Effects of norepinephrine or prostaglandin E2 on extracellular acidification rate of MCG 101, or K1735-M2 tumor cells

We studied by microphysiometry functional effects of two different signalling molecules in the murine tumor cell lines, MCG 101 and K1735-M2, namely norepinephrine (NE) and prostaglandin E2 (PGE2). This methodology implies estimation of intracellular metabolism by measurements of extracellular acidification rate (ECAR). MCG 101 (an undifferentiated, epithelial-like tumor), in contrast to K1735-M2 (a melanoma), has been found to produce great amounts of PGE2. Challenge of MCG 101 cells with PGE2 (0.284 and 2.84 microM for 9 min) elicited an increase in ECAR by about 10 and 41% above basal level, respectively. Pretreatment with indomethacin (0.5 microM) reduced the response to the two PGE2 concentrations by about 70 and 25%, respectively. In contrast, PGE2 caused virtually no response in K1735-M2 cells. Moreover, NE caused increases in ECAR in both cell types, possibly via beta3-adrenoceptors, as investigated pharmacologically in MCG 101, and by immunocytochemistry in both cell lines. The results obtained strongly suggest functional receptors for PGE2 in MCG 101, but not K1735-M2 tumor cells. Functional receptors for NE were demonstrated in both cell lines. There is possibly an autocrine loop in the MCG 101 cells, in which PGE2 activates cyclooxygenase.     

Influence of ferric chloride treated Leucaena leucocephala on metabolism of mimosine and 3-hydroxy 4(1H)-pyridone in growing rabbits

Twenty-five 42-days old New Zealand white rabbits were weaned and accustomed to a control ration in the 1st week and randomly allotted to five groups of five rabbits each. They were offered the control ration (G-1), and in other groups a portion of the control ration was replaced by Leucaena leaf meal (LLM) treated with 1.2% FeCl₃ or untreated i.e. 25% LLM (G-2), 50% LLM (G-3), 25% treated LLM (G-4), and 50% treated LLM (G-5) ration in pelleted form in a 8 weeks feeding cum metabolism trial. Average intake of mimosine and 3,4 DHP (dihydroxypyridone) was 304.6 and 129.5; 680.2 and 212.3; 279.6 and 147.6; and 643.1 and 239.9 mg day⁻¹ in G-2–G-5, respectively. Mimosine and 2,3 DHP were not detected in faeces. The faecal excretion of 3,4 DHP (as % intake of mimosine plus 3,4 DHP) in the rabbits of groups G-4 (43.5) and G-5 (40.6) was significantly (P<0.05) higher due to FeCl₃ treatment as compared to excretion in groups G-2 (30.1) and G-3 (21.4) fed untreated LLM. GOT (Glutamic oxalacetic transaminase), GPT (Glutamic-pyruvic transaminase), T₃ (tri-iodothyronine) and T₄ (thyroxine) levels in blood were within normal physiological range. Mimosine 3,4 DHP and 2,3 DHP, all were excreted through urine. The urinary excretion of 3,4 DHP was significantly lower (P<0.05) in G-4 and G-5. The overall excretion of DHP (2,3 and 3,4 DHP) was similar in all the groups. Severe hepatic and kidney damage occured in G-2 and G-3, while, in G-4 and G-5 very mild or no damage to liver and kidney was recorded. All tissues were devoid of mimosine, but DHP was present in liver, kidney and lungs. The maximum DHP in liver indicated as the primary site of DHP metabolism. In vitro incubation of LLM with caecal contents revealed 72.68–100% microbial degradation of mimosine. The overall DHP degradation ranged from 7.10% to 37.81% being the highest in G-3. The results indicated that, FeCl₃ treated leucaena could be used in commercial meat rabbit rations.     

Purification of N-terminal hexapeptide of big gastrin from human urine

We previously demonstrated that extremely high amounts of N-terminal big gastrin (G-34) fragments are excreted in human urine and three of them are N-terminal octa-, nona-, and decapeptide of G-34. Our subsequent examination revealed that there exists a considerable amount of another N-terminal G-34 fragment in urine, less hydrophobic than the three peptides. We purified this fragment from urine of an achlorhydric patient and determined the structure: less than Glu-Leu-Gly-Pro-Gln-Gly. The purification was carried out by Sep-Pak C18 cartridges, Sephadex G-25, and reverse phase HPLC. The structure was determined by a combination of amino acid analysis, amino acid sequence analysis, and mass spectral analysis. N-terminal hexapeptide of G-34 is the second richest component of urinary N-terminal G-34 fragments next to N-terminal octapeptide of G-34 in normal subjects.     

