Nebracetam (WEB 1881FU) prevents N-methyl-D-aspartate receptor-mediated neurotoxicity in rat striatal slices. The effects of nebracetam were investigated on N-methyl-D-aspartate (NMDA) receptor- and voltage-operated Ca2+ channels (VOCC)-mediated neural dysfunction by directly monitoring the real-time dynamics of dopamine released from rat striatal slices. Nebracetam (10(-5) and 10(-4) M) completely protected against striatal dopaminergic impairment induced by L-glutamate and NMDA, respectively. BAY K-8644-evoked striatal dysfunction was not blocked by nebracetam (10(-4) M). Therefore, nebracetam seems to produce a neuroprotective action by interacting, at least in part, with NMDA receptor-operated Ca2+ channels.


Isolation of substance P binding protein from rat brain. Substance P (SP) binding protein of rat brain was solubilized by digitonin. The solubilized proteins were then purified by sequential gel filtration, concanavalin A lectin Sepharose, and SP-affinity chromatography. The calculated molecular weight of this purified SP binding protein was 76-74 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The rabbits were immunized with the purified protein and resulting polyclonal anti-sera were tested. The immune serum significantly inhibited [3H]SP binding to the 3-[(3-cholamidopropyl)dimethylammonio]-1-propane sulfonate solubilized membrane fractions from rat brain, whereas pre-bleed antiserum failed to inhibit the binding. This polyclonal antibody also inhibited the activity of 45Ca influx into astroglioma cells stimulated by SP, but does not inhibit that stimulated by histamine. Furthermore, this polyclonal antibody recognized the 76-74 kDa band as assessed by Western blotting. These data strongly suggest that this polyclonal antibody could recognize a part of the natural SP receptor site.


Relaxant response of isolated basilar arteries to calcitonin gene-related peptide in stroke-prone spontaneously hypertensive rats. The relaxant effects of calcitonin gene-related peptide (CGRP) and other drugs were compared in basilar artery rings obtained from stroke-prone spontaneously hypertensive rats (SHRSP) and Wistar-Kyoto rats (WKY). In addition, the relaxant effect of CGRP on basilar arteries from spontaneously hypertensive rats (SHR) was examined. Relaxation induced by CGRP was independent of the presence of endothelium, and it was markedly increased in SHRSP when compared to WKY. In contrast, acetylcholine-induced relaxation was endothelium-dependent and did not differ between the two groups. Enhanced CGRP-induced relaxation was also found in SHR when compared to WKY. However, the relaxant response was greater in SHRSP than in SHR. No significant differences were found in the relaxation induced by isoproterenol, forskolin, dibutyryl cyclic AMP, and 3-isobutyl-1-methylxanthine in endothelium-rubbed arteries of WKY and SHRSP. These results suggest that CGRP produces endothelium-independent relaxation in the rat basilar artery, and that the enhanced CGRP-induced relaxation found in SHRSP may not be associated with alterations of vasodilation mediated by cyclic AMP.


[Granulocyte colony-stimulating factor producing lung large cell carcinoma with sarcomatous transformation]. We report a case of granulocyte colony-stimulating factor (G-CSF) producing lung large cell carcinoma with sarcomatous transformation. A 57-year-old man was admitted for evaluation of an abnormal shadow near the right pulmonary hilus on chest X-ray film. Brush cytology specimen from right B6 disclosed large cell carcinoma. His clinical stage was already IIIB (T3N3M0) on admission, so we immediately administered chemotherapy and radiation therapy. However, the patient's condition gradually deteriorated and he died of respiratory failure. Although there was no evidence of infection throughout his clinical course, his peripheral white blood cell count gradually increased and reached 47,000/mm3 (neutrophils 96%) before death. Histological study of the autopsy specimen revealed that the primary tumor was composed of two different elements (large cell carcinoma and spindle cell sarcomatoid element). The spindle cell sarcomatoid element appeared to have arisen from sarcomatous transformation of carcinoma cells, because the transition margin of the carcinoma to the sarcomatoid element was smooth, and special and immunohistochemical staining of both elements showed the same properties. Moreover, immunohistochemical study with monoclonal antibody to human G-CSF, 4A6 clearly demonstrated granular staining of G-CSF in the cytoplasm of tumor cells.


[Adjuvant chemotherapy with MVP-CAB (methotrexate, vincristine, cisplatinum, cyclophosphamide, adriamycin and bleomycin) for epithelial tumors of the upper urinary tract]. Surgery plus adjuvant chemotherapy using MVP-CAB (Day 1; methotrexate 20 mg/m2, vincristine 0.6 mg/m2, cyclophosphamide 500 mg/m2, adriamycin 20 mg/m2, and bleomycin 30 mg, Day 2; cisplatinum 50 mg/m2) was conducted in 12 patients with epithelial tumors of the upper urinary tract who had unfavorable prognostic factors (progressive disease which was pT2 or more, or transitional cell carcinoma of grade 2 and 3). The MVP-CAB regimen was as follows: A total of 3 cycles were given either before or after surgery. MVP-CAB was given at 3- to 4-week intervals before surgery, or after surgery if the patient had macroscopic residual lesions. For the patients with micrometastases detected after radical surgery, MVP-CAB was given every 1 to 2 months. The median survival period of the 10 patients who underwent radical surgery was 17 months (5-59 months). The three-year survival rate of these 10 patients (Kaplan-Meier method) was 100% in grade 2 (5 patients), 100% in progressive cancer greater than pT3 (6), and 80% in grade 3 (5). In two patients, residual macroscopic lesions after surgery were confirmed. One of them initially responded to MVP-CAB but died of cancer 21 months later, while the other one did not respond and died of cancer 8 months later. Two renal pelvis cancer patients for whom radical surgery was considered impossible due to distant metastases showed remarkable tumor reduction after MVP-CAB administration (one showed CR for liver metastases and the other showed PR for lymph node metastases).(ABSTRACT TRUNCATED AT 250 WORDS)


[Intra-arterial COMPA chemotherapy for invasive bladder cancer]. We presented 12 patients with invasive bladder cancer treated by the subselective intra-arterial COMPA chemotherapy. COMPA was administered up to an average of 3.3 courses (ranged from 2 to 6 courses) every 2 or 3 weeks, consisting of cisplatin: 15 mg/M2 on days 4 and 5; vincristine (oncovin): 0.6 mg/M2 on days 1 and 2; methotrexate: 5 mg/M2 on days 2 and 3; peplomycin: 5 mg/body on days 1, 2 and 3; and adriamycin: 10 mg/M2 on day 4. These were injected through a teflon catheter the tip of which was placed just proximal to the aortic bifurcation, and another tip was led through a subcutaneous tunnel from the inguinally punctured area to the anterior chest wall. The 12 patients, 7 men and 5 women, ranged in age from 53 to 73 (mean: 67) years. Histopathologically 11 had transitional cell carcinoma and one had adenocarcinoma. Malignant gradings were grade 2 in 8 patients, and grade 3 in 4. The stagings were T2 in 3 patients, T3 in 5, T4 in 4 and only one had bony metastasis. Of the 12 patients, 10 were alive at the last follow-up with a mean duration of 36 months (range: 16 to 49). Six patients achieved a complete remission, four achieved a partial remission and two were stable. One died of ileus after 16 months and another of progression after 36 months. All the patients received post-chemotherapeutic adjunctive therapies, which were transurethral resection, partial cystectomy, radiation and/or intravesical instillation. The toxicities were not severe, but anorexia, nausea, vomiting, hair loss, numbness of fingers and/or toes, subileus, and leukopenia were noticed.(ABSTRACT TRUNCATED AT 250 WORDS)


[A single session of transurethral microwave thermotherapy for benign prostatic hyperplasia]. A single session of transurethral microwave thermotherapy using Prostatron (Technomed International, France) was performed in 20 patients with benign prostatic hyperplasia, and the clinical efficacy of this modality was evaluated by analysing the subjective and objective responses following the treatment. 1,296 MHz microwaves are delivered to the prostate by an antenna placed in a 20 Fr urethral balloon catheter equipped with a cooling system for preservation of the urethral mucosa. The treatment was performed in a single session for an hour on an outpatient basis. The clinical efficacy was evaluated by a total score of subjective symptoms and objective findings including residual urine and average flow rate, 8 weeks after the treatment. Reduction of subjective symptom score was observed in 26 cases (86.7%), and that of objective findings in 21 (70%). When 25% or more decrease of the total score was defined as good, the overall good result including subjective and objective responses was seen in 24 cases (80%). During the treatment and follow-up period, no severe adverse effect was detected. In conclusion, a single session of thermotherapy by Prostatron is a safe and useful modality as a non-surgical treatment performed on an outpatient basis for benign prostatic hyperplasia.


[Significance of prostatic specific antigen in the mass screening for prostate cancer]. The significance of prostatic specific antigen (PSA) was investigated in the subjects examined by the mass screening for prostate cancer from 1985 to 1990. All subjects was examined by digital rectal examination (DRE) and with prostatic acid phosphatase (PAP) and the subjects in whom prostate cancer (Pca) was suspected from abnormal DRE and/or elevated PAP were recommended to receive the secondary screening to confirm the presence of Pca. PSA was measured by radioimmunoassay using Ball-Elsa-PSA-kit. 1,600 serum samples were obtained from our serum bank. The relationship among PSA, prostate size estimated by DRE and age was investigated. PSA was increased with age and the prostate size, PSA being more closely related with the latter. Therefore, we estimated that PSA has an ability to detect benign prostatic hypertrophy (BPH) in the mass screening. This estimation should be confirmed by using an ultrasound tomography because the prostate size obtained by DRE is inaccurate as compared with that obtained by ultrasound tomography. The cut off level of PSA was determined by control which was composed from the subjects with normal size prostate and one with BPH. When the cut off level was 8.6 ng/ml, the sensitivity, specificity and efficiency as Pca marker was 73.9%, 97.4% and 97.1%, respectively. PSA was more than 8.6 ng/ml in all of Pca with elevated PAP. PSA was expected to improve the Pca detection rate in our mass screening system.


[Three-dimensional display of the prostate based on transrectal ultrasonogram]. A software for a lap top computer to display prostatic contour three-dimensionally based on transrectal ultrasonograms was developed, and its clinical usefulness was examined. The prostatic contour of a case with prostatic cancer showed a typical irregular surface, and that of a case with prostatic hypertrophy had a smooth spherical shape, while that of a normal case had a flat shape. Each showed its characteristic shape. Estimation of the prostatic weight by assuming that the prostatic contour has ellipsoidal contour is a simple method for prostatic weight measurement, but it has a tendency to underestimate the weights, especially in normal cases. Three-dimensional display of the contour of prostatic cancer, prostatic hypertrophy, and normal prostate revealed that it could be estimated as ellipsoid in prostatic cancer or hypertrophy, while the normal prostatic contour was too flat to be calculated as such. Among 52 patients with benign prostatic hypertrophy who underwent transrectal ultrasonotomography before and during the anti-androgen therapy more than three time, nine cases showed transient reduction of more than 30% in prostatic weight and then reenlargement of more than 30%. In these cases, three-dimensional display of prostatic contour was done. The display was useful to visualize clearly which part reenlarged. Therefore, usefulness of the three-dimensional display of the prostate was verified.


