The adaptor Grb7 is a novel calmodulin-binding protein: functional implications of the interaction of calmodulin with Grb7

We demonstrate using Ca2+-dependent calmodulin (CaM)-affinity chromatography and overlay with biotinylated CaM that the adaptor proteins growth factor receptor bound (Grb)7 and Grb7V (a naturally occurring variant lacking the Src homology 2 (SH2) domain) are CaM-binding proteins. Deletion of an amphiphilic basic amino-acid sequence (residues 243-256) predicted to form an alpha-helix located in the proximal region of its pleckstrin homology (PH) domain demonstrates the location of the CaM-binding domain. This site is identical in human and rodents Grb7, and shares great homology with similar regions of Grb10 and Grb14, and the Mig10 protein from Caenorhabditis elegans. We show that Grb7 and Grb7V are present in the cytosol and bound to membranes, while the deletion mutants (Grb7Delta and Grb7VDelta) have less capacity to be associated to membranes. Grb7Delta maintains in part the capacity to bind phosphoinositides, and CaM competes for phosphoinositide binding. Activation of ErbB2 by heregulin beta1 decreases the pool of Grb7 associated to membranes. The cell-permeable CaM antagonist W7 (N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide), but not the CaM-dependent protein kinase II inhibitor KN93, prevents this effect. Highly specific cell-permeable CaM inhibitory peptides decrease the association of Grb7 to membranes. This suggests that CaM regulates the intracellular mobilization of Grb7 in living cells. Direct interaction between enhanced yellow fluorescent protein (EYFP)-Grb7 and enhanced cyan fluorescent protein (ECFP)-CaM chimeras at the plasma membrane of living cells was demonstrated by fluorescence resonance energy transfer (FRET). The FRET signal dramatically decreased in cells loaded with a cell-permeable Ca2+ chelator, and was significantly attenuated when enhanced yellow fluorescent protein-Grb7 chimera (EYFP-Grb7)Delta instead of EYFP-Grb7 was used. Finally, we show that conditioned media from cells transiently transfected with Grb7Delta and Grb7VDelta lost its angiogenic activity, in contrast to those from cells transiently transfected with their wild-type counterparts.     

Providers' compliance with the balanced counseling strategy in Guatemala

The balanced counseling strategy developed in Peru improved family planning care and clients' knowledge of their contraceptive method choice, but few providers adopted it. To expand its use, an algorithm was introduced and training, job aids, and reinforcement were supplied to Ministry of Health providers, most of whom were paraprofessionals, from two areas (40 clinics) in Guatemala. Mystery clients made pretest and post-test visits to these clinics and to providers from a nonequivalent control group (40 clinics). The results showed that the strategy was used in 85 percent of the controlled consultations at the experimental clinics. Use of the strategy improved quality of care regardless of the provider's performance at baseline and regardless of ethnic or regional differences. Counseling session length increased by nine minutes, but real-client load did not change. Guatemalan clients can be expected to benefit from the strategy. The increased session length has not yet caused problems, but it may pose policy dilemmas in the future.     

Absence/disappearance of gallbladder in a patient diagnosed with cholelithiasis

We report the case of a patient with preoperatively demonstrated cholecystitis due to cholelithiasis but with no presence of gallbladder at laparoscopy or on conversion to open surgery. Gallbladder ectopy was ruled out and the possibility of gallbladder agenesia was discussed. However, four preoperative ultrasound scans performed within a protocol of treatment with octreotide, in the setting of acute cholecystitis, ruled out this possibility. We discuss the possibility of gallbladder destruction and digestion after a severe inflammatory process, as well as Frey's criteria for gallbladder agenesia. We believe that this is a case of complete gallbladder lysis due to an inflammatory process.     

Non-alcoholic Steatohepatitis (NASH) and Hepatocellular Carcinoma

Non-alcoholic fatty liver disease (NAFLD) is characterized by an excessive accumulation of fatty acids and triglycerides within the cytoplasm of the hepatocytes of non-alcohol users. The natural history varies according to the initial histological diagnosis. A current consideration is that cryptogenic cirrhosis may be representative of a late stage of non-alcoholic steatohepatitis (NASH), which has lost its features of necroinflammatory activity and steatosis in up to 80% of patients. Since NASH is able to progress to cirrhosis, hepatocellular carcinoma (HCC) development may be an end-stage of this disease. We report below two clinical cases of patients diagnosed with NASH who developed HCC. The relationship between NAFLD and HCC is reviewed.     

Prefrontal cognitive functions in stabilized first-episode patients with schizophrenia spectrum disorders: a dissociation between dorsolateral and orbitofrontal functioning

Specific prefrontal cognitive impairments have been reported in first-episode and chronic schizophrenia. We sought to investigate potential impairments in specific prefrontal cortical cognitive functions among stabilized patients with a first-episode of schizophrenia. A sample of 80 individuals with a first-episode of schizophrenia spectrum disorders and 22 healthy volunteers underwent a neurocognitive battery assessing orbitofrontal (OFC) [The Iowa Gambling Task (GT)], and dorsolateral prefrontal (DLPFC) functions (WAIS III Backward digits, verbal fluency test (FAS), and Trail Making Test). Cognitive data were obtained following stabilization of acute psychotic symptoms. Clinical symptoms after six weeks of treatment were assessed by using the SAPS and SANS scales. While there were no significant group differences in overall scores and in the profile of progress of performance along periods on the GT, patient group showed a significant impairment when performing DLPFC tasks. Only FAS score was correlated to the severity of negative symptomatology. The OFC functions are unimpaired at the early phases of psychosis and in contrast there is a significant deficit in DLPFC functions in first-episode of schizophrenia.     

Catechol-O-methyltransferase Val158Met polymorphism and negative symptoms after acute antipsychotic treatment in first-episode non-affective psychosis

Genetic factors play an important role in the understanding of clinical response to antipsychotic treatments. We aimed to assess the effect of the catechol-O-methyltransferase (COMT) genotype in the short-term (6 weeks) clinical response of 161 first-episode psychosis patients. COMT genotype was not related to clinical response at 6 weeks. Val homozygote patients showed higher negative symptoms than Met homozygote patients. The COMT Val158 genotype seems to be related to the severity of negative symptoms rather than to clinical response.