Synthetic rat V1a vasopressin receptor fragments interfere with vasopressin binding via specific interaction with the receptor

To study the vasopressin receptor domains involved in the hormonal binding, we synthesized natural and modified fragments of V1a vasopressin receptor and tested their abilities to affect hormone-receptor interactions. Natural fragments mimicking the external loops one, two, and three were able to inhibit specific vasopressin binding to V1a receptor. In contrast, the natural N-terminal part of the V1a vasopressin receptor was found inactive. One fragment, derived from the external second loop and containing an additional C-terminal cysteine amide, was able to fully inhibit the specific binding of both labeled vasopressin agonist and antagonist to rat liver V1a vasopressin receptor and the vasopressin-sensitive phospholipase C of WRK1 cells. The peptide-mediated inhibition involved specific interactions between the V1a receptor and synthetic V1a vasopressin receptor fragment since 1) it was dependent upon the vasopressin receptor subtype tested (Ki(app) for the peptide: 3.7, 14.6, and 64.5 microM for displacing [3H]vasopressin from rat V1a, V1b, and V2 receptors, respectively; 2) it was specific and did not affect sarcosin 1-angiotensin II binding to rat liver membranes; 3) it was not mimicked by vasopressin receptor unrelated peptides exhibiting putative detergent properties; and 4) no direct interaction between [3H]vasopressin and synthetic peptide linked to an affinity chromatography column could be observed. Such an inhibition affected both the maximal binding capacity of the V1a vasopressin receptor and its affinity for the labeled hormone, depending upon the dose of synthetic peptide used and was partially irreversible. Structure-activity studies using a serie of synthetic fragments revealed the importance of their size and cysteinyl composition. These data indicate that some peptides mimicking extracellular loops of the V1a vasopressin receptor may interact with the vasopressin receptor itself and modify its coupling with phospholipase C.     

Measurement of synovial lining volume by magnetic resonance imaging of the knee in chronic synovitis

OBJECTIVES: Current methods of monitoring chronic synovitis in a single joint rely on clinical examination derived indices, such as the detection of synovial thickening. This study aimed to develop a reproducible method for quantifying the volume of synovial lining in chronic synovitis using contrast enhanced magnetic resonance (MR) imaging. METHODS: The knees of 18 patients with chronic synovitis were examined (34 studies). A 2D T1 weighted FLASH sequence was used to evaluate the temporal enhancement of synovial structures after intravenous contrast. Synovial lining volume was calculated from subtraction of pre and post-enhancement 3D T1 weighted MP RAGE images with thresholding and pixel counting. Eleven patients were examined before and after intra-articular glucocorticoid (mean interval 14 weeks) and MR data compared with changes in clinical examination derived indices of disease activity. RESULTS: Synovial lining volume varied from 52-267 ml. The coefficient of variation in volume calculation was 3.5% for a single observer and was 3.8% for two observers. Synovial lining volume was quantified in all patients where synovial lining thickening could not be detected clinically. A decrease in synovial lining volume of > 40% was associated with an improvement in synovial lining thickening, detected clinically. Decreases in synovial lining volume were quantified by MR in two of three patients where changes in clinical examination derived indices were inconsistent. CONCLUSIONS: A reproducible method of estimating the volume of synovial lining in patients with chronic synovitis has been developed. MR measurement of synovial lining volume may quantify changes in chronic synovitis that remain unidentified by clinical measures.     

Maternal immunization to Gov system alloantigens on human platelets

BACKGROUND: Immunization to platelet alloantigens can occur during pregnancy or after the transfusion of blood components. Platelet alloantibodies can cause neonatal alloimmune thrombocytopenia and posttransfusion purpura. Transfusion-induced alloimmunization to a novel platelet alloantigen system, Gov, expressed on the 175-kDa glycosyl phosphatidylinositol-anchored platelet glycoprotein, CD109, was previously described. This report describes three unrelated patients who were alloimmunized to Gov(a) or Gov(b) during pregnancy. STUDY DESIGN AND METHODS: Platelets were typed by using radioimmunoprecipitation for HPA-1a, -3a, -5a, -5b, Gov(a), and Gov(b) and by polymerase chain reaction-restriction fragment length polymorphism for HPA-1a, -1b, -3a, and -3b. Maternal sera were screened for platelet antibodies by using radioimmunoprecipitation and the antigen capture assay. RESULTS: Patients 1 and 2 were investigated after the diagnosis of neonatal alloimmune thrombocytopenia in their children, and alloantibodies specific for Gov(b) and Gov(a), respectively, were detected in maternal serum. Serum from patient 3, who had mild idiopathic thrombocytopenia purpura with no detectable autoantibody, was found to contain alloantibodies to Gov(b) and to HPA-5b, presumably as a result of immunization during pregnancy. Platelet typings confirmed that the patients were at risk for alloimmunization to the respective antigen. CONCLUSION: This report of three cases of maternal alloimmunization to antigens in the Gov system indicates that immunization can occur via placental transfer of antigen and that Gov system alloantibodies may be associated with neonatal alloimmune thrombocytopenia.     

Diagnosis of transfusion-associated graft-versus-host disease by polymerase chain reaction in fludarabine-treated B-chronic lymphocytic leukaemia

Transfusion-associated graft-versus-host disease (TA-GVHD), has rarely been reported associated with B-chronic lymphocytic leukaemia (B-CLL). We report a patient diagnosed with B-CLL, previously treated with fludarabine, who developed TA-GVHD after being transfused during surgery for splenectomy. Diagnosis was confirmed by polymerase chain reaction (PCR) detection of donor DNA in the patient, by amplification of Y-chromosome sequence and analysis of minisatellite polymorphisms. B-CLL patients treated with fludarabine appear to be at risk for TA-GVHD and should be regarded as candidates for transfusions with irradiated blood products. This case illustrates that PCR is a rapid technique for the early diagnosis of TA-GVHD.