[The surgical treatment of malignant liver tumors]. From study of the data concerning 133 patients who underwent operation for tumor of the liver the authors found that 43 (32.2%) patients were treated by a radical operation and 34 (25.5%) were subjected to palliative interventions. Among the radical interventions hemihepatectomy was performed on 21 (48.8%) patients. The total number of complications after radical operations was 46.3%, the mortality rate 11.6%. The authors discuss problems of the prevention of some complications, intraoperative hemorrhages in the first place. Different variants of bile diversion accounted for most of the palliative interventions. Survival in radical operations was 38.56 months, in palliative operations 8.34 months, and in exploratory operations 5.9 months.


[Reoperations in primary rectal cancer]. The authors studied the results of repeated radical operations performed in 73 patients who had been subjected earlier to exploratory and palliative operations. During repeated interventions radical operations were conducted on 54 (74%) patients which were of a combined character in 11 (20.4%) of them. Localized-radical palliative resections and extirpations in the presence of remote metastases were performed on 3 (4.1%) patients. Palliative operations were carried out on 16 (21.9%) patients. Postoperative mortality was 6.8%. The data provides evidence that the performance of repeated operations is expedient and justified because they make it possible to raise the resectability index among patients with rectal carcinoma.


Plasma lipoproteins and monocyte-macrophages in a peroxisome-deficient system: study of a patient with infantile refsum disease. Hypocholesterolaemia in infantile Refsum disease (IRD) may link peroxisomes and lipoprotein metabolism. In our patient, plasma cholesterol levels were reduced to 26% and 29% of control in LDL and HDL fractions, respectively. Plasma apolipoproteins B-100 and A-I levels were 52% and 66% of controls, respectively. In the kindred, plasma cholesterol concentration was 61-73% of controls. The HDL-cholesterol/apo A-I ratios were: patient 0.12; kindred 0.17; controls 0.28. Analysis of the IRD patient's lipoprotein revealed compositional abnormalities in all fractions. The patient's LDL demonstrated a substantial reduction in its lipid-to-protein ratio. Alterations in plasma lipoproteins affect their interaction with macrophages. Upon incubation of the patient's LDL with J-774 macrophages, its cellular uptake, measured as cholesterol esterification rate, was only 66% of a control rate. The abnormal LDL of the IRD patient showed also only 25% of control susceptibility to in vitro oxidation. Studies of cellular cholesterol metabolism in the patient's monocyte-derived macrophages (MDM) showed 57% increased cholesterol esterification rate in comparison to normal MDM. The possible link between lipoprotein abnormalities and monocyte-macrophage cholesterol metabolism is discussed.


[Laboratory diagnosis of tumors of the pancreato-biliary area in patients with mechanical jaundice]. Blood investigation of tumor markers CA-19-9, carcinoembryonic antigen and alpha-fetoprotein was performed in patients with mechanical jaundice due to tumor involvement of the pancreatobiliary area and a nontumor process. It was concluded that the use of CA-19-9-marker and carcinoembryonic antigen provided a successful differential diagnosis between jaundice resultant from tumor and nontumor diseases. Alpha-fetoprotein turned to be of no informative value. Analysis of tumor markers contents in patients with pancreatobiliary tumors demonstrated the usefulness of them in the preoperative assessment of surgical intervention practicability.


Expression of acidic fibroblast growth factor in regenerating liver and during hepatic differentiation. Acidic fibroblast growth factor belongs to a family of growth factors that show a high affinity for heparin sulfate proteoglycans. In vitro, it participates in various cellular functions including proliferation, differentiation, angiogenesis, and cell migration, but in vivo, the physiologic role of this growth factor is still not clearly defined. The level of expression and also cellular distribution of transcripts for acidic fibroblast growth factor (aFGF) were studied in adult rat liver after partial hepatectomy and during hepatic differentiation in fetal, neonatal, and adult livers by Northern analysis and in situ hybridization techniques. After partial hepatectomy a significant increase in the transcripts for aFGF was observed at 24 hours, whereas at 4 and 12 hours after the operation, the level of transcripts were similar to those of sham-operated animals. In the postnatal liver a high level of aFGF expression was present when the most evident transition from 2 to 3 cell thick hepatic cords to normal hepatic structure is taking place (Ogawa K, Medine A, Farber E. Br J Cancer 1979;40: 782-90). In contrast during the prenatal period, when the liver is still a hemopoietic organ and only a small number of sinusoids are present, low level of aFGF transcripts could be found. Animals treated with 2-acetylaminofluorene and partial hepatectomy (Evarts RP, Nagy P, Marsden E, Thorgeirsson SS. Carcinogenesis 1987;8:1737-40) displayed a marked increase in hepatic aFGF transcripts at the peak of proliferation of primitive liver epithelial cells (oval cells) and perisinusoidal stellate cells (Ito cells) in addition to hepatocytes. In situ hybridization combined with immunocytochemistry using oval and Ito cell specific antibodies revealed the presence of transcripts both in oval cells and Ito cells. Basophilic areas composed of small hepatocytes had a 3-fold increase in the level of transcripts as compared with the surrounding hepatocytes. These experiments demonstrate that the expression of aFGF is highest during the late stages of hepatic morphogenesis in newborn animals as well as during hepatic differentiation in adult liver.


Markers of complement-dependent and complement-independent glomerular visceral epithelial cell injury in vivo. Expression of antiadhesive proteins and cytoskeletal changes. Visceral glomerular epithelial cells (GEC) are an important component of the glomerular filtration barrier to proteins. While ultrastructural GEC changes have frequently been observed in proteinuric states, no suitable light microscopic markers of GEC injury have yet been identified. We have analyzed in vivo the GEC expression of proteins known to be involved in cell shape changes. SPARC (osteonectin, BM-40) and tenascin (cytotactin, J1, hexabrachion) belong to a group of anti-adhesive glycoproteins, that modulate cell-matrix interactions. We also studied cytoskeletal intermediate filament proteins, including desmin and vimentin. The GEC expression of SPARC, tenascin, desmin, and vimentin was analyzed in various types of GEC injury in the rat, including complement-mediated injury (passive Heymann nephritis, autologous immune complex nephritis, conA anti-conA nephritis), complement-independent injury (nephrotoxic nephritis), toxic injury (aminonucleoside nephrosis) and hypertensive injury (5/6 nephrectomy, angiotensin-II infusion). A complement-mediated model of mesangial cell injury (anti-Thy 1.1 mesangial proliferative nephritis) served as a control. SPARC mRNA and protein were constitutively expressed in normal rat glomeruli. Immunostaining and immunoelectron microscopy primarily localized SPARC to the cytoplasm of GEC. Markedly increased glomerular SPARC synthesis and GEC immunostaining was observed in all instances of complement-mediated GEC injury but in none of the other conditions. In contrast, glomerular immunostaining for tenascin, that also stained in a GEC pattern, either remained unchanged or increased to a minor degree (complement-mediated models). GEC immunostaining for desmin in normal rats was low and variable, and increased significantly in any form of GEC injury but not in anti-Thy 1.1 nephritis. No concomitant increase of GEC immunostaining for vimentin was detectable, which could have been due to the constitutively high expression of vimentin in GEC. SPARC and desmin, but not tenascin or vimentin, are suitable light microscopic markers of GEC injury. The combined staining for these proteins may be useful in differentiating the mechanisms of GEC injury.


A simple, quantitative method for assessing angiogenesis and antiangiogenic agents using reconstituted basement membrane, heparin, and fibroblast growth factor. Blood vessel growth is necessary for normal tissue homeostatis and contributes to solid tumor growth. Methods to quantitate neovascularization should be useful in testing biological factors and drugs that regulate angiogenesis or to induce a vascular supply to promote wound healing. An extract of basement membrane proteins (Matrigel) was found to reconstitute into a gel when injected subcutaneously into C57/BL mice and to support an intense vascular response when supplemented with angiogenic factors. New vessels and von Willebrand factor antigen staining were apparent in the gel 2-3 days after injection, reaching a maximum after 3-5 days. Hemoglobin content of the gels was found to parallel the increase in vessels in the gel allowing ready quantitation. Angiogenesis was obtained with both acidic and basic fibroblast growth factors and was enhanced by heparin. Several substances were tested for angiostatic activity in this assay by coinjection in Matrigel with fibroblast growth factor and heparin. Platelet-derived growth factor BB, interleukin 1-beta, interleukin-6, and transforming growth factor-beta were potent inhibitors of neovascularization induced by fibroblast growth factor. Tumor necrosis factor-alpha did not alter the response but was alone a potent inducer of neovascularization when coinjected with Matrigel and heparin. Consistent with the previously demonstrated importance of collagenase in mediating endothelial cell invasion, a tissue inhibitor of metalloproteinases that also inhibits collagenases was found to be a potent inhibitor of fibroblast growth factor-induced angiogenesis. Our assay allows the ready quantitative assessment of angiogenic and anti-angiogenic factors and should be useful in the isolation of endothelial cells from the capillaries that penetrate into the gel.


Pathways of macromolecular tracer transport across venules and small veins. Structural basis for the hyperpermeability of tumor blood vessels. Blood vessels supplying tumors are hyperpermeable to macromolecules, but the mechanisms responsible are poorly understood. To investigate the structural basis for the leakiness of tumor blood vessels, we performed a transmission electron microscopic study of three syngeneic transplantable carcinomas (mouse ovarian carcinoma and the line 1 and line 10 bile duct guinea pig carcinomas) at early intervals after intravenous injection of several macromolecular tracers. Tracers with widely differing physical properties were studied: horseradish peroxidase, ferritin, 150 kilodalton fluorescein isothiocyanate-dextran and gold-bovine serum albumin. All tracers leaked primarily from venules and small veins at the tumor-host interface, for the most part vessels lined by a continuous endothelium. The predominant pathway by which all four tracers exited venules in all three tumors was by way of a system of smooth membrane-bound, interconnecting vesicles and vacuoles; these tended to cluster together at irregular intervals in the endothelial cell cytoplasm to form organelle-like structures, vesiculo-vacuolar organelles (VVO). In favorable sections, VVO interfaced with both the luminal and abluminal surfaces of endothelial cells. HRP alone crossed venules and small veins through apposed inter-endothelial cell junctions. Tracers also exited vessels by way of endothelial fenestrae where these occurred (rarely) in mouse ovarian tumor-associated venules. VVO occurred with similar frequency and complexity in the continuous endothelium-lined venules and small veins that supplied the normal subcutis of either tumor-bearing or control animals. As in tumor-associated vessels, VVO provided the predominant pathway by which all four tracers exited normal vessels, but VVO labeling and extravasation were both much greater in tumor than in control vessels (p < 0.001 for ferritin). VVO are prominent structures in both tumor-supplying and control vessel endothelial cells and provide the primary pathway for macromolecular extravasation. The large increase in permeability characteristic of tumor vessels is likely attributable to upregulation of VVO function.


