Effect of blood transfusion on gene expression of transforming growth factor beta-1 and the alpha subunit of interleukin-2 receptor in a model of colonic wound healing.

Blood transfusions have been shown in the past to compromise wound healing in experimental models of colonic anastomosis. Transforming growth factor beta (TGFbeta) has been proposed to play a major role in the process of colonic wound healing, and its expression is believed to be modulated by interleukin-2 (IL2). According to an earlier report, the postoperative administration of the histamine receptor-2 antagonist ranitidine increases the blood levels of soluble IL2 receptor in humans undergoing abdominal surgery and could thus affect molecular determinants of colonic anastomosis. In this study we examined the effect of blood transfusions, with and without ranitidine administration, on the gene expression of TGFbeta(1) and the specificity-conveying alpha subunit of the receptor for IL2, in rat perianastomotic tissue. Analysis of gene expression by semiquantitative RT-PCR revealed a tendency for downregulation of both genes, albeit in a statistically nonsignificant manner. This effect was not prevented by the postoperative administration of ranitidine. On the other hand, there appears to be a significant correlation between expression of TGFbeta(1) and that of IL2 receptor alpha subunit, irrespective of treatment.

Plasma atrial natriuretic peptide in essential hypertension after treatment with terazocin.

The aim of this study was to evaluate the medium term effects of the selective alpha1-adrenenergic- blocker terazocin on atrial natriuretic peptide (ANP) levels in patients with moderate essential hypertension. The drug was given orally for 30 days. The daily dose was 1 mg for the first 7 days, 2 mg for the next 7 days and 5 mg for the remaining period of this clinical trial. At the end of this clinical trial, plasma ANP levels increased by 16.40% despite the drop in blood pressure while left atrial and ventricular diameters remained unchanged. These findings indicate that the increase of ANP plasma levels is not the result of a mechanical load on the left cardiac chambers but the result of a pharmacological action. These observations also indicate that terazocin exerts part of its antihypertensive action by increasing ANP plasma levels.

The effect of losartan on calcium uptake by bovine aortic media in vitro.

The action of losartan, an AT1-angiotensin II receptor antagonist, on Ca2+ uptake by bovine aortic media in vitro was studied. This action was compared with that of nifedipine, a Ca2+ entry blocker. Ca2+ uptake values were obtained by measuring small slices of thoracic aorta tissue incubated in Kreb's solution in the presence of 45Ca. After preparation stimulation with angiotensin II (10 microM), losartan (1 microM and 10 microM) was shown to have a weaker inhibiting effect on Ca2+ uptake than nifedipine in equal concentrations. It is known that angiotensin II contracts smooth muscle by mobilizing intracellular Ca2+. The stimulating action on Ca2+ entry might be a secondary mechanism, which was studied for the first time in tissue slices derived from the aorta media. Results seem to be reasonable, since nifedipine also inhibited the spontaneous entrance of Ca2+, while losartan's action was focused only on the effects of angiotensin II.

Plasma atrial natriuretic peptide levels in essential hypertension after treatment with verapamil.

The aim of this study was to evaluate the long term effects of the selective Ca2+-blocker verapamil on atrial natriuretic peptide (ANP) levels in patients with moderate essential hypertension. The drug was given orally in a daily dose of 300 mg for 30 days. At the end of this clinical trial, plasma ANP levels increased by 16.14% despite the drop in blood pressure while left atrial and ventricular diameters remained unchanged. These findings indicate that the increase of ANP plasma levels is not the result of a mechanical load on the left cardiac chambers but the result of a pharmacological action. These observations also indicate that verapamil exerts part of its antihypertensive action by increasing ANP plasma levels.

Tissue structure-specific distribution of glycosaminoglycans in the human penis.

