Further research on the role of prostanoids in controlling renal function in humans in normal potassium balance and acute experimental potassium depletion. I: Studies of normal potassium balance. Effects of indomethacin

The renal function was studied by clearance (cl.) method during hypotonic polyuria (oral water load followed by 5% dextrose solution infusion) and successive relative antidiuresis induced by lysine-8-vasopressin (LVP) administration (5 microU in bolo followed by continuous infusion at a rate of 0.04 microU/min). Four 15 min and two 60 min clearance (cl.) periods were performed during hypotonic polyuria and antidiuresis, respectively. Glomerular filtration rate was estimated by creatinine cl.; the osmotic cl. (Cosm, CH2O), the absolute and fractional excretions of water, sodium, potassium and chloride were determined by usual methods. The urinary PGE2, 6-keto-PGF1 alpha and TxB2 concentrations were determined by RIA method. Fourteen healthy women submitted to a normal sodium and potassium daily intake were studied; in 6 of them paired studies in absence and in presence of indomethacin (100 mg, i.m.), respectively, were performed. LVP induced a significant reduction of creatinine cl., urinary flow rate and of prostanoid excretion. In hypotonic polyuria, indomethacin significantly reduced the creatinine cl. and the diuretic response to the water load; moreover the urinary PGE2 and 6-keto-PGF1 alpha excretions were significantly lower (85.6 +/- 1.9% and 37.7 +/- 3.2%) while the reduction of urinary TxB2 excretion was not significant (34.4 +/- 13%). Indomethacin did not affect significantly the LVP renal effects in normal potassium balance.     

Role of prostanoids in the control of renal function in normal potassium balance and in acute experimental potassium depletion. 4. Relation of extrarenal parameters, renal function parameters and urinary excretion of prostanoids

The renal function has been evaluated by clearance (cl.) method during hypotonic polyuria and successive moderate antidiuresis induced by a low dose of lysine-8-vasopressin; four 15 min and two 60 min cl. periods were performed, respectively. Glomerular filtration rate was estimated by creatinine cl.; the osmotic cl. (Cosm, CH2O), the absolute and fractional excretions of water, sodium, potassium and chloride were determined by usual methods. The urinary concentrations of PGE2, 6-keto-PGF1 alpha (6KPGF) and TxB2 were measured by RIA. The study protocol was applied in normal potassium balance and experimental potassium balance (KD), both in absence and presence of indomethacin. In KD groups with a potassium cumulative deficit of 198.4 +/- 22.2 meq (D3; n = 6) during polyuria significant correlations are consistent with the hypothesis that the lower the plasma potassium concentration is the higher the urinary chloride excretion and the inhibition of distal fractional chloride reabsorption. Moreover, by utilizing the polyuria and antidiuresis data pool, the effects of urine flow rate changes on PGE2 and 6KPGF urinary excretions are blunted as compared to normal potassium balance (n = 14). After indomethacin treatment (D3.I) the following functional relationships are disclosed: a) the lower the kaliemia is the lower the urinary chloride and potassium excretions and the higher the fractional isosmotic reabsorption; b) the lower the urinary potassium excretion is the lower the urinary chloride excretion. In both D3 and D3.I experimental groups the positive correlation between urinary chloride excretion and urinary potassium excretion is significant.     

The structure and evolution of the spider monkey delta-globin gene

We have isolated the delta-globin gene of the New-World spider monkey, Ateles geoffroyi, and compared its nucleotide sequence with those of other primate delta- and beta-globin genes. Among primate delta-globin genes, the rate of nonsynonymous substitutions is much less than the rate of synonymous substitutions. This suggests that primate delta- globin genes may remain under evolutionary conservation, perhaps because hemoglobin A2 has an as yet unknown physiological importance.      

Induction of lymphokine-activated killer activity in rat splenocyte cultures: The importance of 2-mercaptoethanol and indomethacin

Summary The role of 2-mercaptoethanol and indomethacin in the induction of lymphokine-activated killer (LAK) activity by interleukin-2 (IL-2) in rat splenocyte cultures was investigated. Spleens from 4-month-old male rats of five different strains were tested. Splenocytes were cultured for 3–5 days in the presence of IL-2 (1000 U/ml) and LAK activity was assessed by 4-h⁵¹Cr release assays with P815 and YAC-1 cells as targets. LAK activity could be induced by IL-2 in splenocytes from all rat strains, but only when 2-mercaptoethanol was present in the culture medium. Optimal LAK activity was induced when the 2-mercaptoethanol concentration in splenocyte cultures was at least 5 µM. Different rat strains showed differences in levels of in vitro induction of LAK activity. In the presence of 2-mercaptoethanol the level of LAK activity induced by IL-2 was high in BN and Lewis rats, intermediate in Wistar and Wag rats, and low in DZB rats. In the absence of 2-mercaptoethanol no or minimal LAK activity was induced. Furthermore we observed that addition of 50 µm indomethacin to the culture medium in the presence of 2-mercaptoethanol augmented the induction of LAK activity to some extent. In the absence of 2-mercaptoethanol, addition of indomethacin resulted only in low levels or no induction of LAK activity. We conclude that for optimal induction of LAK activity by IL-2 in rat splenocyte cultures 2-mercaptoethanol is essential, while indomethacin can only marginally further improve this induction.     

