A tagging-via-substrate technology for detection and proteomics of farnesylated proteins

A recently developed proteomics strategy, designated tagging-via-substrate (TAS) approach, is described for the detection and proteomic analysis of farnesylated proteins. TAS technology involves metabolic incorporation of a synthetic azido-farnesyl analog and chemoselective derivatization of azido-farnesyl-modified proteins by an elegant version of Staudinger reaction, pioneered by the Bertozzi group, using a biotinylated phosphine capture reagent. The resulting protein conjugates can be specifically detected and/or affinity-purified by streptavidin-linked horseradish peroxidase or agarose beads, respectively. Thus, the technology enables global profiling of farnesylated proteins by enriching farnesylated proteins and reducing the complexity of farnesylation subproteome. Azido-farnesylated proteins maintain the properties of protein farnesylation, including promoting membrane association, Ras-dependent mitogen-activated protein kinase kinase activation, and inhibition of lovastatin-induced apoptosis. A proteomic analysis of farnesylated proteins by TAS technology revealed 18 farnesylated proteins, including those with potentially novel farnesylation motifs, suggesting that future use of this method is likely to yield novel insight into protein farnesylation. TAS technology can be extended to other posttranslational modifications, such as geranylgeranylation and myristoylation, thus providing powerful tools for detection, quantification, and proteomic analysis of posttranslationally modified proteins.     

Effects of Coffee Intake on Incident Chronic Kidney Disease: A Community-Based Prospective Cohort Study

Background

Drinking coffee can raise public health problems, but the association between coffee and kidney disease is unknown. We studied whether coffee intake can affect the development of chronic kidney disease in the general population.

Physiologic Characteristics and Clinical Outcomes of Patients With Discordance Between FFR and iFR

ObjectivesThis study evaluated the physiologic characteristics of discordant lesions between instantaneous wave-free ratio (iFR) and fractional flow reserve (FFR) and the prognosis at 5 years.BackgroundFFR or iFR have been standard methods for assessing the functional significance of coronary artery stenosis. However, limited data exist about the physiologic characteristics of discordant lesions and the prognostic implications resulting from these lesions.MethodsA total of 840 vessels from 596 patients were classified according to iFR and FFR; high iFR–high FFR (n = 580), low iFR–high FFR (n = 40), high iFR–low FFR (n = 69), and low iFR–low FFR (n = 128) groups, which were compared with a control group (n = 23). The differences in coronary circulatory indices including the coronary flow reserve (CFR), index of microcirculatory resistance (IMR), and resistance reserve ratio (RRR) (resting distal arterial pressure × mean transit time / hyperemic distal arterial pressure × hyperemic mean transit time), which reflect the vasodilatory capacity of coronary microcirculation, were compared. Patient-oriented composite outcomes (POCO) at 5 years including all-cause death, any myocardial infarction, and any revascularization were compared among patients with deferred lesions.ResultsIn the low iFR–high FFR group, CFR, RRR, and IMR measurements were similar to the low iFR–low FFR group: CFR 2.71 versus 2.43 (p = 0.144), RRR 3.36 versus 3.68 (p = 0.241), and IMR 18.51 versus 17.38 (p = 0.476). In the high iFR–low FFR group, the CFR, RRR, and IMR measurements were similar to the control group: CFR 2.95 versus 3.29 (p = 0.160), RRR 4.28 versus 4.00 (p = 0.414), and IMR 17.44 versus 17.06 (p = 0.818). Among the 4 groups, classified by iFR and FFR, CFR and RRR were all significantly different, except for IMR. However, there were no significant differences in the rates of POCO, regardless of discordance between the iFR and FFR. Only the low iFR–low FFR group had a higher POCO rate compared with the high iFR–high FFR group (adjusted hazard ratio: 2.46; 95% confidence interval: 1.17 to 5.16; p = 0.018).ConclusionsDifferences in coronary circulatory function were found, especially in the vasodilatory capacity between the low iFR–high FFR and high iFR–low FFR groups. FFR–iFR discordance was not related to an increased risk of POCO among patients with deferred lesions at 5 years. (Clinical, Physiological and Prognostic Implication of Microvascular Status; NCT02186093; Physiologic Assessment of Microvascular Function in Heart Transplant Patients; NCT02798731)     

Prolongation of the period between biopsy and EMR can influence the nonlifting sign in endoscopically resectable colorectal cancers

BACKGROUND: The nonlifting sign is widely used for evaluating the invasion depth of colorectal tumors, and it is commonly accepted that EMR is contraindicated for colorectal tumors with a nonlifting sign because of the probability of massive submucosal invasion. OBJECTIVE: To identify the clinicopathologic factors that affect the nonlifting sign in submucosal invasive colorectal carcinoma (SICC). DESIGN: Details regarding a history of biopsy, postbiopsy days, tumor location, tumor configuration, tumor size, depth of submucosal invasion, histologic type, adenomatous remnants, and angiolymphatic invasion were studied in relation to the nonlifting sign. SETTING: National Cancer Center, Korea. PATIENTS: The study involved 76 patients with SICC treated by endoscopic or surgical resection, in whom the tumor was examined for the nonlifting sign from 2001 to 2006. RESULTS: The nonlifting sign was observed in 15 cases (19.7%). A deep submucosal invasion, a history of biopsy, and the absence of adenomatous remnants were identified as factors affecting the nonlifting sign in univariate and multivariate analyses (P < .05). An increase in the number of postbiopsy days was associated with the nonlifting sign in endoscopically resectable SICC, and all 11 sm1 cancer cases with fewer than 21 postbiopsy days showed lifting. CONCLUSIONS: A history of biopsy and the absence of adenomatous remnants, in addition to deep submucosal invasion, were found to influence the nonlifting sign in SICC. It may be best that mechanical stimulation such as forceps biopsies are minimized before EMR, and EMR should be tried as soon as possible if biopsy was performed.     

