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.     

The effect of eplir on an experimental toxic lesion of the liver

A hepatoprotector of phospholipid nature eplir extracted from silt mud was shown to prevent like essentiale in CCl4-hepatitis in rats the development of the liver parenchyma necroses, to promote the maintenance of the normal activity of enzymes in hepatocytes, to stabilize lysosomes, to stimulate the antitoxic and excretory functions of the liver, to reduce hyperfermentemia. The mechanism of action of eplir is determined by the antioxidant properties, a decrease of lysophosphatide formation and the inclusion of phosphatidylcholine in membranes.     

The normally expressed κ immunoglobulin light chain gene repertoire and somatic mutations studied by single-sided specific polymerase chain reaction (PCR); frequent occurrence of features often assigned to autoimmunity

The expressed human κ light chain gene repertoire utilized by healthy individuals was studied by two different single-sided specific PCR techniques to avoid bias for certain V genes. A total of 103 rearranged κ sequences from peripheral blood mononuclear cells from healthy individuals were cloned from cDNA and assigned to the Vκ and Jκ germ-line genes with the closest overall homology. The use of cDNA rather than genomic DNA focused the analysis on activated B cells rich in mRNA. Accordingly, the sequences represented the applied repertoire and almost all were somatically mutated. V genes from the Jκ-proximal duplication unit of the κ locus were almost exclusively used. A total of 65% of the sequences could be assigned to four or five genes: A27 (humkv325), L6 (Vg), L2 (humkv328), and A3 and/or A19. N additions and P nucleotides were quite common and found in 32% and 21% of the sequences, respectively. Extended CDR3s more than nine residues in length were found in 18% of the sequences, and in 71% of cases this was due to insertion of an extra proline residue. This proline was usually explained from the germ-line sequences involved. These results are in good agreement with those of previous repertoire studies using potentially V-gene-biased techniques. Thus, it is clear that restricted V-gene usage, common N and P additions, and extended CDR3 regions are normal features and not, as has been claimed, characteristics of pathological autoantibodies.     

cDNA Sequencing of the 5′ Noncoding Region (5′ NCR) to Determine Hepatitis C Genotypes in Patients with Chronic Hepatitis C

Reports suggest that response tointerferon-alpha therapy is influenced by both hepatitisC viral genotype and titer. Our aim was to determine ifdirect, automated, cycle sequencing of the PCR productfrom an HCV RNA detection assay could be used toreliably determine HCV genotype. In addition, theapproach was used to determine the HCV genotypedistribution in our patient population and to learn ifthere was a correlation between HCV genotype and RNAtiter that could be used to predict response totreatment. In all 143 consecutive patients were testedfor both HCV RNA titer and genotype. Automated, cycle sequencing of PCR product was highly effectiveand failed to yield a genotype in only 3 (2%) patients.The distribution of HCV genotypes was: 1a (40%), 1b(39%), 2a (2%), 2b (6%), 3a (4%). There were significant differences in the median HCV RNA titersbetween genotypes 1, 2, and 3. 6 High HCV RNA titers>4.4 × 10⁶ copies/ml were only seenin genotype 1. However, the HCV RNA level should not beused as a surrogate marker of genotype because of a significantoverlap of titers within the genotypes.     

Hematopoietic turnover index in reactive and neoplastic bone marrow lesions: quantification by apoptosis and PCNA labeling

 In order to determine the dynamics of hematopoietic cell turnover, proliferative activity and incidence of apoptosis (programmed cell death) were evaluated in bone marrow trephine biopsies. Selection of patients (20 in each group) included in addition to a control group, idiopathic thrombocytopenia (ITP), reactive thrombocytosis (TH), secondary polycythemia-smokers' polyglobuly (PG), primary (essential-hemorrhagic) thrombocythemia (PTH), polycythemia vera (PV), and finally acute myeloid leukemia (AML). Apoptosis was demonstrated by the in situ end-labeling technique (ISEL) and proliferative activity by applying the monoclonal antibody PC10 raised against proliferating cell nuclear antigen (PCNA). To assess dynamic features of hematopoiesis, an index was calculated consisting of the ratio between PCNA-positive nuclei and the apoptotic cell fraction. This factor was termed the hematopoietic turnover index (HTI). Morphometric analysis revealed that the HTI was significantly increased in AML and PV. According to cell culture studies both disorders are characterized by either a prevalent proliferation of the myeloid or erythroid cell mass. On the other hand, PG, PTH, and TH showed no relevant enhancement of this index in comparison to the control specimen. In vitro experiment results are in keeping with the finding that PG and PTH are not associated with a significant expansion of the erythroid lineage (CFU-E). Similar to ITP and TH, in PTH megakaryocyte proliferation (CFU-MEG) is the predominant feature of cell turnover. Differences between PTH and TH are in line with the reduced in vitro formation of CFU-MEG in the latter disorder. In conclusion, our in situ study on turnover rates of the bone marrow in various neoplastic and reactive lesions extends previous experimental data on hematopoietic cell kinetics. Received: 10 March 1997 / Accepted: 18 May 1997     

Emergence and scattering of multiple neurofibromatosis (NF1)-related sequences during hominoid evolution suggest a process of pericentromeric interchromosomal transposition

Type 1 neurofibromatosis (NF1) gene encodes for a member of the GTPase activating protein family and is considered to be a tumor suppressor gene. Its very high rate of de novo mutation in humans led us to study a specific feature of this gene: the presence of numerous NF1-related sequences. According to our results, the human genome contains at least 11 NF1-related sequences, nine of which are scattered near centromeric sequences of seven different chromosomes. These NF1-related sequences, whose extent is quite varied according to loci, are unprocessed copies of the NF1 gene, and bear numerous mutations. A phylogenetic analysis of the six largest sequences indicates that they are all derived from a common ancestor, which would have appeared 22-33 million years ago, and was subsequently duplicated several times during hominoid evolution. The most recent duplication and interchromosomal transposition occurred in the last million years suggesting that the process could still be ongoing. Intriguing similarities between the evolution of alpha- satellite DNA and NF1-related sequences suggest the involvement of a common genetic mechanism for the generation and pericentric spreading of these NF1 partial copies.