Putative tumor suppressor EDD interacts with and up-regulates APC

Adenomatous polyposis coli (APC), whose mutation causes colorectal cancers, is a key player in the Wnt signaling pathway. While the role of APC in inhibition of beta-catenin/LEF1-dependent activation of transformation-inducing genes has been intensively studied and well established, regulation of APC expression at the protein level is only partially understood. Here we report that APC is up-regulated by EDD, the mammalian orthologue of Drosophila melanogaster"hyperplastic discs" gene (hyd) that is considered to be a putative tumor suppressor. Screening of APC immunocomplexes by mass spectrometry identified EDD as a putative APC-interacting protein. Exogenously expressed and endogenous APC interacted with EDD in vivo. Indirect immunofluorescent analyses demonstrated that APC and EDD co-localized in the cytoplasm of the cell. Over-expression of EDD enhanced the protein expression level of APC and its binding partner Axin, resulting in inhibition of Wnt signaling downstream of beta-catenin. Conversely, siRNA knock-down of EDD down-regulated APC at the protein level without altering its mRNA level, causing enhanced protein expression of beta-catenin. Thus, through protein-protein interaction, EDD stabilizes APC and up-regulates APC's function to inhibit beta-catenin, suggesting that EDD could act as a colorectal tumor suppressor.     

Tracheal Aspirate as an Alternative Biologic Sample for Pharmacogenomics Testing in Mechanically Ventilated Pediatric Patients

Patients in the pediatric intensive care unit are exposed to multiple medications and are at high risk for adverse drug reactions. Pharmacogenomic (PGx) testing could help decrease their risk of adverse reactions. Although whole blood is preferred for PGx testing, blood volume in this population is often limited. However, for patients on mechanical ventilation, tracheal secretions are abundant, frequently suctioned, and discarded. Thus, the aim of this pilot study was to determine if tracheal aspirates could be used as a source of human genomic DNA for PGx testing. We successfully extracted DNA from tracheal secretions of all 23 patients in the study. The samples were successfully genotyped for 10 clinically actionable single nucleotide variants across 3 cytochrome P450 genes (CYP2D6, CYP2C19, and CYP3A5). Using DNA from whole blood samples in 11 of the patients, we confirmed the accuracy of the genotyping with 100% concordance. Therefore, our results support the use of tracheal aspirates from mechanically ventilated children as an adequate biospecimen for clinical genetic testing.

Children admitted to the pediatric intensive care unit (PICU) receive an average of 10 different medications per day and receive an average cumulative 20 medications during their stay. ¹ Many of these have clinical guidelines for using pharmacogenomic (PGx) testing to guide the drug choice and/or dose, including voriconazole, selective serotonin reuptake inhibitors, tricyclic antidepressants, and codeine. ² , ³ , ⁴ , ⁵ The US Food and Drug Administration (FDA) has placed genetic testing recommendations and black box warnings on over 100 drug labels, including guidelines on gene‐drug pairs. The Clinical Pharmacogenetics Implementation Consortium (CPIC) has published guidelines for 35 medications with plans for writing over 100 more. ⁶ These initiatives are indicative of the importance of PGx to prescribing practices. The use of PGx has not been routinely utilized in the PICU, but as more evidence and guidelines for PGx‐guided dosing emerges, this could play an important role in the safe delivery of medications to patients in the PICU.The preferred biospecimen for PGx testing is whole blood but saliva and buccal swabs have also been validated and used as an alternative source of DNA. ⁷ , ⁸ , ⁹ , ¹⁰ , ¹¹ However, in small patients, blood volumes can be limited and mechanical ventilation or sedation can impair the collection of saliva or buccal swabs. In pediatrics, most current blood sampling guidelines are not evidence based; so, it is generally recommended to use alternatives to blood to ensure patient safety and maximize clinical benefit. ¹² , ¹³ In addition, blood samples cannot be used for genotyping from children that have had stem cell transplants or recent blood transfusions. Therefore, there is a need to identify an alternative source of DNA that can be used for validating biomarkers in clinical trials and conducting clinical PGx testing.Many patients admitted to the PICU are mechanically ventilated. When they are vented, the patient’s respiratory secretions are frequently suctioned and discarded. As this biological sample is plentiful, easily accessible, and collection is noninvasive, we hypothesized that tracheal aspirates from mechanically ventilated patients could be used as a source of DNA for clinical genotyping. Although one might assume that this would work, there could be too little DNA or it could be to degraded to use for genotyping; so, the feasibility needs to be supported by laboratory data. Thus, the aim of this study was to determine if tracheal aspirates from mechanically ventilated pediatric patients can be used as a source of DNA for PGx testing.     

Control of the inflow and outflow system during liver resection

Control of blood loss is a serious problem during liver resection. Bleeding from the inflow system can be controlled by the Pringle maneuver. The time limit for clamping is up to 10-15?min. A shortcoming of the Pringle maneuver is that it causes blood congestion in the portal vein. To avoid this problem other techniques have been developed including selective vascular occlusion and selective clamping of segmental branches. Bleeding from the outflow system is closely related to central venous pressure (CVP). Lowering the CVP reduces blood loss; in particular, keeping CVP <5?cmH(2)O by anesthesiological management is a simple and effective way to reduce blood loss. CVP remains high in some cases despite anesthesiological efforts, but in these circumstances other techniques are available including inferior vena cava clamping below the liver and intraoperative blood salvage.     

