Adaptation to 5 weeks of intermittent local vascular pressure increments; mechanisms to be considered in the development of primary hypertension?

The aims were to study effects of iterative exposures to moderate elevations of local intravascular pressure on arterial/arteriolar stiffness and plasma levels of vasoactive substances. Pressures in the vasculature of an arm were increased by 150 mmHg in healthy men (n = 11) before and after a 5-wk regimen, during which the vasculature in one arm was exposed to fifteen 40-min sessions of moderately increased transmural pressure (+65 to +105 mmHg). This vascular pressure training and the pressure-distension determinations were conducted by exposing the subjects' arm versus remaining part of the body to differential ambient pressure. During the pressure-distension determinations, venous samples were simultaneously obtained from pressurized and unpressurized vessels. Pressure training reduced arterial pressure distension by 40 ± 23% and pressure-induced flow by 33 ± 30% (P < 0.01), but only in the pressure-trained arm, suggesting local adaptive mechanisms. The distending pressure-diameter and distending pressure-flow curves, with training-induced increments in pressure thresholds and reductions in response gains, suggest that the increased precapillary stiffness was attributable to increased contractility and structural remodeling of the walls. Acute vascular pressure provocation induced local release of angiotensin-II (ANG II) and endothelin-1 (ET-1) (P < 0.05), suggesting that these vasoconstrictors limited the pressure distension. Pressure training increased basal levels of ET-1 and induced local pressure release of matrix metalloproteinase 7 (P < 0.05), suggesting involvement of these substances in vascular remodeling. The findings are compatible with the notion that local intravascular pressure load acts as a prime mover in the development of primary hypertension.NEW & NOTEWORTHY Adaptive responses to arterial/arteriolar pressure elevation have typically been investigated in cross-sectional studies in hypertensive patients or in longitudinal studies in experimental animals. The present investigation shows that in healthy individuals, fifteen 40-min, carefully controlled, moderate transmural pressure elevations markedly increase in vivo stiffness (i.e. reduce pressure distension) in arteries and arterioles. The response is mediated via local mechanisms, and it appears that endothelin-1, angiotensin-II, and matrix metalloproteinase 7 may have key roles.

Purification and partial characterization of a 17 beta-estradiol-binding macromolecule in the human pancreas.

Studies have been performed in order to investigate the presence of estrogen-binding proteins in the human pancreas that may provide the biochemical basis for tissue-specific treatment of pancreatic carcinoma with estrogen-based cytotoxic drugs. Using in vitro techniques, an estrogen-binding macromolecule has been purified from pancreatic cytosol. With estradiol a ligand, Kd was calculated to be 1.7 X 10(-7) M, and this protein was found to constitute about 4% of the total protein content in the cytosol. No metabolism of estradiol was detected under the in vitro conditions used. Competition experiments indicated that, besides estradiol, the protein also had some affinity for estrone and estriol but not for testosterone, progesterone, or dexamethasone. The protein was purified to homogeneity using chromatography on concanavalin A and hydroxylapatite followed by preparative polyacrylamide gel electrophoresis. The purified protein, still able to bind, [3H]-estradiol, gave one single protein-staining band when analyzed using different electrophoretic systems. The steroid-protein and did not bind to phosphocellulose or DNA-cellulose and did not show any similarities to steroid receptor proteins. The complex has a Strokes' radius of 52 A and a sedimentation coefficient of 3S. The biological significance of the macromolecule is known, but the protein is probably synthesized in the pancreas since no similar protein could be detected in serum. Studies are now being carried out to investigate whether this novel protein in the human pancreas may interact with complexes between cytotoxic agents and estrogens and provide the basis for tissue-specific treatment of pancreatic carcinoma.

Pancreas-specific protein. New serum marker for graft rejection in pancreas-transplant recipients.