The aim of this work was to isolate and characterise the glycosaminoglycans present in the different tissue structures of the human penis in view of their potentially significant role in the physiology of erection. Penile tissue samples were obtained from patients who underwent penectomy and were subsequently dissected into individual tissue structures. Total glycosaminoglycans were isolated and purified from tunica albuginea, corpora cavernosa and corpus spongiosum, following tissue mincing, ultrasonication, lipid extraction, extensive digestion with pronase and DNase, treatment with alkali-borohydride and ethanol precipitation. Isolated glycosaminoglycans were separated by cellulose acetate electrophoresis and fractionated by anion exchange chromatography on DEAE Sephacel columns. Different glycosaminoglycan fractions were identified using glycosaminoglycan-degrading enzymes of known specificity. Gradient polyacrylamide gel electrophoresis was used to determine the average molecular mass of the glycosaminoglycans. The corpus cavernosum and the corpus spongiosum extracts contained almost twice the amount of glycosaminoglycan-associated uronic acids as compared to the tunical extracts (1.47+/-0.09, and 1.49+/-0.15 as opposed to 0.75+/-0.15 microg/mg dry defatted tissue, respectively; S.E.M., n=5). With the exception of hyaluronic acid, the relative amount of individual glycosaminoglycan types varied significantly among extracts of different origin. Heparan sulphate was more abundant in cavernosal, dermatan sulphate in tunical, and chondroitin-6-sulphate in corpus spongiosum extracts. No structure-specific differences were detected with respect to the molecular mass distribution of each glycosaminoglycan type. Our study shows that the different structures of the human penis produce distinct profiles of glycosaminoglycans, which are well suited to the individual functional characteristics of these structures.

Benign hyperplasia of the human prostate is associated with tissue enrichment in chondroitin sulphate of wide size distribution.

BACKGROUND: Benign prostatic hyperplasia (BPH) involves qualitative and quantitative alterations in extracellular matrix (ECM) components affecting stromal-epithelial interactions. Glycosaminoglycans (GAGs) are polysaccharide components of the ECM whose role in the development of BPH is under investigation. METHODS: GAGs were extracted from human prostates of normal and BPH origin and were subsequently fractionated through DEAE-sephacel anion exchange chromatography. The isolated GAG fractions were identified through electrophoresis on cellulose acetate membranes and treatment with GAG-degrading enzymes of known specificity. Their size distribution was determined through gradient polyacrylamide gel electrophoresis. RESULTS: Isolated prostatic GAGs included hyaluronic acid (HA), heparan sulphate (HS), and a mixture of dermatan sulphate (DS) and chondroitin sulphate (CS). The CS/DS ratio was significantly higher in hyperplastic as compared to normal prostates. A difference was also observed with respect to the apparent molecular mass of the DS-CS mixture, which reflects the CS enrichment in BPH. GAGs isolated from hyperplastic prostates were more diverse in size as compared to the corresponding glycans from normal prostates. CONCLUSIONS: The apparent increase in CS and decrease in DS content in prostates of patients with BPH is in good agreement with the pathological manifestation of increased cell proliferation in hyperplastic prostate tissue, since these glycan molecules have been reported to increase and decrease cell proliferation, respectively. Identification of the responsible enzymes involved in the homeostasis of CS and DS may provide alternative targets for pharmacological intervention.

A study of the effect of benzylpenicillin on isolated human gallbladder.

The contractile effect of benzylpenicillin on isolated rings of human gallbladder was studied using preparations taken from surgical specimens after gallbladderectomy. The gallbladder was cut into two pieces and one ring was cut from each piece; one ring was mounted in a 100 ml organ bath containing Krebs solution and the isotonic changes of the preparation were recorded for each experiment. Benzylpenicillin (1 x 10(4) to 1 x 10(8) IU/l) was shown to exert a dose-dependent contractile effect on isolated human gallbladder rings which was blocked by atropine (1 x 10(-8) M). The benzylpenicillin-induced contractile effect was analogous to contraction observed with acetylcholine (1 x 10(-7) to 1 x 10(-2) M). The effect of benzylpenicillin on isolated human gallbladder may reflect a possible decrease in gallbladder emptying time in vivo, suggesting a beneficial effect of benzylpenicillin in antimicrobial treatment of gallbladder infections.

A phase I clinical trial of dextromethorphan in intractable partial epilepsy.

We conducted an open-label pilot study of dextromethorphan (DM) in intractable partial epilepsy with the following objectives: a preliminary evaluation of the drug's safety and efficacy in the epileptic patient and a definition of a concentration range which can be safely achieved in future studies. Sixteen patients with drug-resistant, localization-related epilepsies entered the trial. After an 8-week baseline period, DM was added to the existing antiepileptic drugs at a dose of 40 and 50 mg every 6 h (160 and 200 mg/day). Each treatment period lasted 8 weeks. Seizure control improved after administration of DM, especially in the group of intermediate and slow metabolizers. Two patients, however, experienced increased seizure frequency and withdrew from the study. Adverse effects during DM administration were mild and transient. DM was well tolerated even in patients with high plasma levels of the drug (up to 15020 ng/dl). Our results indicate that DM is safe and effective in the treatment of comedicated patients with intractable partial epilepsies.