Loss of expression of protectin (CD59) is associated with complement membrane attack complex deposition in myocardial infarction. Protectin (CD59) is a recently discovered inhibitor of the complement membrane attack complex (MAC). In the present study we investigated expression of protectin in human heart and examined the relationship between MAC deposition and protectin in myocardial infarction. Myocardial tissue specimens were obtained at autopsy from patients who had died of myocardial infarction (n = 10) or other causes (n = 5). MAC and protectin were detected by indirect immunofluorescence microscopy analysis in the heart sections by using antibodies against individual components of MAC, MAC neoantigens and protectin. Myocardial protectin was purified by affinity chromatography and compared with the previously characterized erythrocyte and urinary protectins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis, N-terminal amino acid sequencing, and testing its ability to bind to the terminal complement complex. The possible glycophosphoinositol-type anchorage of protectin in the heart was examined by treating myocardial sections with glycophosphoinositol-specific phospholipase C. Immunoblotting and immunofluorescence analysis showed expression of protectin in the sarcolemmal membranes of normal myocardium. Protectin purified from normal human heart tissue had the same molecular weight and N-terminal amino acid sequence as CD59 purified from urine. In sucrose density ultracentrifugation analysis it was observed to bind efficiently to the SC5b-8 complex. In normal myocardium the expression of CD59 was sensitive to treatment with glycophosphoinositol-specific phospholipase C. The expression of CD59 was lost or clearly diminished in infarcted lesions aged 1-14 days. Loss of CD59 expression was accompanied by concomitant deposition of the MAC within the CD59-negative lesions. In border areas between an infarcted lesion and normal tissue, CD59 often appeared in small vesicles, suggesting shedding as a possible mechanism for its removal. Glycophosphoinositol-anchored CD59 is expressed in the sarcolemmal membranes of normal heart but lost from infarcted myocardium. Acquired loss of resistance to autologous complement and subsequent complement attack may thus be involved in the pathophysiology of myocardial infarction.


Effects of training on muscle capillary pattern: intermittent vs continuous exercise. The effects of two exercise regimens (intermittent and continuous endurance) on the fiber area, and capillarization of mice skeletal muscle (gastrocnemius) was studied. The training programs had a duration of 30 days, 5 days/week, and was performed on a motor-driven treadmill. The intermittent group performed the exercise at 16 m.min-1 with a slope of 5%. The continuous group performed the exercise at 16 m.min-1 with 0%. The fiber area and the capillarization was evaluated in the white portion of the gastrocnemius. A high degree of hypertrophy was observed, and a significant increase in the capillary density, capillary to fiber ratio, and in the number of capillaries sectioned obliquely and/or longitudinally was found. The results suggest that the endurance training, intermittent or continuous, induce cell hypertrophy, and an increase in capillarization especially, based on an alteration of the structural pattern.


Substance P increases in vitro lymphokine-activated-killer (LAK) cell cytotoxicity against fresh colorectal cancer cells. Gut-associated lymphoid cells are modulated by several gut hormones. We postulated that lymphokine-associated-killer (LAK) cell cytotoxicity of lymphocytes isolated from the gut mucosa may be increased by substance P (SP). Intestinal lamina propria mononuclear cells (LPMC) and colonic cancer cells were isolated from operative specimens by successive mechanical and enzymatic dissociation methods. Effector LAK cells were induced by culturing LPMC with recombinant interleukin-2 at a concentration of 250 U/ml. Substance P (10(-5) M) was added to the culture medium. Targets consisted of fresh colon cancer cells, HT-29 (cultured human colon cancer cell line), and control cell lines. After 4 days of incubation, cytotoxicity was measured using a 4-h 51Cr release assay. LAK cells alone showed moderate cytotoxicity against HT-29 and none against fresh colon cancer cells. LAK cells generated in the presence of substance P showed moderate cytotoxicity against HT-29 and strong cytotoxicity against fresh colorectal cancer cells. The percentage of cytotoxicity +/- SEM at various effector to target ratios was [(*) denotes P < 0.05 compared with above]: [table: see text] We conclude that substance P significantly increases LAK cell cytotoxicity against fresh colon cancer cells, but not against cultured cells.


Protease activation following reperfusion of porcine pancreatic allografts. To study the degree of protease activation at reperfusion of a pancreatic allograft after cold storage for 24 hr, 18 porcine whole-organ pancreaticoduodenal allograft transplantations were performed. Twelve grafts were flushed with and stored in Perfadex. In six of these, a hyperosmotic salt solution was injected into the graft aorta at reperfusion. Six grafts were flushed and stored in UW solution. Eleven of twelve grafts in the Perfadex groups were functioning on the first postoperative day, compared with one of six in the UW solution group. There was a significantly more pronounced protease activation among grafts stored in UW solution than in the other groups, with a subsequent breakthrough of the local protease protection barrier made up of protease inhibitors. In surviving pigs (n = 14), biochemical signs of protease activation evolved in plasma, including formation of trypsin-protease inhibitor complexes, a decline in C3 and kininogen levels, and a decline in functionally active alpha 2-macroglobulin, functionally active antithrombin III, and plasma kallikrein inhibitory activity. These biochemical signs of pancreatitis correlated with a deteriorated graft function on the second postoperative day, indicating that graft tissue damage occurred due to protease activation.


Administration of tumor necrosis factor alpha (TNF alpha) inhibitors after exposure to TNF alpha prevents development of the maximal biological effect: an argument for clinical treatment with TNF alpha inhibitors. The cytokine tumor necrosis factor alpha (TNF alpha) is involved in the pathophysiology of a wide variety of diseases. Pretreatment with anti-TNF alpha antibodies has proven its success in animal models of disease. The question, however, whether intervention with anti-TNF alpha antibodies might be useful in the clinical situation in which TNF alpha is already produced is still unanswered. We therefore studied the relation between the duration of TNF alpha/TNF receptor interaction and the extent of the induced biological effects in two different in vitro systems: (1) the slowly induced cytotoxicity of the TNF-sensitive murine cell line L929, and (2) the rapid TNF alpha-induced expression of an adhesion molecule for the polymorphonuclear cell, ELAM-1 on human endothelial cells. The TNF alpha/TNF receptor interaction was interrupted at different times after onset of stimulation, either by washing away TNF alpha or by adding TNF alpha activity-blocking monoclonal antibody. To establish an optimal effect for both TNF alpha-induced cytotoxicity on L929 cells and TNF alpha-induced expression of ELAM-1 on human endothelial cells, the TNF receptor had to be occupied by TNF alpha for at least 30-60% of the full incubation period. This observation provides an argument that clinical intervention with TNF alpha inhibitors can be advantageous, even in a situation in which TNF alpha has already been released into the circulation.


In vitro and ex vivo inhibition of the modification of low-density lipoprotein by indapamide. The effect of an antihypertensive drug, indapamide, on copper- and endothelial cell-induced peroxidation of human low-density lipoprotein (LDL) was studied and compared with that of drugs previously shown to protect LDL against peroxidation: probucol and vitamin E and other thiazidic and nonthiazidic diuretics (clopamide, hydrochlorothiazide, and furosemide). Incubation with indapamide inhibited in a dose-dependent manner LDL peroxidation induced either by copper ions or by cultured endothelial cells. Both electrophoretic mobility and thiobarbituric acid-reactive substances (TBARS) content of LDL returned to almost normal values in the presence of 1 microM indapamide. This drug was at least 10 times more potent than probucol and vitamin E in inhibiting LDL peroxidation. No inhibitory effect has been observed with clopamide, hydrochlorothiazide, and furosemide in the same experimental conditions. Homozygote Watanabe rabbits were treated orally with indapamide (10 mg/kg/d for 3 days) to evaluate the potential protective effect of the compound on LDL peroxidation in vivo. Purified LDL from placebo and treated rabbits were submitted to peroxidation induced by copper ions, and indapamide was effectively able to protect LDL in these experimental conditions. This effect was especially obvious 6 and 8 h after the start of the incubation when LDL of the placebo-treated animals were modified. The mechanism of action of these drugs was examined in vitro using the 1,1-diphenyl-2-picryl-hydrazyl (DPPH) test and in kinetic studies of arachidonic acid photoperoxidation. Indapamide as well as vitamin E and probucol were effective free radical scavengers, but the other diuretic molecules were not.(ABSTRACT TRUNCATED AT 250 WORDS)


Pharmacokinetics and pharmacodynamics of benazepril in hypertensive patients with normal and impaired renal function. The pharmacokinetics and pharmacodynamics of benazepril, an angiotensin converting enzyme (ACE) inhibitor, were investigated after administration of a single oral 5-mg dose and 7 more doses on consecutive days to hypertensive patients with normal renal function (NRF) and those with impaired renal function (IRF). The antihypertensive effect of benazepril was observed as early as 30 min after a single dose, and those effects during consecutive dosing were also sustained for 24 h with a lesser diurnal variation in blood pressure (BP). The time to peak (Tmax) and the apparent elimination half-life (t1/2) for benazepril were 0.6-0.7 h and 0.4-0.8 h, respectively. Tmax for its diacid was 1.5-2.4 h in both groups. The area under the plasma concentration-time curve to 24 h (AUC0-24h) for the diacid was significantly greater in the IRF group than in the NRF group. After consecutive dosing of benazepril, AUC0-24h and plasma peak level (Cmax) were significantly increased in the IRF group. Serum ACE activity was markedly suppressed for 24 h after administration, and the inhibition was closely related to plasma diacid levels. A significant inverse correlation was observed between creatinine clearance and the AUC for the diacid. These results suggest that benazepril is rapidly bioactivated to diacid and exhibits rapid onset and long-lasting antihypertensive effects. Dosage reduction might be required to minimize unnecessary drug accumulation in patients with severe IRF.


Noninvasive estimates of the inodilatory effects of isoprenaline and their inhibition by transdermally delivered mepindolol in healthy men. We analyzed the reproducibility and sensitivity of noninvasive estimates of cardiovascular performance (impedance cardiography and systolic time intervals) to detect and describe the inodilatory responses to isoprenaline (ISO) and their attenuation by transdermally applied mepindolol. The responses to 5-min intravenous (i.v.) infusions of 1 microgram/min ISO in 9 young male healthy volunteers were evaluated repeatedly before and at the end of chronic treatments with daily applied TSD skin patches for transdermal drug delivery with either mepindolol or placebo (double-blind randomized cross-over design). ISO caused a clear increase in mean heart rate (HR), systolic blood pressure (SBP), maximum velocity of impedance changes, estimated stroke volume (SV), and the Heather index, and a reduction in diastolic BP (DBP), estimated systemic vascular resistance (SVR), and a shortening of all systolic time intervals (STIs). These responses were effectively blunted at the end of 1-week treatment with mepindolol TSD patches. Despite overall good agreement of the mean baseline data (under placebo), their within-subject reproducibility nevertheless was too weak to guarantee adequate power to detect small changes under active treatment. Reproducibility of the ISO responses was quite poor, although in a method-independent way. The methods used allowed us to describe the cardiac and vascular components of the ISO responses with adequate detail. Transdermally delivered mepindolol was shown to block ISO responses effectively, but the overall variability of the responses was too large to detect slight changes in them, although the methods in themselves did not appear to be the main cause of this lack of sensitivity.