A radioimmunoassay for a novel human pancreatic protein (pancreas-specific protein, PASP) has been developed. We studied the possibility that serum PASP levels reflect pancreas-graft rejections in human pancreas-transplant recipients. Ten patients subjected to combined pancreas-kidney transplantation and 4 patients subjected to pancreas transplantation alone were studied. Twelve kidney recipients served as control subjects. On several occasions, PASP levels were elevated at kidney rejections in patients with combined pancreas-kidney grafts and then decreased after antirejection therapy, although no other indications for concomitant pancreas-graft rejection were at hand. In the recipients of pancreas grafts alone, PASP levels increased before or at the same time as graft rejections were indicated by current methods. In two cases of chronic graft rejection, PASP rose to high levels long before hyperglycemia occurred. In the control group of kidney-graft recipients, PASP levels were stable and were not affected by high serum creatinine levels, kidney-rejection episodes, or antirejection therapy. This study indicates that PASP may be a good serum marker for pancreas-graft rejection.

Novel assay for pancreatic cellular damage: 1. Characterization of protein profiles in human pancreatic cytosol and purification and characterization of a pancreatic specific protein.

The protein patterns of cytosols from normal human pancreas and pancreatic carcinoma were studied by a two-dimensional separation technique using high-performance liquid chromatography followed by isoelectric focusing on polyacrylamide gels and visualization of the focused proteins by Coomassie Blue staining. Almost identical protein patterns were obtained for 20 different specimens from normal pancreas, whereas quite different protein patterns were found in 12 samples of pancreatic carcinoma. A major protein in normal pancreatic cytosol, not identical to any macromolecule previously tested as a marker for pancreatic function, was selected for further studies. The protein was not found in specimens of pancreatic carcinoma. It was purified by a single step chromatofocusing procedure, focused at pH 6.9, and moved as one single band in sodium dodecyl sulfate-polyacrylamide gel electrophoresis with an apparent molecular weight of 44,500 daltons. Total amino acid analysis revealed a high concentration of glutamic acid, leucine, and lysine. The purified protein had no amylase activity or lipase immunoactivity. It constituted approximately 2% of the total normal pancreatic cytosol protein. Later immunological studies have shown the protein to be highly specific for normal human pancreas, indicating a possible future use as a marker for pancreatic cell damage.

A novel assay for pancreatic cellular damage: IV. Serum concentrations of pancreas-specific protein (PASP) in acute pancreatitis and other abdominal diseases.

Pancreas-specific protein (PASP) was compared with serum amylase in 95 episodes of acute pancreatitis with the diagnoses supported by elevated amylase levels. The etiology was typical for Scandinavian countries, with alcohol as the predominant factor, followed by cholelithiasis. PASP values were clearly raised in all patients, except in three cases found to have high salivary-type amylase levels, and one patient with recurrent alcohol pancreatitis. The rise of PASP levels were in general more pronounced than the corresponding amylase elevations, especially in severe pancreatitis. The elevations were generally parallel for the two analytes, but in 41% of the cases PASP levels remained elevated 2-11 days longer than the corresponding amylase levels. PASP was, however, eliminated from the circulation at a rate comparable to that of amylase. The serum range of PASP for 259 healthy subjects was 15-111 micrograms/L with 95% of the values within 16-98 micrograms/L. The upper reference level was set at 100 micrograms/L. PASP levels were also determined for 291 patients with disorders other than acute pancreatitis. Serum levels in patients with renal insufficiency (n = 12), primary biliary cirrhosis (n = 9), and diabetes mellitus (n = 17) were equal to those in healthy subjects. Eight patients of 173 with acute abdominal disorders and no evidence of pancreatitis had elevated PASP levels as well as 4 patients with prostatic carcinoma (n = 28) and 2 patients with benign prostatic hyperplasia (n = 16). PASP values were low in chronic painful pancreatitis (n = 15) and pancreatic cancer (n = 11).

A novel serum assay for pancreatic cellular damage. II. High tissue specificity of a pancreatic protein.

A previously unknown major protein in human pancreatic cytosol has been purified and partly characterized. The protein, designated pancreas specific protein (PASP), has a molecular weight of 44,500 and a pI of 6.9. A two-dimensional gel separation technique revealed the protein to be specific for normal pancreatic tissue. Antibodies against PASP were raised in rabbits and a radioimmunoassay was developed for the quantitation of this protein. The following concentrations of PASP (mg/kg wet weight) were found in human tissues: normal pancreas 100-1,000; pancreatic carcinoma 0.1-20; prostate 0.5-5; and 13 other tissues less than 0.5. The levels of PASP in peripheral serum were less than 0.1 mg/L in normal subjects, 0.7-3 mg/L in cases of acute pancreatitis, and less than 0.1 mg/L in cases of pancreatic carcinoma, prostatic diseases, and other abdominal diseases investigated. The high tissue specificity and the specific elevation of serum PASP levels in acute pancreatitis may indicate a use of this protein as a marker of this pancreatic condition.