Tumour Necrosis Factor-alpha Production and Immune Cell Activation in Tuberculous Meningitis

The local production of tumour necrosis factor-alpha (TNFalpha) was evaluated in the cerebrospinal fluid (CSF) from ten patients with tuberculous meningitis (TBM). The degree of intrathecal immune activation was also studied by assessing the CSF levels of beta(2)-microglobulin (beta(2)-M) and adenosine deaminase activity (ADA). Results indicate that elevated CSF concentrations of TNFalpha, beta(2)-M and ADA were found in all TBM patients. Moreover, TNFalpha is produced and selectively concentrated for a long period of time, while beta(2)-M and ADA values progressively decline during the course of TBM. Our findings suggest that in TBM patients, after an early activation of immune cells, there is an enhanced and continuous production of TNFalpha at the site of infection.     

Isolation and characterization of gangliosides from Trypanosoma brucei

Gangliosides were isolated from Trypanosoma brucei and analyzed by thin-layer chromatography (TLC) and TLC immunostaining test. Four species of gangliosides, designated as G-1, G-2, G-3, and G-4, were separated by TLC. G-1 ganglioside had the same TLC migration rate as GM3. In contrast, G-2, G-3, and G-4 gangliosides migrated a little slower than GM1, GD1a, and GD1b, respectively. To characterize the molecular species of gangliosides from T. brucei, G-1, G-2, G-3, and G-4 gangliosides were purified and analyzed by TLC immunostaining test with monoclonal antibodies against gangliosides. G-1 ganglioside showed the reactivity to the monoclonal antibody against ganglioside GM3. G-2 was recognized by the anti-GM1 monoclonal antibody. G-3 showed reaction with the monoclonal antibody to GD1a. G-4 had the reactivity to anti-GD1b monoclonal antibody. Using 4 kinds of monoclonal antibodies, we also studied the expression of GM3, GM1, GD1a, and GD1b in T. brucei parasites. GM3, GM1, GD1a, and GD1b were detected on the cell surface of T. brucei. These results suggest that G-1, G-2, G-3, and G-4 gangliosides are GM3 (NeuAc alpha2-3Gal beta1-4Glc beta1-1Cer), GM1 (Gal beta1-3GalNAc beta1-4[NeuAc alpha2-3]Gal beta1-4Glc beta1-1Cer), GD1a (NeuAc alpha2-3Gal beta1-3GalNAc beta1-4[NeuAc alpha2-3]Gal beta1-4Glc beta1-1Cer), and GD1b (Gal beta1-3GalNAc beta1-4[NeuAc alpha2-8NeuAc alpha2-3]Gal beta1-4Glc beta1-1Cer), respectively, and also that they are expressed on the cell surface of T. brucei.     

Chromosomal location of a <Emphasis Type="Italic">Triticum dicoccoides</Emphasis>-derived powdery mildew resistance gene in common wheat by using microsatellite markers

The powdery mildew resistance has been transferred from an Israeli wild emmer (Triticum dicoccoides) accession "G-305-M" into common wheat by crossing and backcrossing (G-305-M/781//Jing 411*3). Genetic analysis showed that the resistance was controlled by a single dominant gene at the seedling stage. Among the 102 pairs of SSR primers tested, four polymorphic microsatellite markers (Xpsp3029, Xpsp3071, Xpsp3152 and Xgwm570) from the long arm of chromosome 6A were mapped in a BC(2)F(3) population segregating for powdery mildew resistance and consisting of 167 plants. The genetic distances between the resistance gene and these four markers were: 0.6 cM to Xpsp3029, 3.1 cM to Xpsp3071, 11.2 cM to Xpsp3152 and 20.4 cM to Xgwm570, respectively. The order of these microsatellite loci agreed well with the established microsatellite map of chromosome arm 6AL. We concluded that the resistance gene was located on the long arm of chromosome 6AL. Based on the origin and chromosomal location of the gene, it is suggested that the resistance gene derived from "G-305-M" is a novel powdery mildew resistance gene and is temporarily designated MlG.     