The differential distribution of hyaluronic acid in the layers of human atheromatic aortas is associated with vascular smooth muscle cell proliferation and migration.

Vascular smooth muscle cells (VSMC), under conditions of induced proliferation, similar to those involved in atherosclerosis, secrete an acidic glycan, 82% of which exhibits structural homology with hyaluronic acid (HA), has a molecular mass of 340 kDa (HA-340) and inhibits VSMC proliferation in vitro. In this study, the expression of glycans was investigated in human atheromatic aortas and evidence is presented that a HA molecule, similar to HA-340, is distinctly expressed in all aortic layers. The isolation of the glycans from human aortas was performed after homogenization of the individual aortic layers (atheromatic plaque, tunica intima, tunica media and tunica adventitia), by lipid extraction and extensive digestion with pronase and DNase. The total glycans were purified from the digestion products by gel filtration on Sephadex G-25 and fractionated on a Superose 6 column. Enzymatic treatment of the ensuing glycan fractions with all known glycosaminoglycan-degrading enzymes, followed by electrophoresis on polyacrylamide gradient gels and cellulose acetate membranes, revealed that, in addition to HA, the tunica intima and the atheromatic plaque also contained dermatan sulfate, while the tunica media and the tunica adventitia also contained chondroitin sulfates and heparan sulfate. The highest concentration of the human aorta HA was found in the tunica media, exhibiting a negative concentration gradient from the tunica media to the atheromatic plaque. Investigation of the biological function of the human aorta HA revealed that this molecule acts as a negative regulator on the PDGF-induced VSMC proliferation and as a positive regulator on the PDGF-induced VSMC migration. The differential expression of HA within the aortic layers correlates with the biological function attributed to this acidic glycan and associates it with key events in the progression of atherogenesis.

Time course of isolated rat fundus response to muscarinic agonists: a measure of intrinsic efficacy.

The establishment of a dose-response relationship and its quantification is the usual procedure for analysing drug action on an isolated organ. However, the time course of the effect seems to be an inherent characteristic of the agonist which produces it. In our study, we have analyzed the time-response curves of four cholinergic agonists (acetylcholine, methacholine, carbachol and bethanechol) which produce tonic contractions of the isolated rat gastric fundus. The order of affinity of agonists to muscarinic receptors on the rat fundus were carbachol > bethanechol > methacholine > acetylcholine (K(A) values: 46 +/- 12, 84 +/- 21, 380 +/- 110 and 730 +/- 120 nM, respectively). The effective concentrations which produced 60% of the maximal response (EC60) were used for establishing the time-response curves. The time-response curves were also recorded after partial alkylation of muscarinic receptors with phenoxybenzamine, after exposure of the isolated rat fundus to physostigmine and after addition of supramaximal concentrations of the agonists. The experimental time-response curve for acetylcholine was on the extreme left, followed by curves for methacholine, bethanechol and carbachol, respectively. Phenoxybenzamine and supramaximal doses of the agonists did not change the order of response development in time, but supramaximal doses shifted all curves to the left and phenoxybenzamine shifted all time-response curves to the right. Only physostigmine shifted the time-response curve for methacholine to the right. The results of our study suggest that the response rate of the isolated rat gastric fundus to cholinergic agonists depends on the intrinsic activity of these agents, but not on their affinity for muscarinic receptors.

Increased type IV collagen-degrading activity in metastases originating from primary tumors of the human colon.

The secretion of specific matrix metalloproteinases (MMPs) is considered a prerequisite step for tumor cell invasion and metastasis. In the present study we investigated the expression of type IV collagen-degrading activity in primary tumors of the human colon in correlation to tumor grade and in comparison to activity expressed in arising metastases. We observed that type IV collagen-degrading activity (MMP-2 and MMP-9), purified by ion exchange, gel filtration and affinity chromatography and characterized by gelatin zymography, correlates to tumor grade. Furthermore, in surgical specimens identified as metastases originating from primary tumors of the colon, we observed that enzyme activity was significantly enhanced, relatively to that identified in the primary tumor. This observation should be considered when targeting MMPs as a therapeutic intervention to prevent cancer progression.