Contrasting cardiovascular responses from intrathecal administration of epinephrine and norepinephrine in conscious rats: role of alpha 1- and alpha 2-adrenoceptors. In conscious rats, intrathecal (i.t.) administration of norepinephrine (NE) produced pressor responses, whereas i.t. epinephrine (Epi) caused depressor responses at low doses (0.1-1 microgram) and pressor responses at a higher dose (10 micrograms). Epi administered i.t. produced bradycardia; however, NE caused tachycardia at low doses and bradycardia at high doses. The cardiovascular responses were dissimilar to those observed after intravenous (i.v.) administration of these doses of NE and Epi. When [3H]NE or [3H]Epi (1.0 microgram, 10 mCi) was injected i.t., minimal radioactivity was detected in peripheral blood (PB) samples, indicating that the effects of i.t.-injected catecholamines on blood pressure (BP) and heart rate (HR) are due to stimulation of central spinal adrenoceptors and not to peripheral effects after leakage. Pretreatment with i.t. administration of the alpha 1-antagonist prazosin (1.0 microgram) attenuated pressor responses and tachycardia produced by i.t. NE (1.0 microgram), whereas i.t. pretreatment with the alpha 2-antagonist yohimbine (10 micrograms) counteracted depressor responses and bradycardia produced by i.t. Epi. Therefore, these spinally released catecholamines appear to produce opposite cardiovascular effects whereby sympathetic preganglionic neurons are excited by NE through spinal alpha 1-adrenoceptors and are inhibited by Epi through spinal alpha 2-adrenoceptors.


Electrophysiologic effects and efficacy of cibenzoline in patients with supraventricular tachycardia. Electrophysiologic effects of intravenous (i.v.) cibenzoline were evaluated in 18 patients with accessory pathways or dual atrioventricular (AV) nodal pathways (12 men and 6 women with a mean age of 44 +/- 18 years). Twelve patients had accessory AV pathways, including 6 patients with a manifest accessory pathway. Six patients had AV nodal reentrant tachycardia (AVNRT). Electrophysiologic studies were performed before and after cibenzoline (1.4 mg/kg i.v.) infusion for 5 min. Sinus cycle length did not change significantly after cibenzoline administration. Cibenzoline increased both the AH (85 +/- 20 vs. 91 +/- 21 ms, p less than 0.05) and HV intervals (41 +/- 10 ms vs. 53 +/- 11 ms, p less than 0.001). Neither the atrial nor ventricular effective refractory period (ERP) was altered by cibenzoline. Complete block in the accessory pathway occurred antegradely in 4 patients and retrogradely in 1 patient. Cibenzoline prevented induction of AV reentrant tachycardia (AVRT) in 3 of 8 patients with sustained orthodromic AVRT by abolishing retrograde accessory pathway conduction or prolonging the retrograde accessory pathway ERP. Of 5 patients who continued to have inducible AVRT before and after cibenzoline administration, the tachycardia cycle length was increased in 3, mainly due to the increase in retrograde accessory pathway conduction time. Cibenzoline prevented induction of sustained AVNRT in 4 of 5 patients by prolonging the minimum pacing cycle length, maintaining 1:1 ventriculoatrial (VA) conduction through the retrograde fast AVN pathway or shortening the antegrade fast AVN pathway ERP equal to the slow AVN pathway. In one patient who had an uncommon type of AVNRT, sustained tachycardia was induced by cibenzoline.(ABSTRACT TRUNCATED AT 250 WORDS)


Calcium antagonistic and binding properties of semotiadil (SD-3211), a benzothiazine derivative assessed in cerebral and coronary arteries. The present experiments were undertaken to elucidate Ca2+ antagonistic and binding properties of semotiadil and its (S)-(-)-enantiomer (SD-3212) in plausible clinical target tissues such as cerebral and coronary arteries. Semotiadil was about six times more potent than D-cis-diltiazem for Ca2+ antagonistic action, with a long-lasting and wide spectrum of inhibitory effects on contraction of dog cerebral arteries elicited by various spasmogens and mechanical stretch. Semotiadil exhibited a weak, negative, and heterotropic allosteric effect on (+)-[3H]PN 200-110 binding to pig coronary artery membranes: Scatchard analysis of saturation isotherms indicated that semotiadil increased the equilibrium dissociation constant (Kd) of (+)-[3H]PN-200-110 binding without causing a significant change in the maximum binding density (Bmax). Furthermore, semotiadil significantly increased the dissociation rate (k-1) of (+)-[3H]PN 200-110 from the specific binding site. The enhanced binding of (+)-[3H]PN 200-110 to the coronary artery caused by D-cis-diltiazem was attenuated when semotiadil was present, whereas binding inhibited by verapamil was not affected in the presence of semotiadil. The results suggest that semotiadil exerts a potent Ca2+ antagonistic action by binding to a site in the Ca2+ channel distinct from the 1,4-dihydropyridine recognition site and interacts with the 1,4-dihydropyridine binding site in a negative, heterotropic, allosteric manner.


Release of multiple endothelium-derived relaxing factors from porcine coronary arteries. Using a chemiluminescence method in the present study, we measured nitric oxide and one-electron oxidation products of nitric oxide (NOX) released from porcine coronary artery segments in response to bradykinin, ADP, and the calcium ionophore A23187. Total NOX was compared with the bioactivity of endothelium-derived relaxing factors (EDRF) by a biodetector ring preparation before and after inhibition of L-arginine-dependent nitric oxide synthesis and in the presence of indomethacin. Under basal conditions, arterial segments released NOX and relaxed biodetector rings. Bradykinin, ADP, and A23187 elicited vasorelaxation greater than that observed basally; A23187, but not bradykinin or ADP, caused additional release of NOX greater than that measured basally. Hemoglobin completely reversed vasorelaxation elicited by all three agonists. We compared the amount of nitric oxide released under basal conditions and after stimulation with bradykinin, ADP, and A23187 with the amount of authentic nitric oxide necessary to elicit a bioequivalent response. Authentic nitric oxide did not account for the observed bioactivity as compared with the amount of nitric oxide actually measured in arterial segment effluent. To investigate whether a second non-nitric oxide-containing compound was responsible for the increased bioactivity and the discrepancy between the bioactivity and quantity of nitric oxide measured, we exposed arterial segments to omega-nitro-L-arginine methyl ester to inhibit L-arginine-dependent synthesis of nitroso compounds. The drug completely abolished the nitric oxide signal derived from both basally released and A23187-stimulated relaxing factor and completely reversed vasorelaxation. In contrast, omega-nitro-L-arginine methyl ester only partially reversed bradykinin-stimulated vasorelaxation.(ABSTRACT TRUNCATED AT 250 WORDS)


Role of lipoxygenase and cytochrome P-450 in production of endothelium-derived relaxing factors in canine femoral veins. We wished to determine whether the metabolism of arachidonic acid, through lipoxygenase and cytochrome P-450 pathways, is involved in production of endothelium-derived relaxing factor(s) (EDRFs) in canine femoral veins. Veins were removed from anesthetized dogs and cut into rings. Endothelium was deliberately removed from some rings. In separate sets of experiments, rings were incubated with either AA861 (10(-5) M) or TMK777 (10(-6) M), inhibitors of 5-lipoxygenase, nordihydroguaiaretic acid (NDGA 3 x 10(-6) M), an inhibitor of lipoxygenase or proadifen (SKF 525A, 10(-6) M), an inhibitor of cytochrome P-450. In addition, some rings were incubated with a combination of indomethacin (10(-5) M) and NG-monomethyl-L-arginine (L-NMMA 10(-4) M) or, where appropriate, a solvent control. Concentration-response curves were obtained for acetylcholine, adenosine diphosphate, thrombin, A23187, and nitric oxide in rings contracted with a submaximal concentration of prostaglandin F2 alpha. AA861 and TMK777 did not alter endothelium-dependent relaxations to the agonists, whether with or without indomethacin and L-NMMA. However, indomethacin plus L-NMMA reduced endothelium-dependent relaxations to thrombin. These results suggest that metabolism of arachidonic acid, through lipoxygenase and cytochrome P-450 pathways, does not produce an EDRF in veins. However, thrombin receptor-activated relaxations are mediated in part by products of the cyclooxygenase pathway and nitric oxide.


OPC-13340, a new dihydropyridine calcium channel blocker attenuates rapid vascular smooth muscle cell growth in spontaneously hypertensive rats. We investigated the mechanism of the antimitotic effects of calcium channel blockers in vascular smooth muscle cells (VSMC) from spontaneously hypertensive rats (SHR). VSMC from SHR exhibited rapid proliferation through a quick transition from the G0/G1 to the DNA synthetic (S) phase and from the S to the G2/mitotic (M) phase, whereas the DNA synthetic rate itself was equal to that of Wistar-Kyoto rats (WKY). OPC-13340, a new dihydropyridine calcium channel blocker, dose-dependently decreased incorporation of [3H]thymidine into the DNA fragments in randomly cycling VSMC in SHR. Cell cycle analysis showed that the rapid transition from the S to the G2/M period was restored by OPC-13340 to the control level in WKY, whereas the quick transition from G0/G1 to S was unaffected. This antimitotic effect of OPC-13340 was reflected by attenuation of enhanced cellular protein synthesis during the G2/M period. Protein synthesis in the G0/G1 period was not influenced by OPC-13340. Thus, these data indicate that the calcium channel blocker OPC-13340 mitigates the enhanced proliferation observed in randomly cycling VSMC from SHR and that this effect is primarily due to normalization of the premature mitosis during the G2/M period.


Effect of amlodipine pretreatment on ischaemia-reperfusion-induced increase in cardiac endothelin-1 binding site density. Endothelin-1 (ET-1) may be implicated in the pathophysiology of myocardial ischaemia. To determine whether the long-acting calcium antagonist amlodipine attenuates the ischaemia- and reperfusion-induced increase in cardiac ET-1 binding sites, hearts from rats pretreated with amlodipine (0.25 or 0.5 mg/kg) 2 or 5 h before they were killed were made ischaemic for 20 or 40 min, reperfused, and subfractionated. Twenty- and 40-min ischaemia caused a time-dependent increase in ET-1 binding site density (Bmax) identified with [125I]ET-1. Amlodipine pretreatment attenuated this increase in a time- and dose-dependent manner. 0.25 and 0.5 mg/kg amlodipine also suppressed the reperfusion-induced increase in [125I]ET-1 binding site density, even when the 0.5-mg/kg pretreatment series reperfusion was administered after 40-min ischaemia.