A new mannoheptaose containing alpha and beta-(1-->2) linkages isolated from the mannan of Torulaspora delbrueckii: ELISA inhibition studies

Torulaspora delbrueckii starin IFO 0955 was examined with respect to its structural and serological properties of the cell wall mannan (Td-0955-M). Td-0955-M revealed significant reactivities with sera from a commercially available factor serum kit (Candida Check) in ELISA. Td-0955-M was investigated for its chemical structure by acetolysis under conventional and mild conditions. NMR and GC techniques were used as analytical techniques. The mannooligosaccharide fractions eluted from a Bio-Gel P-2 column were found to consist of Man(alpha1-2)Man, M2, Man(alpha1-2)Man(alpha1-2)Man and Man(beta1-2)Man(alpha1-2)Man, M3, Man(alpha1-2)Man(beta1-2)Man(beta1-2)Man(alpha1-2)Man, M5, and a new mannoheptaose, which possesses the structure, Man(alpha1-2)Man(beta1-2)Man(beta1-2)Man(beta1-2)Man(beta1-2)Man(alpha1-2)Man, M7. The results of the inhibition ELISA showed that the M7 oligosaccharide significantly inhibited the reactivities in the Td-0955-M-factor serum systems.     

Thromboxane (TX)A3 and prostaglandin (PG)I3 are formed in man after dietary eicosapentaenoic acid: identification and quantification by capillary gas chromatography-electron impact mass spectrometry

The low incidence of myocardial infarction in Greenland Eskimos has been related to their traditional marine diet rich in eicosapentaenoic acid. However, whether dietary eicosapentaenoic acid is indeed transformed in man to antiaggregatory PGI3 and weakly proaggregatory TXA3 has not been clarified. In our studies we ingested either cod liver oil or mackerel both rich in eicosapentaenoic acid. Formation of TXB3, the hydrolysis product of TXA3, in platelet-rich plasma stimulated ex vivo with collagen was traced by capillary GC/EIMS. Via external standard, TXB3 formation in platelets was estimated to be 5-15% of TXB2 formation. From urine we extracted dinor metabolites of PGI according to a selective method. We utilized delta 17-2,3-dinor-6-keto-PGF1 alpha (PGI3-M) as an index of total body production of PGI3 in analogy to 2,3-dinor-6-keto-PGF1 alpha (PGI2-M), the major urinary metabolite of PGI2. We separated PGI2-M and PGI3-M as the Me, MO, Me3Si derivatives by capillary gas chromatography and identified PGI3-M by EI mass spectrometry. Excretion of PGI3-M, which was not detectable under control conditions, was 83 +/- 25 ng/24 h (SD) after ingestion of cod liver oil and 134 +/- 38 ng/24 h after mackerel ingestion, while excretion of PGI2-M was 162 +/- 52 ng/24 h and 236 +/- 32 ng/24 h, respectively. Our findings with diets rich in EPA show that it is possible in man to change in vivo the spectrum of biologically active prostanoids by nutritional means and alter it in a favourable direction.     

Enzymatic detection of urinary conjugated steroids after gel chromatography

An enzymatic detection method is described for urinary conjugated steroids after chromatographic fractionation with Sephadex G-25. The principle of the method is as follows. Part of a 24-h urine sample, (1–2 ml of urine) is applied directly, to a short column of Sephadex G-25 and eluted with acetate buffer solution. Steroid conjugates in each fraction are hydrolyzed with steroid sulfatase—β-glucuronidase. After enzymatic hydrolysis, an enzymatic color development reagent for steroids, either 3α-hydroxysteroid dehydrogenase or 3β-hydroxysteroid oxidase, are added and the dye formed is measured spectrophotometrically. Excretion patterns of steroid-3β-sulfates, and steroid-3α-glucoronides and steroid-3α-sulfates are shown with some patients' samples. A precision of the assay values for steroid-3α-glucuronide, steroid-3α-sulfate and steroid-3β-sulfates in urine samples and assay values for normal subjects are also studied.This simple enzymatic method for detecting the excreption patterns of urinary conjugated steroids may have a diagnostic value for clinical tests.     

The in vitro interactions of rat pancreatic elastase and normal and inflammatory ray serum.