Biochemical pathways in prokaryotes can be traced backward through evolutionary time

For the first time, a credible prokaryotic phylogenetic tree is being assembled by Woese and others using quantitative sequence analysis of oligonucleotides in the highly conservative rRNA. This provides an evolutionary scale against which the evolutionary steps that led to the arrangement and regulation of contemporary biochemical pathways can be measured. This paper presents an emerging evolutionary picture of aromatic amino acid biosynthesis within a large superfamily assemblage of prokaryotes that is sufficiently developed to illustrate a new perspective that will be applicable to many other biochemical pathways.      

Noninvasive papillary transitional-cell tumors. Karyometric and DNA-content analysis

An investigation was undertaken to quantify and correlate the nuclear and DNA-content modifications in the different grades of noninvasive urothelial papillary carcinoma. Nuclear area and a roundness factor were studied by histomorphometry in 35 cases, and the DNA content was analyzed in 27 cases. The values of the nuclear area and the roundness factor increased from normal-looking urothelium to grade 3 tumors. The DNA-content analysis indicated a progressive change of ploidy and the presence of an aneuploid (greater than 4c) population in some of the grade 2 and grade 3 lesions. A high coefficient of correlation between the nuclear abnormalities and the DNA content was found.     

Ultrastructural organization of freeze-fractured interphase nuclei

The morphology of intact or membrane-deprived interphase nuclei has been analysed by freeze-fracture electron microscopy. This method appears particularly useful for providing information on the distribution and organisation of chromatin and ribonucleoproteins in the absence of dehydration and embedding artifacts of conventional electron microscope techniques which, among other effects, appear to affect heterochromatin distribution, inducing its aggregation along the nuclear envelope. The main levels of chromatin superstructure, from nucleosome to solenoid fibres, are detectable in the replicas of freeze-fractured nuclei on the basis of the size of their shadow, a parameter particularly suitable for automated image analyses.     

The evolutionary history of the amylase multigene family in Drosophila pseudoobscura

In Drosophila pseudoobscura, the amylase (Amy) multigene family is contained within a series of inversions, or gene arrangements, on the third chromosome. The Standard (ST), Santa Cruz (SC), and Tree Line (TL) inversions are central to the phylogeny of arrangements, and have clusters of other arrangements derived from them. The gene arrangements belonging to each of these three clusters have a characteristic number of Amy genes, ranging from three in ST to two in SC to one in TL. This distribution pattern can reflect a history of either duplications or deletions, although the data available in the past did not permit a decision between these alternatives. We provide unambiguous evidence that three Amy genes were present before the divergence of the ST, SC, and TL arrangements. Thus, the current status of the Amy multigene family is the result of deletions in the TL and SC arrangements, which created three new pseudogenes: TL Amy2-psi, TL Amy3-psi, and SC Amy3- psi. Analysis of pseudogene sequences revealed that, in the SC and ST arrangements, pseudogene evolution has been retarded, most likely due to the homogenization effect of gene conversion. Finally, by determining the original copy number, we have reconstructed the evolutionary history of the Amy multigene family and linked it with the evolution of the central gene arrangements.      

Immunocytochemical analysis of phosphatidylinositol-specific phospholipase C in PC12 cells: predominance of the δ isoform during neural differentiation

The rat pheochromocytoma PC12 cell line, which differentiates into sympathetic neurons under nerve growth factor (NGF) treatment, contains at least three phosphoinositidase C (PIC) isozymes, PIC , PIC , PIC . These isozymes have been previously shown to display a different subcellular localization. To determine whether or not NGF induces changes in the presence and/or distribution of PIC isozymes during PC12 neural differentiation, studies were carried out by means of in situ immunocytochemistry. After NGF administration the proliferative activity was progressively reduced to very low levels, as measured by bromodeoxyUridine incorporation, and a neuron-like morphology was displayed by almost all cells. In unstimulated PC12 cells, PIC was detected in the nucleus whereas PIC was only cytoplasmic; PIC was found in both cell compartments. In cells treated with NGF for 3 days, neural processes extended to twice the diameter of the cell body; the isoform was concentrated near the nucleus, while the immunoreactivity of the form remained constant and the form was increased. After 10 days of treatment with NGF, PIC was hardly detectable and PIC immunostaining was considerably decreased. On the contrary, PIC progressively increased and, after 14 days of NGF exposure, fully differentiated cells displayed an intense labelling of cell body and neurites. In the same cells, PIC and PIC were almost negative. These results suggest that NGF dependent neural differentiation is related to the selective down regulation of PIC and and the increase of PIC isozyme associated with the decrease of cell proliferation.