Liver regeneration and restoration of liver function after partial hepatectomy: the relation of fibrosis of the liver parenchyma

BACKGROUND/AIMS: Patients who survive partial hepatectomy sometimes have unsatisfactory liver regeneration and restoration of liver function. Although the extent of resection should be adjusted to attain favorable liver regeneration and restoration of liver function, a guiding principle for this has not been established. METHODOLOGY: Seventy patients with hepatic tumors associated with liver disorders of various severity who underwent hepatectomy were studied. We calculated the removal rate of the liver and the regeneration rate of the remnant liver using computed tomography. The liver function was investigated using ICG R-15 (retention rate of indocyanine green). Liver disorder was classified into 4 groups, according to the severity of fibrosis. RESULTS: The regeneration rates of the remnant liver indicated a significant decline in patients with severe fibrosis. In the no fibrosis and mild fibrosis groups, an increased removal rate was associated with increased regeneration rate, and post-operative ICG R-15 improved with time. However, in the moderate fibrosis and severe fibrosis groups, an increased removal rate was not associated with increased regeneration rate, and post-operative ICG R-15 showed no change or became worse with time. CONCLUSIONS: Severe fibrosis of the liver parenchyma is associated with poorer regeneration of the remnant liver leading to poor restoration of post-operative liver function. The severity of fibrosis is useful as a predictive factor for liver regeneration and restoration of liver function after partial hepatectomy.     

Increment of Tal positive cells in peripheral blood from patients with rheumatoid arthritis

Recently it was demonstrated that the Tal antigen is a marker for activated T cells and the population of Tal+ cells includes memory T cells. Our study was undertaken to analyze Tal+ cells in peripheral blood from patients with rheumatoid arthritis (RA) and examine the correlation between the proportion of Tal+ cells and disease activity. Using a dual immunofluorescent method, T cell subsets were analyzed in peripheral blood from 46 patients with RA, 10 patients with osteoarthritis (OA) and 10 healthy subjects. Patients with RA had a significantly higher percentage of T cells bearing the Tal antigen than patients with OA and healthy subjects. Furthermore, the percentage of CD3+Tal+ cells in peripheral blood from patients with RA was significantly correlated with disease activity including erythrocyte sedimentation rate, morning stiffness and the Lansbury index, whereas CD3+ HLA-DR+ cells did not correlate. In serial observations in patients with RA, the percentage of CD3+ Tal+ cells decreased in accordance with the improvement of disease activity. Thus, we suggest that the percentage of Tal+ cells may be a marker of the disease activity in patients with RA.     

Left ventricular hypertrophy is associated with arterial stiffness and vascular calcification in hemodialysis patients.

Left ventricular hypertrophy (LVH) is the most frequent cardiac abnormality in patients with end-stage renal disease (ESRD). Recent studies have shown that arterial stiffness is associated with mediacalcinosis in these patients. However, whether arterial stiffness and vascular calcification are associated with the LVH in patients with ESRD has not been well established. Forty-nine patients on chronic hemodialysis participated in this study. 1) To better understand the mechanism underlying the increased incidence of LVH, we studied the relation between LVH and each of arterial wall stiffness, aortic calcification, and numerous clinical parameters in 49 patients on chronic hemodialysis. 2) To evaluate the contribution of arterial stiffness and arterial calcification to LVH in hemodialysis patients, we performed the present clinical analysis on 49 patients on chronic hemodialysis. We used an automatic device to measure arterial pulse wave velocity (PWV) as an index of arterial wall stiffness. The aortic calcification index (ACI) was quantified morphometrically by CT scan. The left ventricular mass index (LVMI) was estimated by M-mode echocardiography. To understand the mechanism underlying the increased incidence of LVH, we examined the factors contributing to LVMI in these patients. The correlation between each of the study parameters and LVMI as an indicator of LVH was then examined. The LVMI value was correlated positively with PWV (r=0.439, p=0.0014), systolic blood pressure (r=0.421, p=0.0023), and ACI (r=0.467, p=0.0006). A stepwise linear regression analysis showed that PWV, systolic blood pressure, and ACI were independently associated with LVH in our subjects. These results suggest that LVH is associated with hypertension, increased arterial stiffness, and the extent of vascular calcification in hemodialysis patients, with vascular calcification being the most important contributor to the development of LVH. Alteration of pulsatile dynamics contributes to an increase in left ventricular load and thus is also related to the LVH in these patients. These results suggest that LVH is associated with hypertension, increased arterial stiffness, and the extent of vascular calcification in hemodialysis patients. Vascular calcification, which alters the pulsatile dynamics and thereby contributes to an increase in left ventricular load, is the most important contributor to the development of LVH in patients undergoing hemodialysis.     

Portable-type signal-averaged electrocardiography with dipyridamole to detect patients with coronary artery disease.

BACKGROUND: In a retrospective study portable-type signal-averaged electrocardiography (SAECG) with dipyridamole stress was found to identify patients with coronary artery disease (CAD) at their bedside with high sensitivity and specificity, so the utility of this method was prospectively investigated in the present study. METHODS AND RESULTS: Standard 12-lead QRS wave SAECG was performed before and after dipyridamole stress at the bedside in 71 patients with chest pain (43 males, mean age 63 +/-9 years). The filtered QRS duration (fQRSd) before and after dipyridamole stress was determined by multiphasic oscillation method for each of the standard 12 leads, and the maximal value of changes in fQRSd (MAX DeltafQRSd) among the 12 leads was determined. The positive test was defined as MAX DeltafQRSd >or=5 ms, and negative as MAX DeltafQRSd <5 ms based on the previous study. Selective coronary arteriography was performed next. In the positive group (n=31), 25 patients had significant stenosis of the coronary artery and 6 did not. In the negative group (n=40), 5 patients had significant stenosis and 35 did not. The sensitivity, specificity, positive predictive accuracy and negative predictive accuracy for CAD detection by SAECG was 83%, 85%, 81% and 88%, respectively. CONCLUSIONS: Dipyridamole-stress portable SAECG is useful for detecting CAD at the patient's bedside with high sensitivity and specificity.