The partition of labelled rat pancreatic elastase (EC 3.4.21.11) between the different protease inhibitors of rat plasma was studied at different levels of saturation of the inhibitors of rat plasma was studied at different levels of saturation of the inhibitor capacity of plasma with the enzyme. The reaction mixtures were analysed by immunoelectrophoretic methods utilizing specific antisera against the different inhibitors and by gel filtration on Sephadex G-200. Rat serum was shown to contain four elastase binding proteins. alpha 1-antitrypsin, alpha 1-macroglobulin and alpha 2-acute phase protein and alpha 1-inhibitor 3 which exhibits immunologic cross-reaction with human inter-alpha-trypsin inhibitor and is of similar molecular weight. With minute amounts of labelled elastase the partition among the binding protein was alpha 1-macroglobulin 60%, alpha 1-antitrypsin 24% and alpha 1-I3 16%. The 60% value of alpha 1-M bound radioactivity in normal serum corresponds to the sum of alpha 1-M and alpha 2-AP labelling in inflammatory serum.     

Critical evaluation of α1- and β2-microglobulins in urine as markers of cadmium-induced tubular dysfunction

The purpose of the study was to examine the validity of alpha1-microglobulin (alpha1-MG) in comparison with popularly used beta2-microglobulin (beta2-MG). A database was revisited to select ca. 7,500 spot urine samples (of adequate urine density) from non-pregnant, non-lactating and never-smoking adult women. The validity of the MGs was examined in terms of stability of the MG-uria prevalence in urine samples of various creatinine (CR or cr) concentration or specific gravity (SG or sg). Comparisons were made for MGs as observed (e.g., alpha1-MGob), as corrected for CR (e.g., alpha1-MGcr) and as corrected for SG of 1.016 (e.g., alpha1-MGsg). A cut-off value of 5.7 mg/g cr (or mg/l) for alpha1-MG was deduced from a cut-off value of 400 microg/g cr (or mcirog/l) for beta2-MG, because the correlation between alpha1-MGcr and beta2-MGcr was statistically significant. The prevalence of a 1-MGsg-uria was essentially unchanged (i.e., from a low of 13.6% to a high of 17.0%, or 1.2 times) except for in very dense or very thin urine samples, in contrast, beta2-MGcr-uria showed a substantial increase (from 0.0% to 2.8% with an infinite rate) as a reverse function of a decrease in CR in urine. The prevalence of uncorrected markers, i.e., alpha1-MGob-uria and beta2-MGob-uria, showed even greater CR- or SG-dependent changes. Thus, it appeared prudent to consider a alpha-MGsg rather than beta2-MGcr as a marker of tubular dysfunction among a general population with various urine density.     

In vivo metabolism of 2,2'-diaminopimelic acid from gram-positive and gram-negative bacterial cells by ruminal microorganisms and ruminants and its use as a marker of bacterial biomass.

Cells of Bacillus megaterium GW1 and Escherichia coli W7-M5 were specifically radiolabeled with 2,2'-diamino[G-3H]pimelic acid ([3H]DAP) as models of gram-positive and gram-negative bacteria, respectively. Two experiments were conducted to study the in vivo metabolism of 2,2'-diaminopimelic acid (DAP) in sheep. In experiment 1, cells of [3H]DAP-labeled B. megaterium GW1 were infused into the rumen of one sheep and the radiolabel was traced within microbial samples, digesta, and the whole animal. Bacterially bound [3H]DAP was extensively metabolized, primarily (up to 70% after 8 h) via decarboxylation to [3H]lysine by both ruminal protozoa and ruminal bacteria. Recovery of infused radiolabel in urine and feces was low (42% after 96 h) and perhaps indicative of further metabolism by the host animal. In experiment 2, [3H]DAP-labeled B. megaterium GW1 was infused into the rumens of three sheep and [3H]DAP-labeled E. coli W7-M5 was infused into the rumen of another sheep. The radioactivity contents of these mutant bacteria were insufficient to use as tracers, but the metabolism of DAP was monitored in the total, free, and peptidyl forms. Free DAP, as a proportion of total DAP in duodenal digesta, varied from 0 to 9.5%, whereas peptidyl DAP accounted for 8.3 to 99.2%. These data reflect the extensive metabolism of bacterially bound DAP within the gastrointestinal tracts of ruminant animals and serve as a serious caution to the uncritical use of DAP as a marker of bacterial biomass in the digesta of these animals.     

