Prostacyclin is not a circulating hormone

Gas chromatography with electron-capture detection of the extensively purified pentafluorobenzyl derivative of 6-oxo-PGF_1α was used to determine prostacyclin in blood. Neither human peripheral plasma or whole blood, nor blood drawn directly from the human heart (blood from the right and left atrium which is comparable to pulmonary artery and vein blood), contained any detectable prostacyclin (< 20 pg/ml). Even hyperventilation did not result in detectable PGI₂-formation. During intravenous infusion of PGI₂ into one arm, large amounts were found in blood drawn from the other arm. Increased levels were also found during severe infection and in endotoxin shock. These results lend no support to theories based on the concept that prostacyclin is a circulating hormone under normal conditions.     

Atrial natriuretic factor is a circulating hormone

The radioimmunoassay of atrial natriuretic factor (ANF) has been applied for determination of immunoreactive ANF (IR-ANF) in rat plasma. Immunoreactive ANF has been extracted from rat plasma by immunoaffinity column on Sepharose-4B anti-ANF or by Vycor glass. The mean concentrations of IR-ANF in ether anesthetized rats were found to be 1.61 +/- 0.14 ng/ml in female and 1.25 +/- 0.21 ng/ml in male rats when extracted on Sepharose-4B anti-ANF, and 1.21 +/- 0.10 ng/ml in females and 1.02 +/- 0.11 ng/ml in males when extracted by Vycor glass. A close linear correlation has been observed between the plasma IR-ANF concentrations in aorta and jugular vein. The described results indicate that atrial cardiocytes secrete atrial natriuretic factor into plasma. The heart is, therefore, an endocrine organ.     

Prostacyclin does not affect insulin secretion in humans

The effects of Prostacyclin (PGI₂) infusion on insulin secretion and glucose tolerance were investigated in 7 healthy subjects. PGI₂ infusion caused no statistically significant changes of either glucose or insulin concentration, over the range 2.5–20 ng/Kg/min. A constant PGI₂ infusion (10 ng/Kg/min) did not inhibit acute insulin responses to a glucose (20 g i.v.) pulse (response before PGI₂ = 612±307%; during PGI₂ = 515±468%, mean ± SD, mean change 3–5 min insulin, % basal; P=NS). Glucose disappearance rates were similar after the first and second glucose pulse.Thus, in contrast to PGE₂, PGI₂ does not affect insulin secretion nor glucose disposal at doses producing platelet and vascular changes. It is hypothesized that an altered PGI₂/PGE₂ balance in diabetes may represent a link between vascular, platelet and metabolic changes.     

25-Hydroxyvitamin D3 3-sulphate is a major circulating form of vitamin D in man

25-Hydroxyvitamin D3 3 beta-sulphate has been identified in human plasma. The compound was isolated by anion-exchange chromatography and following hydrolysis it was characterized by high-performance liquid chromatography and gas chromatography-mass spectrometry. The mean concentration of sulphated 25-hydroxyvitamin D3 in plasma from 60 patients was 16.7 +/- 7.1 ng/ml and the levels often exceeded those of the corresponding free compound. The study also shows that unconjugated 25-hydroxyvitamin D3 is not readily sulphated by man in vivo.     

Prostacyclin inhibition of phosphatidic acid synthesis in human platelets is not mediated by protein kinase C

The activation of protein kinase C in human platelets by phorbol-12, 13- dibutyrate (PDBu) results in the phosphorylation of a 40,000 dalton protein. This phosphorylation is time- and concentration-dependent. Maximal phosphorylation is rapid and is not affected by indomethacin or prostacyclin. PDBu does not promote activation of the phosphodiesteratic cleavage (phospholipase C) of the inositol phospholipids and the subsequent formation of 1,2-diacylglycerol or its phosphorylated product, phosphatidic acid. If platelets exposed to PDBu are subsequently stimulated with thrombin, this stimulus does not initiate further 40,000 dalton protein phosphorylation but will promote the formation of phosphatidic acid and also the phosphorylation of a 20,000 dalton protein (myosin light chain). However, prostacyclin will prevent the subsequent stimulation of phosphatidic acid synthesis by thrombin in a concentration-dependent manner. The fact that prostacyclin can affect the response to thrombin, even in the presence of phorbol ester, supports the idea that the enzymes related to the formation of phosphatidic acid or inhibition of its synthesis are not related to the phosphorylated 40K protein.     