An Arabidopsis histone H2A mutant is deficient in Agrobacterium T-DNA integration

Agrobacterium tumefaciens genetically transforms plant cells by transferring a portion of the bacterial Ti-plasmid, the T-DNA, to the plant and integrating the T-DNA into the plant genome. Little is known about the T-DNA integration process, and no plant genes involved in integration have yet been identified. We characterized an Arabidopsis mutant generated by T-DNA insertional mutagenesis, rat5, that is resistant to Agrobacterium root transformation. rat5 contains two copies of T-DNA integrated as a tandem direct repeat into the 3' untranslated region of a histone H2A gene, upstream of the polyadenylation signal sequence. Transient and stable beta-glucuronidase expression data and assessment of the amount of T-DNA integrated into the genomes of wild-type and rat5 Arabidopsis plants indicated that the rat5 mutant is deficient in T-DNA integration. We complemented the rat5 mutation by expressing the RAT5 histone H2A gene in the mutant plant. Overexpression of RAT5 in wild-type plants increased Agrobacterium transformation efficiency. Furthermore, transient expression of a RAT5 gene from the incoming T-DNA was sufficient to complement the rat5 mutant and to increase the transformation efficiency of wild-type Arabidopsis plants.     

Biliary-duodenal fistula following radiofrequency ablation therapy for hepatocellular carcinoma]

Hepatocellular carcinoma (HCC) is one of the most common malignant neoplasms occurring worldwide. Surgical resection currently provides the best chance of long-term tumor free survival, but the most HCCs are not candidates for surgical excision due to poor liver function or poor medical background. Numerous noninvasive alternatives to surgical resection have been introduced to treat liver cancers. Radiofrequency thermal ablation has begun to receive much attention as an effective and minimally invasive technique for the local control of HCC. The biliary system related complications after radiofrequency ablation has rarely been reported. We report a case of biliary-duodenal fistula with liver abscess after radiofrequency ablation for HCC. The case was treated by abscess drainage and antibiotics.     

Biomechanics of cartilage articulation: effects of lubrication and degeneration on shear deformation

OBJECTIVE: To characterize cartilage shear strain during articulation, and the effects of lubrication and degeneration. METHODS: Human osteochondral cores from lateral femoral condyles, characterized as normal or mildly degenerated based on surface structure, were selected. Under video microscopy, pairs of osteochondral blocks from each core were apposed, compressed 15%, and subjected to relative lateral motion with synovial fluid (SF) or phosphate buffered saline (PBS) as lubricant. When cartilage surfaces began to slide steadily, shear strain (Exz) and modulus (G) overall in the full tissue thickness and also as a function of depth from the surface were determined. RESULTS: In normal tissue with SF as lubricant, Exz was highest (0.056) near the articular surface and diminished monotonically with depth, with an overall average Exz of 0.028. In degenerated cartilage with SF as lubricant, Exz near the surface (0.28) was 5-fold that of normal cartilage and localized there, with an overall E(xz) of 0.041. With PBS as lubricant, Exz values near the articular surface were approximately 50% higher than those observed with SF, and overall Exz was 0.045 and 0.062 in normal and degenerated tissue, respectively. Near the articular surface, G was lower with degeneration (0.06 MPa, versus 0.18 MPa in normal cartilage). In both normal and degenerated cartilage, G increased with tissue depth to 3-4 MPa, with an overall G of 0.26-0.32 MPa. CONCLUSION: During articulation, peak cartilage shear is highest near the articular surface and decreases markedly with depth. With degeneration and diminished lubrication, the markedly increased cartilage shear near the articular surface may contribute to progressive cartilage deterioration and osteoarthritis.     

Independent association between glycated hemoglobin and arterial stiffness in healthy men

Aims/Introduction

Many studies have reported that high levels of glycated hemoglobin (HbA1c) are strongly associated with an increased risk of cardiovascular disease. Many researchers have not studied the association of HbA1c with various subclinical atherosclerosis phenotypes. We evaluated the impact of HbA1c on arterial stiffness and atherosclerosis in healthy Korean healthy men.

Materials and Methods

The study population included healthy adult men who participated in health check‐ups. All participants fasted for at least 8 h before taking the blood sample for fasting blood glucose, HbA1c, total cholesterol, high‐density lipoprotein cholesterol, low‐density lipoprotein cholesterol and triglyceride. Arterial stiffness was measured using brachial‐ankle pulse wave velocity. Hierarchical regression analysis allowed us to determine the relationship between brachial‐ankle pulse wave velocity (baPWV) and potential risk factors for cardiovascular disease.

Results

Age and HbA1c were significantly related to baPWV, in model 1. In model 2, blood pressure was added to model 1. Systolic blood pressure was a more significant variable, which was more affected on baPWV than diastolic blood pressure. In the case of model 3, we included all variables regarding arterial stiffness. According to model 3, the most explainable variable was age, and then systolic blood pressure, body mass index and triglyceride, respectively.

Conclusions

We analyzed the associations between HbA1c, which is one of the risk factors, and arterial stiffness independently. An arterial stiffness measurement using baPWV can show the level of severity of the arteriosclerosis. When the level of stiffness has been increased, we could assume that the risk of arteriosclerosis would be increased. It can also be related to the increase of the risk of cardiovascular disease.