NH2-terminal big gastrin immunoreactivity in human urine

The concentrations and molecular forms of urinary and plasma gastrin from normal subjects were studied by radioimmunoassays using two region-specific antisera. Urinary concentration of NH2-terminal big gastrin (G-34) immunoreactivity was several hundred times as great as that of COOH-terminal gastrin immunoreactivity. Fractionation of urine extract showed a broad giant peak of NH2-terminal G-34 immunoreactivity (gastrin fragments "U") eluting in a later position than G-34(1-17) by Sephadex G-50 column chromatography. HPLC revealed that urinary NH2-terminal G-34 immunoreactivity was composed of four fragments including G-34(1-8), G-34(1-9), and G-34(1-10). Sephadex G-50 column chromatography of plasma extract revealed two or three peaks of NH2-terminal G-34 immunoreactivity, and a major peak eluted in the same position as urinary gastrin fragments "U". These results and data on renal clearances suggest that most of all gastrin fragments "U" in plasma are excreted in urine without renal reabsorption, whereas almost all of plasma COOH-terminal gastrin peptides including G-34 and little gastrin (G-17) are removed and metabolized in the kidney.     

Metabolism of oestradiol-17 beta and progesterone in the white rhinoceros (Ceratotherium simum simum)

14C-Labelled oestradiol-17 beta and progesterone (50 mu Ci each) were injected i.v. into an adult female white rhinoceros and all urine and faeces collected separately over the next 4 days. The total recovery of injected label was 61%, 25% being present in the urine and 36% in the faeces. Of the radioactivity recovered, 69% was excreted on Day 2 of the collection period. Repeated extraction of samples obtained on Day 2 showed that, of the radioactivity in faeces, 92.4% was associated with unconjugated steroids whereas in the urine the proportion of conjugated and unconjugated steroids were similar (41.2% and 51.4% respectively). After phenolic separation of urinary steroids, HPLC followed by derivatization and recrystallization techniques identified progesterone as the major component of the unconjugated portion with 4-pregnen-20 alpha-ol-3-one as the principal metabolite in the conjugated fraction. Pregnanediol was not present. Oestrone appeared to be the most abundant oestrogen metabolite with smaller but significant amounts of oestradiol-17 beta and oestradiol-17 alpha in the unconjugated and conjugated fractions respectively. Small amounts of progesterone were found in the faecal extract in which the radioactivity consisted mainly of oestradiol-17 alpha and oestradiol-17 beta. The results have established the major excreted metabolites of oestradiol-17 beta and progesterone in the white rhinoceros and the development of more appropriate assay methods for monitoring ovarian function in African rhinoceroses should now be possible.     

Characterization of human brain cDNA encoding the general isoform of beta-spectrin.

The complete primary structure of the general form of human beta-spectrin (beta G) has been deduced from cDNAs isolated from human brain. beta G-Spectrin is encoded by a gene located on human chromosome 2. beta G-Spectrin and erythrocyte beta-spectrin (beta R) share identical domain organization, with sequence identity of 60% and sequence similarity of 77%. beta-Spectrins have closely related N-terminal domains implicated in binding to actin, and 17 copies of a 106-residue repeat motif with consensus residues that are highly conserved between beta-spectrins as well as alpha-spectrins. C-terminal domains of beta G and the 270-kDa beta R-spectrins are candidate regions to associate with alpha-spectrin, and exhibit 75% similarity. beta G- and beta R-spectrins exhibit different patterns of expression in tissues and follow different developmental programs in those tissues where they are co-expressed. beta G-Spectrin is present in all tissues examined except for erythrocytes, while beta R-spectrin could be detected only in erythrocytes, brain, and heart. beta G- and beta R-Spectrins are both expressed in brain, but beta R appeared later in post-natal development and was highly enriched in cerebellum in contrast to the broad regional distribution of beta G-spectrin. beta-Spectrins are likely to perform related but distinct functions, with beta G in a general, constitutive role and beta R-spectrin involved in more specialized activities of differentiated cells.     

Asymmetric contribution of alpha and beta subunits to the activation of alphabeta heteromeric glycine receptors

This study investigated the role of beta subunits in the activation of alphabeta heteromeric glycine receptor (GlyR) chloride channels recombinantly expressed in HEK293 cells. The approach involved incorporating mutations into corresponding positions in alpha and beta subunits and comparing their effects on receptor function. Although cysteine-substitution mutations to residues in the N-terminal half of the alpha subunit M2-M3 loop dramatically impaired the gating efficacy, the same mutations exerted little effect when incorporated into corresponding positions of the beta subunit. Furthermore, although the alpha subunit M2-M3 loop cysteines were modified by a cysteine-specific reagent, the corresponding beta subunit cysteines showed no evidence of reactivity. These observations suggest structural or functional differences between alpha and beta subunit M2-M3 loops. In addition, a threonine-->leucine mutation at the 9' position in the beta subunit M2 pore-lining domain dramatically increased the glycine sensitivity. By analogy with the effects of the same mutation in other ligand-gated ion channels, it was concluded that the mutation affected the GlyR activation mechanism. This supports the idea that the GlyR beta subunit is involved in receptor gating. In conclusion, this study demonstrates that beta subunits contribute to the activation of the GlyR, but that their involvement in this process is significantly different to that of the alpha subunit.     