Effects of R 56865 and phenytoin on mechanical, biochemical, and morphologic changes during ouabain intoxication in isolated perfused rabbit heart. The Na+/Ca2+ overload inhibitor R 56865 (N-[1-[4-(4-fluorophenoxy)-butyl]-4-piperidinyl)-N-methyl-2- benzothiazolamine) has been reported to prevent or attenuate ischemia- as well as ouabain-induced cellular sodium and calcium load. We investigated the potency of this compound in preventing mechanical, biochemical, and ultrastructural consequences of ouabain (OUA) intoxication in isolated rabbit heart. The protective effect of the digitalis antidote phenytoin (PHT) on the consequences of ouabain intoxication was examined for comparison. In isolated perfused rabbit heart, OUA (0.4 microM) caused an increase in left ventricular end-diastolic pressure (LVEDP) that was accompanied by depletion of high-energy phosphates (80% less than in control), accumulation of tissue lactate (12-fold) and damage of contractile elements and mitochondria. Accumulation of lactate was associated with a decrease in oxygen consumption by the isolated perfused heart. R 56865 (1.0 microM) and phenytoin (60 microM) prevented increase in LVEDP, breakdown of the energy-rich phosphates creatine phosphate (CrP) and ATP, accumulation of lactate, and morphologic changes induced by OUA. The above-mentioned toxic effects of OUA are interpreted as consequences of mitochondrial failure finally leading to breakdown of the oxidative phosphorylation. Thus, we conclude that the protective action of both compounds, R56865 and PHT, may be attributed to prevention or attenuation of mitochondrial failure due to OUA-induced disturbance of ion homeostasis.


Placebo-controlled trial of doxazosin in management of patients with hypertension and hypercholesterolaemia. The blood pressure (BP)- and lipid-lowering activities of the alpha 1-antagonist, doxazosin, were investigated in hypertensive, hypercholesterolaemic patients. Thirty-one patients satisfactorily completed the study, and there was no significant difference between doxazosin and placebo in terms of reported adverse events. After 3-month treatment, BP was significantly reduced by doxazosin by 24/14 mm Hg supine and by 33/22 mm Hg erect as compared with corresponding reductions of 2/9 and 2/2 mm Hg with placebo. There were concomitant improvements in the plasma lipid profile with, in particular, significant net reductions of 30% for triglycerides and 20% for apoprotein B. There was no adverse effect on glucose metabolism. The principal aim of this study was assessment of the clinical utility and acceptability of doxazosin in a heterogeneous population of patients with several cardiovascular risk factors. The results confirm that doxazosin is an effective antihypertensive agent that has modest additional beneficial effects on the plasma lipid profile.


Terbutaline-induced desensitization of beta 2-adrenoceptor in vivo function in humans: attenuation by ketotifen. Use of beta-adrenoceptor agonists in long-term treatment of patients with chronic asthma bronchiale or heart failure is of limited value because beta-adrenoceptor desensitization develops. The antiallergic drug ketotifen prevents beta-adrenoceptor agonist-induced desensitization of rat and human pulmonary and lymphocyte beta 2-adrenoceptors. In 10 healthy volunteers in a double-blind, placebo-controlled study, we investigated whether ketotifen also prevents beta-adrenoceptor agonist-induced desensitization of beta 1- and/or beta 2-adrenoceptor-mediated physiologic in vivo effects. beta 1-Adrenoceptor-mediated effects were isoprenaline (ISO) infusion-induced increase in systolic blood pressure (SBP) and bicycle exercise-induced increase in heart rate (HR); beta 2-adrenoceptor-mediated effects were ISO infusion-induced increase in plasma norepinephrine (NE) and decrease in diastolic blood pressure (DBP); ISO infusion-induced increase in HR was assessed as mixed beta 1- and beta 2-adrenoceptor-mediated effect. These parameters were assessed before and after a 14-day treatment with the beta 2-adrenoceptor agonist terbutaline (5 mg three times daily) with or without simultaneous administration of ketotifen (1 mg twice daily). Terbutaline desensitized all in vivo effects involving beta 2-adrenoceptors (ISO-induced decrease in DBP and increase in plasma NE and, to a minor extent, the mixed beta 1- and beta 2-adrenoceptor-mediated increase in HR), but did not affect beta 1-adrenoceptor-mediated in vivo effects; concomitant treatment of the volunteers with ketotifen markedly blunted terbutaline-induced desensitization of beta 2-adrenoceptor in vivo function. We conclude that ketotifen prevents, or at least attenuates, beta-adrenoceptor agonist-induced desensitization of beta 2-adrenoceptor in vivo function.(ABSTRACT TRUNCATED AT 250 WORDS)


Alpha-adrenoceptor blockade in patients with mild to moderate hypertension: long-term renal effects of doxazosin. Using constant infusion technique and a water-loading procedure, we investigated renal hemodynamic and excretional variables in 15 essential hypertensive patients [diastolic blood pressure (DBP) 102 +/- 10 mm Hg] after 3 weeks of placebo and after 16 weeks of treatment with a postjunctional alpha 1-adrenoceptor-antagonist, doxazosin (1-16 mg) once daily. A minor decrease in supine DBP (p less than 0.05) but no significant changes in systolic BP (SBP) and heart rate (HR) were observed. No significant changes were noted in glomerular filtration rate (GFR), renal plasma flow (RPF), and renal vascular resistance (RVR). The mean renal excretion rate of sodium, potassium, uric acid, and albumin for the entire group was unaffected by the treatment, but the individual changes in sodium clearance correlated significantly with changes in mean BP (r = 0.64, n = 15, p less than 0.05). Six patients showed an increase in sodium excretion after treatment, whereas nine showed a decrease. No decrease in mean body weight was noted, but the BP reduction after 5 months of treatment correlated significantly with the changes in body weight (r = 0.62, n = 15, p less than 0.01). The results indicate that long-term treatment with doxazosin had no deleterious effect on renal function, but the effects on BP were rather modest. The individual BP response is probably determined by the degree of fluid retention even if an intact pressure-natriuresis relationship could still be demonstrated during chronic therapy.


Mechanism of halothane attenuation of isometric tension induced by serotonin in isolated canine coronary artery rings. We explored the mechanism of halothane's interaction with the serotoninergic contractile response of isolated canine coronary artery rings. The serotoninergic contractile response of both intact and denuded rings was measured with and without halothane. In some experiments, rings were pretreated with methiothepin, a 5-HT1 and 5-HT2 antagonist, or ketanserin, a 5-HT2 antagonist. The contractile responses to 5-carboxamidotryptamine (5-CT) and alpha-methylserotonin, a 5-HT1 and a 5-HT2 receptor agonist, respectively, were measured with and without halothane. Finally, the response to prostaglandin F2-alpha, another spasm mediator, was also measured with and without halothane. Halothane attenuated the coronary artery response to serotonin (5-hydroxytryptamine, 5-HT), and specific 5-HT1 and 5-HT2 agonists, and prostaglandin F2 alpha (PGF2 alpha). Its inhibitory effect on the serotoninergic response was abolished in vessels pretreated with either 5-HT1 or 5-HT2 blockers. These data suggest that halothane is not a direct smooth muscle depressant, that it is not a specific 5-HT1- or 5-HT2-subtype antagonist in canine coronary arteries, and that it might interfere with intracellular pathways activated by agonist-receptor interactions.


Steady-state pharmacokinetics and pharmacodynamics of cilazapril in the presence and absence of cyclopenthiazide. Twenty-two patients with essential hypertension received a single dose of 2.5 mg cilazapril and were then randomised into a double-blind parallel group study to receive either placebo, 1.25 mg cilazapril + 0.5 mg cyclopenthiazide (CPTZ), 2.5 mg cilazapril + 0.5 mg CPTZ, or 2.5 mg cilazapril alone for 1 month. After oral administration of a single dose of 2.5 mg cilazapril, the active diacid cilazaprilat appeared rapidly in the plasma (Tmax 2.0 +/- 0.2 h). With the radioinhibitor assay used in this study, a single elimination phase of cilazaprilat was evident, with a half-life (t1/2) of 2-3 h. At steady state, the pharmacokinetics of cilazaprilat were similar to single-dose administration and were not altered by CPTZ. The Cmax and area under the curve (AUC) of cilazaprilat were directly proportional to dose. Cilazapril administration in the dose range of 1.25-2.5 mg produced a dose-proportional inhibition of angiotensin-converting enzyme (ACE) activity that was maximum 2 h after drug administration. The degree of ACE inhibition correlated with the plasma concentration-time profile of cilazaprilat and the maximum decrease in blood pressure (BP). The EC50 for ACE inhibition by cilazaprilat was 7.7 ng/ml after acute treatment and was not significantly altered during chronic administration or by concomitant administration of CPTZ. There was no evidence of a dose-related antihypertensive effect of cilazapril at steady state and, with the small numbers of subjects used in this study, there was no evidence of 24-h BP control with monotherapy.


Electrophysiologic and hemodynamic effects of H 234/09 (almokalant), quinidine, and (+)-sotalol in the anesthetized dog. The electrophysiologic and hemodynamic effects of H 234/09 (Almokalant), a novel class II antiarrhythmic agent, were studied in the anesthetized dog. H 234/09 (1.0 mumol/kg i.v.) significantly prolonged the atrial and ventricular effective refractory periods, the ventricular monophasic action potential duration, and the paced QT interval. At this dose, atrial, ventricular, and atrioventricular conduction was not affected, aortic blood pressure was not changed, and contractile force was transiently increased. The effects on cardiac repolarization and refractoriness induced by H 234/09 were both larger and more long lasting than the effects observed after quinidine (11.8 mumol/kg) and (+)-sotalol (9.7 mumol/kg). However, both quinidine and (+)-sotalol significantly reduced the aortic blood pressure and (+)-sotalol also decreased cardiac contractility. The effect of H 234/09 on atrial refractoriness was very little influenced by the paced heart rate and was twice as large as the corresponding effect in the ventricle. In conclusion, H 234/09 has electrophysiological properties suggestive of a class III antiarrhythmic. H 234/09 may have a favorable therapeutic profile compared to both quinidine and (+)-sotalol, especially for the treatment of atrial arrhythmias.


Neuropeptide Y inhibits relaxation of guinea pig cerebral, coronary, and uterine arteries: blockade by D-myo-inositol-1,2,6-triphosphate. Interactions between neuropeptide Y and perivascular vasodilator agents were studied in guinea pig cerebral, coronary, and uterine arteries. In all three types of arteries, vessel segments precontracted with prostaglandin F2 alpha or histamine relaxed concentration dependently upon application of acetylcholine (ACh), substance P (SP), and vasoactive intestinal peptide (VIP). Neuropeptide Y (NPY: 10(-8)-10(-7) M) caused inhibition of relaxations produced by ACh, SP, and VIP in all three types of segments; however, the effective concentration varied between vessel type. Thus, cerebral and uterine arteries were approximately 10 times more sensitive to NPY than the coronary artery. D-myo-inositol-1,2,6-triphosphate (PP56) was a potent inhibitor of the NPY effect in all three vessel types. Thus, NPY, which is colocalized not only with norepinephrine in sympathetic perivascular fibers but also with VIP and ACh in some parasympathetic neurons, can greatly reduce the vasodilatory effect of ACh and VIP, as well as of the sensory peptide SP. This further illustrates the complex interactions NPY has with perivascular neuroeffector mechanisms.


