and an enhanced computed tomography performed 6 months after the surgery revealed a dilation in the ventral branch of the right posterior inferior segmental bile duct. The patient's liver function remained normal, and she had no symptoms of cholangitis 42 months after the surgery. CONCLUSIONS: Although cholecystohepatic duct is a rare anomaly compared to other aberrant hepatic ducts, surgeons performing cholecystectomy should always keep its existence in mind to avoid serious postoperative complications. Ideally, preoperative detection of cholecystohepatic duct is preferable, but even if it is detected during surgery, the appropriate management according to the drainage area is also important.

Succinimidyl (3-[(benzyloxy)carbonyl]-5-oxo-1,3-oxazolidin-4-yl)acetate on a triazole-bonded phase for the separation of dl-amino-acid enantiomers and the mass-spectrometric determination of chiral amino acids in rat plasma.

Changes in the levels of amino-acid enantiomers are associated with some serious diseases; consequently, amino acid monitoring in peripheral blood can be used to diagnose and predict the onset of disease. Herein, we report the design and synthesis of a new chiral derivatization reagent, namely succinimidyl (4S)-(3-[(benzyloxy)carbonyl]-5-oxo-1,3-oxazolidin-4-yl)acetate ((S)-COXA-OSu), for the separation of dl-amino-acid enantiomers. The usefulness of (S)-COXA-OSu was examined as a derivatization reagent for LC-MS/MS following certification of its total optical purity (>99%). The enantiomeric separations of amino-acid derivatives tagged with the reagent were examined using a triazole-bonded phase. (S)-COXA-OSu enabled the simultaneous enantiomeric separation of more than 40 α-amino acids. (S)-COXA-amino-acid derivatives were efficiently converted into their product ions, from which formaldehyde (CH2O) was eliminated [M-30] from the oxazolidinone moiety of COXA by collision-induced dissociation during LC-MS/MS. Limits of detection were in the 0.0138-0.518 pmol/injection range. For precise and accurate quantitation, we synthesized and used a stable-isotope-labeled (S)-COXA-OSu that was used as an internal standard in LC-MS/MS-determination experiments. Finally, changes in plasma amino-acid levels in rats, following administration of S-methyl-l-cysteine, an alanine-serine-cysteine transporter-1 (Asc-1) inhibitor, were successfully detected by LC-MS/MS using (S)-COXA-OSu.

Amino acid analyses of the exosome-eluted fractions from human serum by HPLC with fluorescence detection.

OBJECTIVES: Amino acid levels in serum or plasma are used for early detection and diagnosis of several diseases. The objective of this study was to analyze amino acid levels in serum exosomes, which have not been previously reported. DESIGN AND METHODS: We investigated the amino acid composition of exosomes from human serum using HPLC with fluorescence detection. RESULTS: The composition ratios of His, Arg, Glu, Cys-Cys, Lys, and Tyr were significantly increased in the exosomes compared with those in the corresponding native serum. d-Ser, an endogenous co-agonist of the N-methyl-d-aspartate receptor, was also enriched in the exosome-eluted fraction. CONCLUSIONS: Our results suggest that certain amino acids are enriched in the exosome-eluted fraction from human serum. These differences could have future diagnostic potential.