The use of resonance energy transfer in high-throughput screening: BRET versus FRET

Bioluminescence resonance energy transfer has developed in recent years as a new technique to study protein-protein interactions. Protein partners of interest are tagged with either luciferase or green fluorescent protein (GFP). Non-radiative energy transfer between the excited luciferase and the GFP permits the study of spatial relationships between the two partners. This technique constitutes an important tool for the study of the functional activity of different types of receptors, and can be used in sensitive, homogenous high-throughput screening assays.     

Interaction of the insulin receptor with the receptor-like protein tyrosine phosphatases PTPalpha and PTPepsilon in living cells

The interactions between the insulin receptor and the two highly homologous receptor-like protein tyrosine phosphatases (PTPase) PTPalpha and PTPepsilon were studied in living cells by using bioluminescence resonance energy transfer. In human embryonic kidney 293 cells expressing the insulin receptor fused to luciferase and substrate-trapping mutants of PTPalpha or PTPepsilon fused to the fluorescent protein Topaz, insulin induces an increase in resonance energy transfer that could be followed in real time in living cells. Insulin effect could be detected at very early time points and was maximal less than 2 min after insulin addition. Bioluminescence resonance energy-transfer saturation experiments indicate that insulin does not stimulate the recruitment of protein tyrosine phosphatase molecules to the insulin receptor but rather induces conformational changes within preassociated insulin receptor/protein tyrosine phosphatase complexes. Physical preassociation of the insulin receptor with these protein tyrosine phosphatases at the plasma membrane, in the absence of insulin, was also demonstrated by chemical cross-linking with a non-cell-permeable agent. These data provide the first evidence that PTPalpha and PTPepsilon associate with the insulin receptor in the basal state and suggest that these protein tyrosine phosphatases may constitute important negative regulators of the insulin receptor tyrosine kinase activity by acting rapidly at the plasma membrane level.     

Do health causal attributions and coping strategies act as moderators of quality of life in peritoneal dialysis patients?

OBJECTIVE: The present study aimed at testing the relationships between health causal attribution and coping mechanisms with quality of life (QOL) in patients who have end-stage renal disease (ESRD) undergoing a peritoneal dialysis (PD) treatment. It was hypothesized that QOL should be negatively associated with the severity of the disease. Problem-focused coping, internal health-related locus of control (HRLOC) and medical power HRLOC were hypothesized as positive moderators preserving a better QOL, after controlling for the severity of the disease. METHODS: A total of 47 PD patients completed the Kidney Disease Quality of Life (KD-QOL) scale encompassing the Medical Outcomes Study Short-Form (MOS SF-36) self-administered questionnaire, the Multidimensional Health Locus of Control scale and the Ways of Coping Check-List (WCCL) scale. RESULTS: Low scores for all QOL scores were found except for pain dimension, as compared with scores available from a general French population. Globally, QOL was not related to the severity of the disease. Univariate analysis showed that the physical component score (PCS) of QOL was positively associated with internal HRLOC (r=.35; P<.05), and negatively with medical power HRLOC (r=-.36; P<.05). Multivariate analysis adjusting for age confirmed these results. Mental component score (MCS) was negatively associated with the use of emotion-focused coping and seeking social support (r=-.45; P=.001 and r=-.30; P<.05, respectively), the first association persisting in multivariate analysis. Neither PCS nor MCS was linked to the use of problem-focused coping. CONCLUSION: These results suggest that physical QOL is all the more preserved when patients are more convinced that their behaviour can influence their health condition and that psychological QOL is all the more impaired when health condition is perceived as less controllable, requiring emotion-focused coping (avoidance strategies). Health causal attributions and coping act respectively as moderators of physical and psychological components of QOL.     

Von Hippel-Lindau disease and obstetric anaesthesia: 3 cases report

Von Hippel-Lindau (VHL) disease is a rare autosomal dominant genetic disorder with retinal and nervous system haemangioblastomas, phaeochromocytomia, kidney, pancreas and endolymphatic tumors. The management of childbirth, including epidural anaesthesia is controversial. The possible presence of vascular malformations (retinal and central nervous system and spinal haemangioblastomas) increases the risk of disrupting central nervous system haemangioblastoma during delivery and when epidural anaesthesia is initiated. We report 3 cases of patient with VHL disease with successful epidural anaesthesia 2 for delivery and 1 for Caesarean section.     

Intravitreal triamcinolone for refractory pseudophakic macular edema

PURPOSE: To evaluate the efficacy of intravitreal triamcinolone in refractory pseudophakic cystoid macular edema. DESIGN: A prospective, interventional case series. METHODS: Three eyes of three patients with longstanding pseudophakic cystoid macular edema following uncomplicated cataract surgery, refractory to any medication, were treated with 8 mg of intravitreal triamcinolone. All three eyes were evaluated before injection and throughout follow-up with the Early Treatment Diabetic Retinopathy Study's visual acuity chart, fluorescein angiography, and macular mapping using optical coherence tomography. RESULTS: A month after intravitreal triamcinolone injection, a dramatic decrease in macular thickness was noted by optical coherence tomography in all three eyes (from a mean of 502-233 microm). Mean improvement in visual acuity was 3.7 Snellen lines. Two to 4 months after triamcinolone injection, however, the edema recurred in all cases, to the same degree as before the injection, combined with a decrease in vision. Two eyes underwent a second injection of triamcinolone, and macular thickness decreased, but the edema again recurred 3 months after injection. CONCLUSION: Intravitreal injection of triamcinolone induces striking regression, within 1 month, of chronic refractory macular edema. This regression appears to be transient, however, even after a second injection.     

Involvement of superior mesenteric vessels and portal vein in pancreatic adenocarcinoma: Detection with CT during arterial portography

A prospective study was designed to determine the utility of computed tomography (CT) during arterial portography (CTAP) in the detection of superior mesenteric vessels and portal vein involvement in patients with pancreatic adenocarcinoma. Eighteen patients with adenocarcinoma of the head of the pancreas and eight patients with benign pancreatic disease were investigated with CTAP, dynamic contrast-enhanced CT, and angiography. Appropriate review was made to determine presence or absence of superior mesenteric vessels and portal vein involvement. Final diagnosis was obtained in all cases by surgical explorations. The overall accuracy rate for detecting or excluding superior mesenteric vessels and portal vein involvement was 96% (25 of 26 patients) with CTAP, 88% (23 of 26 patients) with dynamic contrast-enhanced CT, and 85% (22 of 26 patients) with angiography. No statistically significant difference in accuracy was found among the three techniques. Our results suggest that the use of CTAP is not indicated in the preoperative detection of superior mesenteric vessels and portal vein involvement in patients with pancreatic adenocarcinoma.