Interaction of fluoxetine with the human placental serotonin transporter

The interaction of fluoxetine, a non-tricyclic antidepressant, with the human placental serotonin transporter was investigated by studying its influence on [3H]paroxetine binding to the transporter and on [3H]serotonin uptake via the transporter. These studies were done using brush-border membrane vesicles purified from normal term human placentas. Fluoxetine inhibited binding of paroxetine to the membrane vesicles in a concentration-dependent manner, with a Ki value of 3 nM. Kinetic analysis revealed that the inhibition was competitive because the presence of 10 nM fluoxetine increased the Kd for paroxetine from 72 to 461 pM, but had no effect on the Bmax. Fluoxetine also caused a time-dependent dissociation of paroxetine already bound to the transporter. The dissociation followed first-order kinetics. Uptake of serotonin in these membrane vesicles was also inhibited by fluoxetine. The inhibition was concentration dependent with a Ki value of 66 nM at pH 7.5 and 80 nM at pH 6.5. The effect of fluoxetine on the uptake kinetics was to increase the apparent dissociation constant (Kt) for serotonin without influencing the maximal transport capacity (Vmax). The results demonstrate that fluoxetine is a high-affinity ligand and a potent inhibitor of the serotonin transporter found in the human placental brush-border membrane.     

Comparison of Snellen letter and Vistech grating charts as refraction targets

Snellen letter and Vistech sine wave grating charts were used as refraction targets to determine differences in times required to accomplish subjective refractions, endpoint refractive values, and subject preferences. Thirty hyperopes and 30 myopes, divided into three age groups, were selected as subjects. Two examiners tested each subject using each chart. Both timed their subjective refractions and recorded their findings. After all data were recorded, each subject completed a questionnaire to determine chart preference. There were no clinically significant mean differences between the charts in terms of endpoint refractive data, but, on average, the Snellen chart was faster to use. Chart rating depended on the subject's refractive status with more myopes preferring the Snellen chart and more hyperopes rating the grating chart higher.     

Lung inflation with direct injection of agarose: a technique for simultaneous molecular and morphometric measurements

The usual methods for preparing lungs for morphologic study involve the instillation of fixatives that modifyproteins and RNA such that the tissue is unsuitable for molecular studies. To develop a technique suitable for molecular studies, pieces of adult rat lungs were infiltrated with agarose, glutaraldehyde, or paraformaldehyde and the consistency of alveolar inflation was compared to lungs inflated with 10% formalin. Only direct injection with 1% agarose resulted in comparable inflation of lung tissue and preserved RNA and protein. Thus, this technique enables simultaneous molecular and morphometric analysis of the lung on small pieces of lung tissue in heterogeneous lung diseases.     

Impaired transforming growth factor-beta signaling in idiopathic pulmonary arterial hypertension

Mutations in transforming growth factor-beta family receptor-II, bone morphogenetic protein receptor-2, and activin-like kinase-1 have been associated with pulmonary hypertension. In the present study, we determined that pulmonary arteries in normal lungs and in lungs of patients with emphysema and idiopathic pulmonary arterial hypertension comparably expressed transforming growth factor-beta receptors I and II, Smad(1, 5, 8), Smad2, Smad3, Smad4, phosphorylated Smad(1, 5, 8), and phosphorylated Smad2 (the latter two both indicative of active in vivo signaling) in endothelial cells, as assessed by immunohistochemistry and quantitative morphometry. Medial or intimal smooth muscle cells had weak or absent expression of these molecules. In clear contrast to endothelial cell expression in pulmonary arteries and in endothelial cells lining incipient vessels within plexiform lesions of hypertensive lungs, endothelial cells present in the core of the lesions lacked expression of all examined members of the signaling molecules. These findings were made irrespective of the mutation status of bone morphogenetic protein receptor-2 in hypertensive patients. Our findings suggest that pulmonary artery endothelial cells in both normal and severely hypertensive lungs have active transforming growth factor-beta family signaling, and that loss of signaling might contribute to the abnormal growth of endothelial cells in plexiform lesions in idiopathic pulmonary arterial hypertension.     

Systemic hematologic effects of PEG-rHuMGDF-induced megakaryocyte hyperplasia in mice

PEG-rHuMGDF injected daily in normal mice causes a rapid dose-dependent increase in megakaryocytes and platelets. At the same time that platelet numbers are increased, the mean platelet volume (MPV) and platelet distribution width (PDW) can be either decreased, normal, or increased depending on the dose and time after administration. Thus, PEG-rHuMGDF at a low dose causes decreases in MPV and PDW, MGDF at an intermediate dose causes an initial increase followed by a decrease in MPV and PDW, and PEG-rHuMGDF at higher doses causes an increase in MPV and PDW followed by a gradual normalization of these platelet indices. In addition to the expected thrombocytosis after 7 to 10 days of daily injection of high doses of PEG-rHuMGDF, a transient decrease in peripheral red blood cell numbers and hemoglobin is noted accompanied in the bone marrow by megakaryocytic hyperplasia, myeloid hyperplasia, erythroid and lymphoid hypoplasia, and deposition of a fine network of reticulin fibers. Splenomegaly, an increase in splenic megakaryocytes, and extramedullary hematopoiesis accompany the hematologic changes in the peripheral blood and marrow to complete a spectrum of pathologic features similar to those reported in patients with myelofibrosis and megakaryocyte hyperplasia. However, all the PEG-rHuMGDF-initiated hematopathology including the increase in marrow reticulin is completely and rapidly reversible upon the cessation of administration of PEG-rHuMGDF. Thus, transient hyperplastic proliferation of megakaryocytes does not cause irreversible tissue injury. Furthermore, PEG-rHuMGDF completely ameliorates carboplatin-induced thrombocytopenia at a low-dose that does not cause the hematopathology associated with myelofibrosis.