A novel assay for pancreatic cellular damage: III. Use of a pancreas-specific protein as a marker of pancreatic graft dysfunction in humans.

Pancreas-specific protein (PASP) is a recently isolated and partially characterized major protein in the human pancreas. It has not been described previously. Serum levels of PASP and amylase were analyzed in 21 patients subjected to combined renal and segmental pancreatic transplantation with both organs obtained from the same donor and in eight kidney transplant patients. In the pancreas transplant patients, PASP and amylase levels were elevated in episodes of graft pancreatitis. With chronic graft rejection, PASP rose to high levels long before other indications. In episodes of renal rejection, the levels of PASP, but not always of amylase, were elevated on several occasions. They decreased after antirejection therapy. This may indicate accompanying pancreatic graft rejection. PASP and amylase levels were stable in kidney transplant patients and were not affected by serum creatinine levels, renal rejection, or antirejection therapy. The results support earlier observations that renal rejection in combined pancreas and renal transplant patients may or may not be accompanied by a rejection process in the pancreatic graft. PASP may be the means by which to tell when the pancreatic graft is involved.

Metabolism of estrone sulphate by benign prostatic hyperplasia in vitro.

Homogenates of benign prostatic hyperplastic (BPH) tissue and abdominal muscle were incubated with [3H] estrone sulphate in the presence of NADH and NADPH. The metabolites were extracted with chloroform, isolated by thin layer chromatography (TLC) and identified by addition of 14C-labelled standards followed by sequential chromatography and acetylation. Both tissues transformed the substrate into [3H] estrone and [3H] estradiol-17 beta, but BPH tissue by far exceeded the abdominal muscle in this respect. Estriol and other polar metabolites could not be detected. The specific ability of BPH tissue to convert the major peripheral estrogen estrone sulphate into the terminal biologically active estrogen estradiol-17 beta further indicates an etiological role of estrogens in the development of human BPH.

Serum hormone levels in benign prostatic hyperplasia.

The peripheral serum levels of FSH, LH, testosterone and total estrone (sum of free + conjugated estrone, mainly estrone sulphate) were determined in 20 patients with benign prostatic hyperplasia (BPH) aged 59--79 years and in 49 healthy men aged 58--79 years. There were no significant differences in the levels of FSH, LH and testosterone. The level of total estrone was significantly higher in the BPH group (p less than 0.05). The results are in accordance with our previous study on the urinary estrogen excretion in BPH and give additional support to the hypotheses concerning a role of the estrogens in the etiology of BPH.

Influence of aging upon the urinary hormone excretion in the male.

The urinary excretion of LH, low polar oestrogens, neutral C19 and C21 steroids was measured in normal healthy males aged 20-79 years. No significant changes could be noted for the excretion of LH, pregnanediol and 17-ketogenic steroids between 50 and 79 years. The excretion of androsterone and aeticholanolone was significantly lower in the oldest group (70-79 years). A significant drop in the excretion of DHA was noted at approx. 60 years. The excretion of low polar oestrogens remained constant from 20 to 55-59 years, but showed a highly significant (P less than 0.001) decrease at about 60 years. This drop might reflect a sudden decrease in the plasma levels of oestrone sulphate due to a decreased sulphurylating activity in the liver.

Further characterization of the estrogen binding macromolecule in human pancreas.

The estrogen binding protein in human pancreas has been purified from pancreatic cytosol by chromatography on Concanavalin-A-Sepharose and hydroxyl-apatite followed by ion-exchange chromatography carried out using a fast-protein liquid chromatography apparatus (FPLC). The purified protein, still able to bind labelled [3H]estradiol, appeared as one single band corresponding to 31 K in SDS-gel electrophoresis. Total amino acid analysis revealed high levels of histidine, glutamic acid and leucine. The capacity of the purified protein to bind estrogens could be increased more than 4-fold by addition of a cytosolic factor, probably being a small peptide, that is present in crude cytosol, but lost during the purification procedure. The iodinated protein does not bind to DNA-cellulose or phosphocellulose, and shows no similarities to estrogen receptor proteins.