Nitrite is produced by elicited but not by circulating neutrophils

The generation of nitrite (NO(2) (-)) was used as an index of the production of nitric oxide by human and rat polymorphonuclear leukocytes (PMN) and rat peritoneal macrophages. Human peripheral blood PMN did not produce significant levels of NO(2) (-). Attempts to induce NO(2) (-) generation in human PMN by incubation with GM-CSF (1 nM), TNFalpha (0.3 nM), endotoxin (1 mug/ml) or formyl-Met-Leu-Phe (100 nM) for up to 16 h were not successful. Addition of human PMN primed by GM-CSF (1 nM) to rabbit aortic ring preparations precontracted with phenylephrine had no effect on tone. In contrast to these observations, PMN, isolated from the peritoneum of oyster glycogen treated rats, generated NO(2) (-) via a pathway sensitive to inhibition by the nitric oxide synthase inhibitor, N(G)-monomethyl L-arginine. However, peripheral blood rat PMN obtained from the same animals did not produce NO(2) (-), even during prolonged incubation for periods of up to 16 h. It is suggested that detectable NO production by PMN requires NO synthase activity to be induced either by the process of PMN migration or by exposure to certain cytokines produced locally at the site of inflammation.     

Red pigment-concentrating hormone is not limited to crustaceans

A peptide that was previously assumed to occur exclusively in crustaceans is found in the corpora cardiaca of the stinkbug, Nezara viridula. The sequence of the peptide was deduced from the multiple MS(N) electrospray mass data as that of an octapeptide: pGlu-Ile/Leu-Asn-Phe-Ser-Pro-Gly-Trp amide. This peptide with Leu at position 2 is known as crustacean red pigment-concentrating hormone and code-named Panbo-RPCH. The ambiguity about the amino acid at position 2, Leu or Ile, was solved by isolating the peptide in a single-step by reversed-phase HPLC and establishing co-elution with authentic Panbo-RPCH but not with the Ile(2)-analog. When injected into stinkbugs, synthetic Panbo-RPCH elicited an increase of lipids in the haemolymph. Thus, it is assumed that Panbo-RPCH functions in the stinkbug as a lipid-mobilizing hormone.     

U6 is not a suitable endogenous control for the quantification of circulating microRNAs

Recently, microRNAs have been detected in serum and plasma, and circulating microRNA (miRNA) profiles have now been associated with many diseases such as cancers and heart disease, as well as altered physiological states. Because of their stability and disease resistance, circulation miRNAs appear to be an ideal material for biomarkers of diseases and physiological states in blood. However, the lack of a suitable internal reference gene (internal reference miRNA) has hampered research and application of circulating miRNAs. Currently, U6 and miR-16 are the most common endogenous controls in the research of miRNAs in tissues and cells. We performed microarray-based serum miRNA profiling on the serum of 20 nasopharyngeal carcinoma patients and 20 controls to detect the expressions of U6 and miRNAs. Profiling was followed by real-time quantitative Polymerase Chain Reaction (qPCR) in 80 patients (20 each with gastric cancer, nasopharyngeal carcinoma, colorectal cancer, and breast cancer) and 30 non-cancerous controls. qPCR was also performed to detect miRNAs in serum with repeated freezing and thawing. The results of microarray showed that with the exception of U6, Ct values of miR-16, miR-24, miR-142-3p, miR-19b and miR-192 in serum samples of nasopharyngeal carcinoma were greater than control samples. The results of 110 cases showed large fluctuations in U6 expression. The difference between the greatest and the least levels of expression was 3.29 for delta Ct values, and 1.23 for miR-16. The expressions of U6, miR-16 and miR-24 in serum subjected to different freeze–thaw cycles showed that U6 expression gradually decreased after 1, 2, and 4 cycles of freezing and thawing, while the expression of miR-16 and miR-24 remained relatively stable. Collectively, our results suggested that U6 is unsuitable as an internal reference gene in the research of circulating miRNAs.     

Decreased circulating growth hormone levels following centrally administered insulin-like growth factor-1 is not mediated by somatostatin in the pig fetus.