Dose-response curve of angiographically smooth human epicardial vessel segments to intracoronary injections of isosorbide dinitrate. The coronary vasodilator properties of isosorbide dinitrate are well established but the doses generally used (1,000-2,000 micrograms) are still empirical. We studied, with the use of quantitative coronary arteriography (CAESAR System), the response of smooth vessel segments (greater than 1.85 mm diameter), preconstricted with methylergometrine (400 micrograms i.v.), to intracoronary injections of graded doses (5-100 micrograms) of isosorbide dinitrate and the effects of these injections on systemic hemodynamic parameters in 10 patients undergoing diagnostic coronary angiography. Six further patients, in whom the injections of isosorbide dinitrate were replaced by equivalent volumes of normal saline, served as controls. Relative to the diameter 5 min after injection of methylergometrine, the diameter increased by a mean +/- SD of 9 +/- 7, 26 +/- 12, 33 +/- 15, 38 +/- 14, and 39 +/- 16% after injections of 5, 15, 60, 240, and 1,000 micrograms, respectively, of isosorbide dinitrate. After a cumulative dose of 80 micrograms, subsequent doses did not cause further significant increases in diameter. Injection of saline in the control group did not alter the coronary diameter. A significant fall in systolic arterial pressure, compared to the control group, occurred at a cumulative dose of 320 micrograms. The mean arterial pressure and heart rate were unchanged. Significant coronary vasodilation occurs with intracoronary doses of isosorbide much smaller than those currently employed. Cumulative doses of 320 micrograms or more cause systemic hemodynamic changes without producing additional coronary vasodilation. During interventional cardiac procedures, where systemic hypotension is undesirable, the use of smaller doses of intracoronary isosorbide dinitrate than currently employed may be feasible and should be investigated further.


Positive inotropic effects due to partial agonistic activity of the beta-adrenoceptor antagonist celiprolol following amplification of cAMP formation in failing human myocardium. The present study investigated the effects of celiprolol a novel beta 1-antagonist with partial beta 2-agonist activity on the human failing heart. Experiments were performed on isolated electrically driven atrial and ventricular cardiac preparations and in membrane preparations from the left ventricles of nine patients (four with dilated cardiomyopathy; five with ischemic cardiomyopathy) undergoing cardiac transplantation for terminal heart failure. Celiprolol produced a negative inotropic effect in atrial and ventricular heart muscle. However, in the presence of forskolin--which activates the catalyst of the adenylate cyclase-or the cAMP phosphodiesterase inhibitor milrinone, celiprolol produced concentration-dependent positive inotropic effects and positive lusitropic effects. Experiments with the beta 1-and beta 2-selective antagonists CGP 207.12A and ICI 118.551, respectively, suggest that the positive inotropic response is mediated by beta 2-adrenoceptors. In radioligand binding experiments, a selectivity of 15.7 [-Gpp(NH)p] or 23.9 [+Gpp(NH)p] as judged from the Ki values--of binding to beta 2-adrenoceptors was measured in the failing human ventricular myocardium. Competition curves with celiprolol alone and in the presence of the guanine nucleotide Gpp(NH)p revealed no evidence for agonist activity at beta 1- or beta 2-adrenoceptors. It is concluded that amplification of the cAMP response is able to unmask partial agonist activity of celiprolol in the failing human heart at beta 2-adrenoceptors. The inotropic measurements are a more sensitive approach than radioligand binding studies. Whether the pharmacological profile of celiprolol will be useful in conditions like heart failure is questionable with respect to the potential downregulation of beta 2-adrenoceptors by its partial agonist activity.


Effect of lisinopril and isosorbide-5-mononitrate on hemodynamics and mortality in rats with permanent coronary artery occlusion. We studied the hemodynamic effects of lisinopril and isosorbide-5-mononitrate in rats with permanent coronary occlusion. Rats (n = 35) underwent left coronary occlusion, and ECGs were recorded before and after occlusion. Ventricular arrhythmias were observed in 57% (20 of 35) of animals. Treatment was given immediately after coronary occlusion and for 2 subsequent days. The control group received 100 mg/kg lactose (i.e., 80% vehicle for isosorbide-5-mononitrate). Lisinopril (100 mg/kg body weight) and isosorbide-5-mononitrate (400 mg/kg body weight) reduced systolic blood pressure (SBP) from 134 +/- 9 to 115 +/- 9 mm Hg (p less than 0.05) and from 137 +/- 9 to 126 +/- 9 mm Hg, respectively; the hypotensive effect lasted 2-3 days. No effect on BP was noted in the control group. Overall mortality was 23%; 8 of 35 animals died within 10-15 min of coronary occlusion. Survival after 4 weeks was similar in each group (approximately 80%). Left ventricular pressure (LVP) was measured 4 weeks after coronary artery occlusion and was similar in each group. However, dP/dt was lower in hearts with infarction than in hearts with none (12,608 +/- 906 vs. 8,992 +/- 1,242 mm Hg/s). The extent of the infarction was the same in groups with coronary artery occlusion. Lisinopril is more effective than isosorbide-5-mononitrate in reducing BP after acute myocardial infarction (AMI). Medium-term survival (4 weeks) is not jeopardized by effective treatment with angiotensin-converting enzyme (ACE) inhibitors or long-acting nitrates.


The neutral endopeptidase inhibitor, SCH 34826, reduces left ventricular hypertrophy in spontaneously hypertensive rats. SCH 34826, i.e., (S)-N-(N-(2,2[(2,2-dimethyl-1,3-dioxolan-4- yl)methoxy]-2-oxo-1-(phenyl-methyl)ethyl)-phenylalanyl)-beta-alanine, is a potent and selective inhibitor of neutral endopeptidase 24.11 (NEP), an enzyme that degrades the atrial natriuretic peptide (ANP). The effects of SCH 34826 on hypertension and left ventricular hypertrophy (LVH) in spontaneously hypertensive rats (SHRs) were evaluated following 1 month of treatment by measuring the blood pressure, cardiac weight, and left ventricular fibrosis. Adult SHRs were treated with SCH 34826 at 10, 30, or 100 mg/kg given orally twice daily or with vehicle. The systolic blood pressure (SBP) and heart rate (HR) were recorded weekly by the tail-cuff method. Cardiac structural damage was determined by morphometric analysis. Over the dose range examined, the drug produced no significant changes in either blood pressure or heart rate. Despite the lack of antihypertensive activity, SCH 34826 at 100 mg/kg reduced both the cardiac mass (-10%) and the amount of fibrotic tissue present in the left ventricle (-42%). These data indicate that chronic inhibition of NEP by SCH 34826 interacts with mechanisms underlying myocardial hypertrophy and cardiac remodeling.


Single-dose and steady-state pharmacokinetics and pharmacodynamics of perindopril in hypertensive subjects. In a double-blind, placebo-controlled, parallel group study, 24 essential hypertensive subjects were randomised to receive either placebo or 2, 4, or 8 mg perindopril. Perindopril, its deesterified metabolite, perindoprilat, and perindoprilat glucuronide were separated with an ion-exchange resin and determined by a radioimmunoassay (RIA). Pharmacokinetic and pharmacodynamic parameters were estimated for 96 h after the first dose and after 4-week once-daily treatment. Perindopril peak levels were achieved in less than or equal to 2 h after dosing with an elimination t1/2 of 1-2 h. Peak levels of perindoprilat were achieved more slowly, reaching a maximum level 5-8 h after dosing, and had an elimination t1/2 of 40 h. Levels of the perindopril glucuronide peaked approximately 0.5 h later than perindopril, with an elimination t1/2 of approximately 2 h. Perindopril, perindoprilat, and its glucuronide conjugate followed linear kinetics in the dose range of 2-8 mg, and there was no evidence of accumulation with chronic dosing. Perindopril 4 and 8 mg produced significant decreases in predose blood pressure (BP) with chronic dosing, with maximal decreases occurring 5-7 h after dosing. Perindopril also produced a prolonged dose-dependent inhibition of plasma angiotensin-converting enzyme (ACE) activity that was maximum after 4 h and had not fully recovered by 48 h after a single dose.


The relevance of cellular to clinical electrophysiology in classifying antiarrhythmic actions. The division of class I antiarrhythmic agents (sodium-channel blockers) into Ia, Ib, and Ic subgroups was based on clinical observations. Lidocaine, mexiletine, and tocainide (Ib) did not alter the QRS or H-V interval in sinus rhythm, but prolonged effective refractory period (ERP) in spite of some shortening of the J-T interval. Encainide, flecainide, and lorcainide (Ic) widened the QRS and prolonged H-V in sinus rhythm and at low concentration, but had little effect on the ERP or J-T. These clinical findings could be explained by fast onset/offset kinetics of Ib drugs, that when used in high concentrations, blocked most sodium channels during the action potential plateau; therefore, at the beginning of diastole, insufficient drug-free channels were available to support conduction, and the ERP was prolonged. Rapid dissociation of the drugs after repolarization insured that by the end of diastole most channels were again drug free, so that the QRS and H-V were normal. The Ic compounds were more potent, but of slow onset, so that a steady-state block of Na channels was not achieved until after many beats. Offset was also slow, so that a proportion of channels was persistently unavailable, Na current was reduced, and conduction slowed, causing widening of the QRS and lengthening of H-V. Because the remaining drug-free channels were normal, they recovered rapidly from inactivation, and the ERP was not prolonged. By clinical criteria, moricizine also must be classed as Ic, and its offset/onset kinetics are much slower than those of Ib drugs.(ABSTRACT TRUNCATED AT 250 WORDS)


RP 58866 and its active enantiomer RP 62719 (terikalant): blockers of the inward rectifier K+ current acting as pure class III antiarrhythmic agents. The present article presents an overview of the pharmacologic profile of the benzopyran derivative RP 58866, a racemic mixture, and of RP 62719 (terikalant), its active enantiomer. In normal cardiac tissues studied in vitro, both drugs dose-dependently prolonged the atrial and ventricular action potential but affected neither the upstroke of the action potential nor the diastolic potential. Patch-clamp experiments demonstrated that the prolongation of the action potential induced by the drugs is due to a specific blockade of the inward rectifier K+ current. In vivo, intravenous administration to anesthetized dogs of low doses of RP 62719 consistently induced bradycardia and prolonged the atrial, nodal, and ventricular refractory periods, but did not affect the conduction velocity. Because of these properties, RP 58866 and RP 62719 exert potent antiarrhythmic and antifibrillatory actions both at the atrial and ventricular levels in various experimental models of arrhythmia. Our results demonstrate that RP 58866 and RP 62719 are K(+)-channel blockers acting as pure class III antiarrhythmic drugs.


Statistical problems in the design of antiarrhythmic drug trials. Although many aspects of design, monitoring, and analysis in trials of antiarrhythmic drugs are similar to those of other intervention studies, several features are different. These include decisions on how to identify the subjects, timing of the intervention with respect to the disease process, time of randomization, methods for measurement of outcome, and interim monitoring for harm or benefit. The resolution of these issues depends almost entirely on the specification of the question of interest.