A nicked beta-subunit of human chorionic gonadotropin purified from pregnancy urine.

Human chorionic gonadotropin (hCG) is a glycoprotein hormone composed of two dissimilar subunits (alpha and beta) and normally excreted in urine of pregnant women. An uncommon beta-subunit of hCG was purified from fresh early normal pregnancy urine by Sepralyte C8, resin adsorption. Sephadex G-100 column chromatography, and reverse-phase HPLC. SDS-PAGE under non-reducing conditions showed that the apparent molecular weight (39,000) of this beta-subunit was extremely similar to that of the native beta-subunit, which is known to consist of 145 amino acid residues and carbohydrates. However, SDS-PAGE, under reducing conditions, resulted in two bands with apparent molecular weights of 22,000 and 18,000, indicating that it consisted of two peptide fragments connected with disulfide bridge(s). These two peptide fragments, separated and purified from the reduced and carboxymethylated protein, were subjected to amino acid and N-terminal sequence analyses. It was found that this beta-subunit consisted of two polypeptide chains composed of residues 1-47 disulfide-bridged to residues 48-145 of the beta-subunit, which may be produced by nicking of the beta-subunit at the one site (Gly47-Val48). This beta-subunit was termed a nicked beta-subunit of hCG (N-hCG beta). It was also found that N-hCG beta was present in urine as an alpha beta dimer, indicating that an intrachain nicking of this site in the beta-subunit does not inhibit alpha beta dimer formation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Metabolism of [14C]glucose by regenerating spheroplasts of Candida albicans

Spheroplasts of Candida albicans were regenerated in [14C]glucose and buffered magnesium sulphate (0.1 M-Tris/HCl; 0.5 M-MgSO4, pH 7.2) at 35 degrees C. Uptake of glucose by spheroplasts was faster than that by intact yeast cells. After 6 h, 65% of the glucose taken up by the yeast appeared as CO2 and 30% was incorporated into the cellular material. With spheroplasts, 55% of the glucose taken up was expired as CO2, 25% was excreted into the medium as other metabolites and 20% was incorporated into the cells. The regenerating spheroplasts excreted 14C-labelled carbohydrates into the medium which were fractionated on a Sephadex G-15 column. Acid hydrolysis of the low molecular-weight fraction yielded the following sugars: mannose (75.7%), fucose (3.8%), arabinose (3%), galactose (2.1%) and an unidentified monosaccharide (14%). Spheroplasts did not incorporate mannoprotein into the regenerated wall. The wall carbohydrate from regenerated spheroplasts was fractionated on the basis of solubility in sodium hydroxide. The alkali-insoluble fraction was analysed by sequential enzyme hydrolysis; 40% of the incorporated counts were associated with beta (1----3)-linked glucan and 50% with a mixed glucan comprising beta (1----3)- and beta (1----6)-linkages and chitin.     

Comparison of the urinary metabolites of prostacyclin and thromboxane of the 2- and 3-series in a Japanese fishing and a Japanese farming village

We measured PGI2-, PGI3-, and TXA2/3-M (the main urinary metabolites of prostacyclin and thromboxane of the two and three series) in 24-hour urine in a fishing village (21 participants) and a farming village (19 participants) in Japan, expecting to find more PGI3-M in the fishing village than in the farming village. The food consumption for three consecutive days prior to a blood collection was recorded and analyzed for eicosapentaenoic acid (EPA) content. Urine was collected for 24 hours prior to the blood sampling. When our studies were performed (April, 1985), catches of fish were the lowest on record around the fishing village, and the consumption of EPA in the fishing village was less than half of that in the farming village. EPA levels in red cell membrane phospholipids were higher in the fishing village than in the farming village. PGI2-, PGI3-, and TXA2/3-M levels were higher in the farming village than in the fishing village. There was a significant correlation between PGI2- and PGI3-M (r=0.80, n=40) and between PGI3-M and the EPA consumption during the day of urine collection (r=0.52, n=40). We conclude that PGI3 production is probably dependent less on the levels of EPA in tissues estimated by the EPA levels in red cell membranes than on the EPA consumption at the moment.