Twenty-six pig fetuses (at 94 days gestational age) were fitted with carotid artery catheters. Eight fetuses were given 1,500 ng of IGF-1 (in 100 microliters) directly into lateral cerebral ventricle; 7 further fetuses received the IGF-1 together with 150 microliters of a potent specific anti-somatostatin serum into a ventricle, 5 other fetuses received the anti-somatostatin serum alone while 6 controls received normal sheep serum. Administration of IGF-1 caused a rapid decrease in circulating growth hormone (pGH) levels but there was no significant change in plasma levels of somatostatin immediately following the IGF-1 administration, suggesting that the decrease in pGH was not mediated by somatostatin secretion. Further evidence that somatostatin was not involved in this effect was provided by the lack of effect of concurrent antisomatostatin serum on the IGF-1-induced decrease in pGH. Thus the high circulating levels of GH in the fetus may result from a lack of feedback of IGF-1, but not through the somatostatin-pituitary axis.     

Is 24,25-dihydroxycholecalciferol a calcium-regulating hormone in man?

Small doses (1-10 microgram daily) of 24,25-dihydroxycholecalciferol (24,25-(OH)2D3), a renal metabolite of vitamin D of uncertain function, increased intestinal absorption of calcium in normal people and in patients with various disorders or mineral metabolism, including anephric subjects. In five of six patients studied, calcium balance increased, but, unlike 1,25-dihydroxycholecalciferol, 24,25-(OH)2D3 did not increase plasma or urinary calcium concentrations. These results suggest that 24,25-(OH)2D3 may be an important regulator of skeletal metabolism in man with potential value as a therapeutic agent.     

Measurement of circulating parathyroid hormone

The measurement of parathyroid hormone (PTH) is important in the investigation of disorders of mineral metabolism. Though the first immunoassay for PTH was described more than 25 years ago, it is only recently that methods have been developed which provide the required sensitivity and specificity to detect small changes in hormone secretion in normal subjects. These new methods take advantage of modern labelled antibody technology and have already been shown capable of yielding improvements in the diagnosis and monitoring of disorders which involve changes in PTH secretion. In addition, they are now providing fresh insight into the basic physiology of PTH secretion.     

Cell membrane, but not circulating, carcinoembryonic antigen is linked to a phosphatidylinositol-containing hydrophobic domain

Carcinoembryonic antigen is present in the cell membrane of most tumors of colorectal origin and in the plasma of patients with colorectal cancer and other malignancies. In this paper we demonstrate that carcinoembryonic antigen can be released from HT-29 cells by phosphatidylinositol specific phospholipase C. Triton X-114 phase separation shows that phospholipase C converts the antigen into a water soluble protein. In addition, plasma carcinoembryonic antigen behaves as the cleaved antigen in phase separation experiments. This strongly suggests that carcinoembryonic antigen is attached to cell membranes by a glycosyl-phosphatidylinositol anchor and that it can be released in vivo by enzymatic cleavage of the hydrophobic tail.     

Atrial natriuretic hormone is not elevated during dopamine induced natriuresis

To evaluate the possibility that atrial natriuretic hormone (ANH) is involved in dopamine induced natriuresis and diuresis, we studied five normal male volunteers. Each was studied on two occasions. During the first two hours of each study, normal saline, 25 ml/hr, was infused. During the second two hours either normal saline, 25 ml/hr, or dopamine, 4 micrograms/kg/min, in normal saline, was infused. Dopamine infusion caused prominent and significant natriuresis and diuresis but plasma levels of immunoreactive ANH levels did not change. We conclude that the ANH is not involved in dopamine induced natriuresis and that dopaminergic stimulation is not responsible for ANH secretion.     

The circulating metabolic regulator FGF21 is induced by prolonged fasting and PPARalpha activation in man

FGF21 is a critical metabolic regulator, pivotal for fasting adaptation and directly regulated by PPARα in rodents. However, the physiological role of FGF21 in man is not yet defined and was investigated in our study. Serum FGF21 varied 250-fold among 76 healthy individuals and did not relate to age, gender, body mass index (BMI), serum lipids, or plasma glucose. FGF21 levels had no diurnal variation and were unrelated to bile acid or cholesterol synthesis. Ketosis induced by a 2 day fast or feeding a ketogenic diet (KD) did not influence FGF21 levels, whereas a 74% increase occurred after 7 days of fasting. Hypertriglyceridemic nondiabetic patients had 2-fold elevated FGF21 levels, which were further increased by 28% during fenofibrate treatment. FGF21 circulates in human plasma and increases by extreme fasting and PPARα activation. The wide interindividual variation and the induction of ketogenesis independent of FGF21 levels indicate that the physiological role of FGF21 in humans may differ from that in mice.     