Effects of tamoxifen on the serum levels of oestrogens and adrenocortical steroids in postmenopausal breast cancer patients.

Peripheral serum levels of cortisol, dehydroepiandrosterone (DHA), dehydroepiandrosterone sulphate (DHA-S), oestradiol-17 beta and total oestrone were measured in 17 postmenopausal breast cancer patients and on age-matched healthy controls. In the breast cancer patients analyses were performed before and after 1, 2, 4, 8 and 12 weeks of treatment with tamoxifen. Basal DHA levels were significantly higher in breast cancer patients than in controls; otherwise no significant differences were noted. Treatment with tamoxifen resulted in significantly elevated levels of cortisol throughout the period of treatment; this effect was probably solely due to an increase in transcortin levels. A transient but significant elevation in total oestrone, probably related to liver function, was observed after one week of treatment. No significant effects of tamoxifen were noted on serum DHA, DHA-S or oestradiol-17 beta. The results indicate that tamoxifen does not affect the adrenocortical steroid biosynthesis in postmenopausal women. The oestrogenic effects of tamoxifen upon FSH, prolactin and the oestrogen induced Pregnancy Zone Protein (PZP) in postmenopausal women, described in a previous communication from this group, are probably not due to any increase in the serum oestrogens since this increase is transient and hardly impressive. The weak oestrogenic effect of the tamoxifen-receptor complex itself may thus be sufficient to produce net oestrogenic effects in subjects with low endogenous oestrogens such as postmenopausal women.

Analysis of an estrogen binding macromolecule in human pancreas by radioimmunoassay.

A radioimmunoassay for quantitation of the human pancreatic estrogen binding protein (hEBP) was developed using polyclonal rabbit hEBP antiserum and iodinated purified hEBP. Parallel dose-response curves were obtained when serial dilutions of human serum and of cytosols obtained from human pancreas, prostate and colon were analyzed simultaneously with serial dilutions of purified hEBP standard. Very high levels of hEBP (500-1000 mg/kg wet weight) were found in normal pancreas. High as well as medium levels were found in pancreatic carcinoma tissue and medium values (0.1-1 mg/kg wet weight) in prostate, colon and ovarian tissue. Other tissues and serum from healthy volunteers showed low values, usually below 0.1 mg/kg. When serial dilutions of rat pancreatic cytosol were analyzed in the hEBP assay, [125I]hEBP was displaced by the rat preparation, but the dose-response curves were not parallel to the standard curves, indicating similarity but non-identity between the estrogen binding proteins in human and in rat pancreas.

The estrogen binding protein in human pancreas: concentrations in subcellular fractions of normal pancreatic tissue, in duodenal juice during pancreatic stimulation and in peripheral serum in normal and pathological conditions.

The steroid binding properties of the human pancreatic estrogen binding protein (hEBP) in cytosol were studied by equilibrium dialysis. A high ligand specificity of the protein was revealed. hEBP in cytosol binds unconjugated steroid estrogens with a medium affinity (Kd = 10(-7) M) but does not bind conjugated estrogens or unconjugated androgens, gestagens, glucocorticoids or cholesterol. Quantitation of hEBP by radioimmunoassay in subcellular fractions of human pancreatic tissue indicated that the protein is translocated into different subcellular compartments. Duodenal juice taken from patients following stimulation of pancreatic secretion by food ingestion (Lund's test) showed high hEBP concentrations, and the levels of hEBP changed concomitantly with the levels of pancreatic isoamylase, indicating that hEBP secretion was stimulated by food ingestion. The levels of hEBP in peripheral serum from healthy subjects showed no sex difference, but were positively correlated to age. Highly elevated (10-20-fold) hEBP levels were found in serum from patients with acute pancreatitis, while normal serum hEBP values were found in other abdominal diseases. It is speculated that hEBP might have a specific role in the transport of estrogens from the peripheral circulation via the pancreas to the duodenum. The elevated hEBP levels in patients with acute pancreatitis indicate that this protein may be used as a marker of cellular damage in the pancreas.