Development of class III antiarrhythmic agents. Currently available antiarrhythmic agents that act by lengthening the action potential duration and refractory period exert most of their effect during bradycardia (class IIIB). Unfortunately, when these agents are most needed, i.e., during tachycardia, they lose most of their effect. In my opinion, ideally, antiarrhythmic agents need not change the electrophysiology of the normal sinus beat, but should lengthen refractoriness upon acceleration (class IIIA) of the heart until the refractory period exceeds the cycle length of the tachycardia. Three examples of how such class IIIA effects can be achieved are: (i) an upstroke-dependent activator of inward currents could form a universal class IIIA agent, (ii) a use-dependent blocker of iTO could be a great class IIIA agent in the atria, and (iii) a frequency-dependent blocker of late repolarizing currents (e.g., iKs) could be a good target for a ventricular agent.


Clinical implications of new insights into mechanism of antiarrhythmic drug action. A classification system of antiarrhythmic drugs should help the clinician to select the optimal antiarrhythmic drug for a particular arrhythmia in an individual patient. This goal cannot be achieved by the Vaughan Williams classification system. Other important goals of this classification system were to provide the clinician with some information about proarrhythmic effects and about the possibility of combining particular antiarrhythmic agents. To date, a subdivision of class III antiarrhythmic agents has not been accomplished.


How to evaluate class III antiarrhythmic drug efficacy clinically: the benefits and shortcomings of the noninvasive approach. Holter monitoring is the most commonly used noninvasive method for assessing antiarrhythmic drug therapy. It can easily be performed and we know the exact criteria for the definition of drug efficacy and the proarrhythmic effect. However, the value of Holter monitoring to predict clinical outcome in patients with malignant ventricular arrhythmias is controversial. Several authors claim that Holter monitoring is useless to evaluate antiarrhythmic drug effects. Theoretical considerations will explain that it is not the technique by itself that is useless; it is the way it is applied. Merely by changing the efficacy criteria, the value of Holter monitoring to assess antiarrhythmic effects can be improved significantly. Nevertheless, Holter monitoring is far from being the ideal method. It focuses only on arrhythmia density, denying that other risk factors such as heart rate variability, ischemia, and QT changes may be of additional prognostic value. Further studies with Holter monitoring should concentrate on the role of these parameters.


How to evaluate class III antiarrhythmic drug efficacy clinically: the benefits and shortcomings of the invasive approach. Ventricular tachycardia and ventricular fibrillation are frequent complications of organic heart disease. There is sufficient evidence that serial electrophysiologic testing is able to predict long-term efficacy of antiarrhythmic agents in patients with malignant ventricular tachyarrhythmias. This approach has not only been useful for the evaluation of class I drugs, but recent studies have shown that this invasive method may also be useful for the management of patients undergoing treatment with class III antiarrhythmic agents such as amiodarone and sotalol. The results of several studies suggest that class III agents are more effective than class I drugs in patients presenting with ventricular tachycardia or ventricular fibrillation. Proarrhythmic complications in patients treated with class III antiarrhythmic drugs are mainly characterized by torsades de pointes. Their incidence does not exceed 5%. Further studies are necessary to elucidate the mechanisms underlying this type of proarrhythmia. By the use of currently available stimulation techniques, patients who might develop torsades de pointes while on therapy with a class III agent cannot be identified.


The utility of serum drug level monitoring during therapy with class III antiarrhythmic agents. Serum drug level monitoring facilitates dosage optimization of agents with a narrow therapeutic index. Concentration-dependent antiarrhythmic and toxic effects have been demonstrated for several class I drugs, whereas the concentration-effect relationships of class III antiarrhythmics are less well established. The efficacy and toxicity of amiodarone, in particular, may be determined more by extensive tissue deposition than by its serum concentration. Furthermore, accumulation of the active metabolite desethylamiodarone seems to play an additional role. The antiarrhythmic serum concentration range of sotalol is wide; monitoring is not required. The clinical utility of serum concentration measurements with the newly developed class III antiarrhythmic agents remains to be evaluated in properly planned therapeutic trials in patients with supraventricular and ventricular arrhythmias.


Therapy with and assessment of class III antiarrhythmic agents in different patient populations. The present article reviews the therapeutic efficacy of class III antiarrhythmic agents, particularly amiodarone, in patients with atrial and ventricular arrhythmias in the setting of different cardiac disorders. Both Holter monitoring and electrophysiologic studies appear to constitute reliable methods for evaluating the effects of class III agents and the long-term outcome in patients with a history of sustained ventricular tachycardia or fibrillation. The efficacy and tolerance of amiodarone in different patient populations is discussed.


Clinical efficacy of long-term treatment with amiodarone for life-threatening arrhythmias. The present article reviews the experience with amiodarone therapy in life-threatening ventricular tachyarrhythmias. In 13 studies, 1,594 patients were treated with amiodarone in average doses of 300 mg/day. The mean annual death rate was 6.2%; the annual withdrawal rate averaged 8%. Risk factors for arrhythmia recurrence despite amiodarone therapy were low left ventricular ejection fraction, inducible ventricular tachycardia by programmed electrical stimulation or persisting ventricular tachycardia on Holter monitoring, and cardiac arrest before the institution of amiodarone therapy. In these patients, additional antiarrhythmic measures such as an automatic implantable cardioverter/defibrillator or other surgical interventions must be considered.


Efficacy of intravenously administered amiodarone for short-term control of serious arrhythmias. Intravenously administered amiodarone exerts substantial antiarrhythmic activity that is based on several different electrophysiologic mechanisms. Of these, the most important consists of an antitachycardic effect together with a significant prolongation of repolarization. The acute intravenous administration of amiodarone has been found to be effective in supraventricular and ventricular tachyarrhythmias resistant to conventional antiarrhythmic agents. Because amiodarone is free of detrimental hemodynamic effects in the majority of patients, intravenous administration of this compound seems to be particularly suitable in patients with serious arrhythmias in the setting of compromised left ventricular function.


Drug combinations and interactions with class III agents. Drug combinations with class III antiarrhythmic agents have not been studied systematically. There are data indicating that enhanced efficacy as evaluated by electrophysiologic studies can be obtained by combining sotalol and a type Ia drug or amiodarone with low-dose beta-blocking drugs. Combining amiodarone with a type Ia or Ic drug or with a beta-blocking drug can slow the rate of ventricular tachycardia to make the ventricular tachycardia hemodynamically well tolerated. Pharmacokinetic drug interactions with amiodarone are extensive. Recently, it has been shown that hepatic metabolism of amiodarone can be induced to enhance the conversion of amiodarone to N-desethylamiodarone. The physiologic effect of this drug transformation is uncertain. There should be few pharmacokinetic drug interactions with sotalol because this drug is almost completely absorbed, does not bind to plasma proteins, and is excreted unchanged by the kidneys.


Antiarrhythmic actions of DL-sotalol in ventricular and supraventricular arrhythmias. DL-Sotalol is a unique antiarrhythmic agent combining potent beta-blocking properties with the propensity to prolong cardiac repolarization in all myocardial tissues. Its beta-blocker-associated negative inotropic effects are attenuated by its action potential-lengthening effect. The drug has an elimination half-life of 10-15 h. Sotalol's major electrophysiologic profile constitutes the summed effects of beta-blockade and prolonged repolarization. It exerts a potent antifibrillatory action modulated by its antiadrenergic effects. The effectiveness and tolerance of sotalol in a variety of atrial and ventricular dysrhythmias is reviewed in the present article.


Differential analysis of frequency-dependent effects of antiarrhythmic drugs: importance of the saturation behavior of frequency-dependent sodium-channel blockade. Frequency-dependent effects of class I antiarrhythmic drugs on Vmax reported in the literature are analyzed with respect to periodical drug binding to sodium channels. The analysis reveals that class I action can be differentiated according to the onset kinetics as well as to the saturation behavior of frequency-dependent sodium-channel blockade at increasing frequencies. As will be shown, class I drugs even of the same subclass (Ic) may differ markedly from each other with respect to the saturation behavior of frequency-dependent block. These findings may be of interest in view of the Cardiac Arrhythmia Suppression Trial (CAST) because the results found with flecainide and encainide in this study are usually extrapolated to other Ic drugs. Additionally, the influence of postrepolarization refractoriness caused by a class I drug on the action potential shortening during repetitive premature stimulation is compared with the effects of prolongation of absolute refractoriness induced by a class III drug.


Electrophysiologic and antiarrhythmic effects of D-sotalol. There is extensive experimental and clinical experience concerning the antiarrhythmic potency of DL-sotalol, a beta-adrenergic antagonist that lengthens the cardiac action potential duration. More recently, its dextrorotatory isomer, D-sotalol, has been evaluated in experimental studies and preliminary clinical trials. Whereas the isomer is almost free of beta-blocking activity, both experimental and clinical data demonstrate that potent class III activity of D-sotalol. Further investigations, particularly in patients with life-threatening arrhythmias, are needed to delineate the clinical usefulness of this compound.


Dofetilide, a novel class III antiarrhythmic agent. Dofetilide is a potent and selective class III antiarrhythmic agent that is under development for the treatment of re-entrant tachyarrhythmias (ventricular tachycardia/ventricular fibrillation, atrial fibrillation/atrial flutter, and paraoxysmal supraventricular tachycardia). In animal studies, dofetilide selectively inhibits the rapid component of the time-dependent outward potassium current (IKr) and therefore increases the effective refractory period and action potential duration without affecting the fast inward sodium current. Studies in dogs have shown that dofetilide (a) prolongs the effective refractory period in a dose-dependent manner, (b) elevates ventricular fibrillation threshold, (c) facilitates conversion of electrically induced ventricular fibrillation or fibrilloflutter to sinus rhythm, (d) does not influence conduction within the His-Purkinje system or within the myocardium, (e) does not impair cardiac contractility, and (f) reduces dispersion of ventricular repolarization. Dofetilide has been administered to healthy volunteers as well as to patients with ischemic heart disease or with supraventricular arrhythmias; the compound has generally been well tolerated. Side effects have occasionally been reported, but have generally been transient and mild and occur in placebo-treated subjects as well. No clinically significant changes in laboratory safety tests have been detected. The pharmacokinetic profile of dofetilide both in healthy volunteers and patients includes a linear dose-plasma concentration relationship and also a linear plasma concentration-QTc relationship. The terminal plasma elimination half-life is approximately 9-10 h and systemic bioavailability in the region of 100%. The elimination pattern is balanced, with 50% being excreted unchanged via the kidney, the remaining 50% being metabolized in the liver to inactive metabolites, with greater than 90% of circulating drug-related material being unchanged dofetilide. After intravenous administration of the compound, a slight hysteresis in the plasma drug level-QTc relationship has been detected. Pharmacodynamic data demonstrate dose- and concentration-dependent effects on myocardial repolarization as evidenced by prolongations of the QTc interval. This is reflected in significant prolongations in the effective and functional refractory periods and monophasic action potential duration throughout the myocardium. No effects on sinus node function, conduction parameters, or cardiac contractility have been detected in any of the clinical studies, supporting the contention that dofetilide is a highly selective class III antiarrhythmic agent.