Circulating CXCL16 is not related to circulating oxLDL in patients with rheumatoid arthritis

CXCL16 acts as a scavenger receptor for oxLDL in its membrane-bound form and induces migration of activated T cells in its soluble form. Due to these properties, CXCL16 has been suggested to play a role in both atherosclerosis and rheumatoid arthritis (RA). Our aim was to evaluate the contribution of soluble CXCL16 to the scavenging of oxLDL and its potential as a marker for cardiovascular disease (CVD) in patients with RA. We found that circulating CXCL16 was not correlated with plasma oxLDL or ApoB and was not related to the presence of CVD in RA patients. Moreover, CXCL16 did not bind and scavenge oxLDL in an in vitro setting. These data suggest that binding of oxLDL by soluble CXCL16 does not play a role in atherosclerosis and, although confirmation in larger studies is needed, that circulating CXCL16 is not related to the presence of CVD in patients with RA.     

Insulinhypoglycemia but not arginine infusion stimulates circulating plasma growth hormone-releasing hormone (GHRH) concentrations in children

The effect of insulinhypoglycemia and arginine infusion on circulating concentrations of plasma growth hormone-releasing hormone (GHRH) and growth hormone (GH) has been studied in 24 children (4.4 to 14.3 years). Plasma GH and GHRH concentrations were determined by RIA. Basal plasma GHRH levels were detectable in the plasma of all patients ranging from 6.8 to 27.1 pg/ml. Injection of 0.1 U/kg body wt. insulin i.v. resulted in an increase of plasma GHRH levels (11.1 +/- 1.4 pg/ml vs. 18.8 +/- 2.6 pg/ml; P less than 0.01) preceding that of plasma GH (1.5 +/- 0.4 ng/ml vs. 13.6 +/- 1.3 ng/ml; P less than 0.01). Infusion of 0.5 gm/kg body wt. arginine hydrochloride did increase GH concentrations (2.0 +/- 0.6 ng/ml vs. 13.9 +/- 2.3 ng/ml; P less than 0.01) but did not change circulating plasma GHRH levels. Since the source of peripheral GHRH concentrations is not known the importance of these findings remains to be determined.     

A new polymorphism in the type II deiodinase gene is associated with circulating thyroid hormone parameters

Type II deiodinase (D2) is important in the regulation of local thyroid hormone bioactivity in certain tissues. D2 in skeletal muscle may also play a role in serum triiodothyronine (T(3)) production. In this study, we identified a polymorphism in the 5'-UTR of the D2 gene (D2-ORFa-Gly3Asp). We investigated the association of D2-ORFa-Gly3Asp, and of the previously identified D2-Thr92Ala polymorphism, with serum iodothyronine levels. D2-ORFa-Gly3Asp was identified by sequencing the 5'-UTR of 15 randomly selected individuals. Genotypes for D2-ORFa-Gly3Asp were determined in 156 healthy blood donors (age 46.3 +/- 12.2 yr) and 349 ambulant elderly men (age 77.7 +/- 3.5 yr) and related to serum iodothyronine and TSH levels. D2-ORFa-Asp(3) had an allele frequency of 33.9% in blood bank donors and was associated with serum thyroxine (T(4); Gly/Gly vs. Gly/Asp vs. Asp/Asp = 7.06 +/- 0.14 vs. 6.74 +/- 0.15 vs. 6.29 +/- 0.27 microg/dl, P = 0.01), free T(4) (1.22 +/- 0.02 vs. 1.16 +/- 0.02 vs. 1.06 +/- 0.04 ng/dl, P = 0.001), reverse T(3) (P = 0.01), and T(3)/T(4) ratio (P = 0.002) in a dose-dependent manner, but not with serum T(3) (P = 0.59). In elderly men, D2-ORFa-Asp(3) had a similar frequency but was not associated with serum iodothyronine levels. This new polymorphism in the 5'-UTR of D2 is associated with iodothyronine levels in blood donors but not in elderly men. We hypothesize that this might be explained by the decline in skeletal muscle size during aging, resulting in a relative decrease in the contribution of D2 to serum T(3) production.