Chromosome aberrations of human small cell lung cancer induced by a new 111In-bleomycin complex. A new 111Indium labeled bleomycin complex (111In-BLMC) was prepared and found to be effective for tumor imaging and therapy both in mouse glioma and human small cell lung cancer (SCLC) cells. Chromosome aberrations were studied in human SCLC cells to explore its mechanisms of killing cancer cells. SCLC cells (N417) were exposed to 111In-BLMC, BLM, or 111InCl3 (for control) for 1 hour, treated with colcemid, and chromosomal changes were analyzed. A dramatic increase in chromatic gaps, breaks, chromosome breaks, double minutes, rings, triradii, quadriradii, and chromosome stickiness were observed in the cells treated by 111In-BLMC compared to BLM or 111InCl3. These results indicated that 111In-BLMC has therapeutic potential for combination chemo-radiotherapy of cancer (e.g., by Auger electrons and local energy deposition).


Early palliation of univentricular hearts with subaortic stenosis and ventriculoarterial discordance. The arterial switch option. The optimal Fontan-type operation greatly depends on appropriate initial palliation. Several surgical techniques have been used in infancy to palliate patients with univentricular hearts, ventriculoarterial discordance, and subaortic stenosis. The two most common are pulmonary artery banding and Damus-Norwood procedures. Palliative arterial switch operation is another surgical option that was used in this early series of seven infants. The principle of this operation is to "switch" the subaortic obstruction into a subpulmonary obstruction; the coronary artery relocation on the large pulmonary trunk creates a harmonious aortic root and the connection of the rudimentary ventricular chamber to the pulmonary artery trunk creates a natural protection of the pulmonary vascular bed through the restrictive bulboventricular foramen. Seven infants with univentricular hearts, ventriculoarterial discordance, and subaortic stenosis underwent a palliative arterial switch operation. All infants had an associated aortic arch obstruction of various degrees, including one with interrupted aortic arch, five with coarctation with severe arch hypoplasia, and one with isolated arch hypoplasia. There were three with double-inlet left ventricle, three with tricuspid atresia, and one with transposition of the great arteries with ventricular septal defect and severe right ventricular hypoplasia. The subaortic obstruction was patent at birth in five patients who underwent a palliative switch operation in the first 2 months of life, and rapidly occurred following a previous neonatal pulmonary artery banding associated with arch repair in two patients who underwent a switch operation at 5 and 8 months of age, respectively. The operation includes aortic arch repair without prosthetic material, an atrial septectomy, and the arterial switch. An associated pulmonary shunt was required in five patients and a pulmonary artery banding in one. There was one early death in a patient with [S,L,L] anatomy and congenital atrioventricular block, leading to an early mortality of 14% (95% confidence limits: 1% to 28%). There was one late death. Four survivors are waiting for a Fontan-type procedure, and one survivor had satisfactory right ventricular growth. Early palliative arterial switch operation offers several advantages: reconstruction of a harmonious aortic root, natural protection of the pulmonary bed through the restrictive bulboventricular foramen, prevention of deleterious myocardial hypertrophy, and arch reconstruction without the introduction of a foreign material. This aggressive technique may provide a satisfactory palliation in infants with univentricular hearts and ventriculoarterial discordance, when the bulboventricular foramen/aortic anulus ratio is less than 0.8 or when the subaortic stenosis is severe enough to be associated with an arch obstruction.(ABSTRACT TRUNCATED AT 400 WORDS)


Combination use of suboptimal dose of FK 506 and cyclosporine in canine lung transplantation. The immunosuppressive potency and the side effects of combination therapy with FK 506 and cyclosporine A were studied in dogs that had undergone lung transplantation. The animals were divided into four groups: group A (one third optimal FK 506 dose: FK 506, 0.03 mg/kg intramuscularly) (n = 5), group B (one third optimal cyclosporine dose: cyclosporine 6 mg/kg orally) (n = 5), group C (one third FK 506 and one third cyclosporine optimal doses): FK 506, 0.03 mg/kg intramuscularly plus cyclosporine 6 mg/kg orally) (n = 5), and group D (half FK 506 plus half cyclosporine optimal doses: FK 506, 0.05 mg/kg intramuscularly, plus cyclosporine, 10 mg/kg orally) (n = 10). Assessments including chest x-ray film, fiberoptic bronchoscopy, hematologic and biochemical tests, FK 506 and cyclosporine blood trough level measurement, right pulmonary artery occlusion test, and histopathologic observations were performed. In group A two of five dogs survived 28 days and three died on postoperative days 7, 14, and 21. In group B one dog survived 28 days and four died on postoperative days 9 (two dogs), 14, and 21. Histologic examination showed severe rejection in both group A and group B. In group C all five dogs survived 28 days but showed mild rejection. In group D one dog died of intestinal bleeding on postoperative day 7 and nine survived 28 days. No pathologic changes were observed except in one case of mild rejection. The ventilation function of the transplanted lung was poor in groups A, B, and C but good in group D. No abnormal rise of FK 506 and cyclosporine trough levels was observed. There were no significant side effects and abnormal hematologic and biochemical data except in one dog in group D. We concluded (1) the combination of FK 506, 0.03 mg/kg, and cyclosporine, 6 mg/kg, is much more effective than either drug used singly, (2) the combination of FK 506, 0.05 mg/kg, and cyclosporine, 10 mg/kg, prevents rejection with tolerable side effects, and (3) no worse side effects are caused by combination therapy with FK 506 and cyclosporine than by either one used singly.


An improved HRP method for tracing axons in whole-mount preparations during early stages of regeneration in peripheral nerves of adult animals. An improved method using HRP for visualizing axons in whole-mount preparations is described. Application of this method to preparations of regenerating frog peripheral nerves maintained in organ culture was used to show that regeneration following nerve crush begins after about 3 days.


Quantitative microdialysis of dopamine in the striatum: effect of circadian variation. Two quantitative microdialysis methods were used to determine the concentration of extracellular dopamine in the anterior striatum of the rat. In the first method, the slow perfusion flow rate method, perfusion was at 57 nl/min and dialysate samples were collected every 90 min for 18 h and assayed for dopamine (DA), DOPAC (3,4-dihydroxy-phenylacetic acid), homovanillic acid (HVA) and 5-hydroxy-indoleacetic acid (5-HIAA). There was a significant increase in the concentration of dopamine during the dark cycle compared with the light cycle (14.7 +/- 1 nM vs. 9.3 +/- 0.7 nM; mean +/- SEM; P less than 0.0001), indicating possible circadian variations in the extracellular concentration of DA. There was a steady decrease in the level of DOPAC and HVA, and no change in the level of 5-HIAA. For the point of no-net-flux method, animals were perfused with 4 concentrations of DA or DOPAC, bracketing the extracellular concentrations. The extracellular concentrations of DA and DOPAC using this method were 10.2 +/- 1.7 nM and 17.4 +/- 2.6 microM, respectively. The in vivo recoveries for DA and DOPAC as derived from the slope of the linear regression curves were 72 +/- 3% and 43 +/- 5%. These values were shown to be significantly different (P less than 0.001). Both methods gave similar results for the level of DA in the striatum.


Enzyme-histochemistry of the foreskin microvasculature using fructose-1,6-diphosphatase. Microvascular capillaries (blood and lymph) in human foreskin were examined for staining characteristics for fructose-1,6-diphosphatase (FDPase). Based on enzyme-histochemistry and electron microscopy, both blood and lymph vessels and microvessels as well as dermal smooth muscle all stain positively but with varying intensity for FDPase.


Inhibitors of proteases prevent endonucleolysis accompanying apoptotic death of HL-60 leukemic cells and normal thymocytes. Exposure of human promyelocytic leukemic HL-60 cells to the topoisomerase I inhibitor camptothecin (CAM) triggers endonucleolytic activity and apoptotic death of these cells. The nucleolytic effect is seen 2-4 h after drug addition and is highly selective to cells progressing through S phase. Concomitant with degradation of DNA, which is preferential to the nucleosomal DNA linker sections, extensive proteolysis takes place in these cells. Cellular RNA, however, is initially degraded to a much lesser degree than DNA or protein. Both endonucleolysis and proteolysis triggered by CAM in S-phase HL-60 cells can be prevented by the protease inhibitors N-tosyl-L-phenylalanylchloromethyl ketone (TPCK), N-tosyl-L-lysylchloromethyl ketone (TLCK) or partly by N-tosyl-L-arginine methyl ester (TAME), added simultaneously with CAM, or up to 30 min after exposure to CAM, at their respective concentrations known to inhibit proteases. The protective effect of these protease inhibitors on DNA degradation cannot be due to the suppression of cell progression through S phase because cells still replicate DNA in their presence, albeit at a reduced rate. Furthermore, TPCK and TLCK protect rat thymocytes against endonucleolysis induced by prednisolone. In the latter cell system, (considered a classic model of apoptosis), endonucleolysis, which primarily affects G0/G1 cells, is unrelated to cell progression through S phase. The present data suggest that the endonucleolysis and proteolysis which accompany apoptotic cell death are coupled, and the proteolytic step is needed for DNA degradation to occur.


The significance of CD34 and TdT determinations in patients with untreated de novo acute myeloid leukemia. The results of intensive chemotherapy given to 247 adults at the University of Maryland Cancer Center with previously untreated de novo acute myeloid leukemia (AML) were reviewed with respect to expression of terminal deoxynucleotidyl transferase (TdT) and CD34. Of the 228 patients with data for TdT, 32 (14%) had > 5% of the leukemia cells positive by an immunofluorescence assay. The median age of the TdT-positive patients was approximately 10 years less than the TdT-negative patients (50 versus 60 years). Patients with TdT-positive AML had similar median survival (12 versus 10.5 months) and complete remission (CR) rates (53 versus 59%), but a greater frequency of long-term complete responders (60 of complete remitters versus 20%, p = 0.08) than TdT-negative patients. Of 126 patients tested, 59% were CD34-negative (< 20% reactivity with leukemia cells). These 74 patients (median age 60 years) had a greater CR rate (71 versus 48%, p = 0.008) than the 52 CD34-positive patients (median age 60 years), and improved survival (p = 0.013 by Wilcoxon) although there was no difference in the duration of CR between the CD34-positive and negative groups. Of CD34-positive patients 12/52 remain in continuous CR, and 16/74 CD34-negative patients remain in continuous CR. None of eight patients strongly positive for CD34 (> 70% reactivity) remain disease-free. Positivity for TdT or CD34 was associated with less differentiated AML. Of CD34-positive patients, 44% had FAB M0/M1 morphology versus 13% of CD34-negative patients (p = 0.0001); similarly, 47% of TdT-positive patients were FAB M0/ML1 versus 25% of TdT-negative patients (p = 0.01). Of seven patients with FAB M4E0, five were CD34-positive. Of the 12 CD34-positive survivors, four had FAB M4E0. Thus CD34 expression predicts for CR rate and overall survival in adults with AML. TdT expression does not significantly affect overall outcome but may be associated with longer CR durations.