Role of mitochondrial calcium in hypochlorite induced oxidative damage of cells.

Hypochlorite (HOCl) is one of the most important mediators of inflammatory processes. Recent evidence demonstrates that changes in intracellular calcium pool play a significant role in the damaging effects of hypochlorite and other oxidants. Mitochondria are shown to be one of the intracellular targets of hypochlorite. But little is known about the mitochondrial calcium pool changes in HOCl-induced mitochondrial dysfunction. Using isolated rat liver mitochondria, we showed the oxidative damage of mitochondria (GSH oxidation and mixed protein-glutathione formation without membrane lipid peroxidation) and alterations in the mitochondrial functional parameters (decrease of respiratory activity and efficiency of oxidative phosphorylation, NADH and FADH coenzyme levels, and membrane potential) under hypochlorite action (50-300 µM). Simultaneously, the mitochondrial calcium release and swelling were demonstrated. In the presence of EGTA, the damaging effects of HOCl were less pronounced, reflecting direct involvement of mitochondrial Ca2+ in mechanisms of oxidant-induced injury. Furthermore, exposure of HeLa cells to hypochlorite resulted in a considerable increase in cytoplasmic calcium concentrations and a decrease in mitochondrial ones. Applying specific inhibitors of calcium transfer systems, we demonstrated that mitochondria play a key role in the redistribution of cytoplasmic Ca2+ ions under hypochlorite action and act as mediators of calcium release from the endoplasmic reticulum into the cytoplasm.

Position Impact of Hydroxy Groups on Spectral, Acid-Base Profiles and DNA Interactions of Several Monohydroxy Flavanones.

Structure-related biological activities of flavanones are still considered largely unexplored. Since they exhibit various medicinal activities, it is intriguing to enter deeper into their chemical structures, electronic transitions or interactions with some biomolecules in order to find properties that allow us to better understand their effects. Little information is available on biological activity of flavanone and its monohydroxy derivatives in relation to their physicochemical properties as spectral profiles, existence of protonated/deprotonated species under pH changes or interaction with Calf Thymus DNA. We devoted this work to research demonstrating differences in the physicochemical properties of the four flavanones: flavanone, 2'-hydroxyflavanone, 6-hydroxyflavanone and 7-hydroxyflavanone and linking them to their biological activity. Potentiometric titration, UV-Vis spectroscopy were used to investigate influence of pH on acid-base and spectral profiles and to propose the mode of interaction with DNA. Cyclic voltammetry was applied to evaluate antioxidant potentiality and additionally, theoretical DFT(B3LYP) method to disclose electronic structure and properties of the compounds. Molecular geometries, proton affinities and pKa values have been determined. According to computational and cyclic voltammetry results we could predict higher antioxidant activity of 6-hydroxyflavanone with respect to other compounds. The values of Kb intrinsic binding constants of the flavanones indicated weak interactions with DNA. Structure-activity relationships observed for antioxidant activity and DNA interactions suggest that 6-hydroxyflavanone can protect DNA against oxidative damage most effectively than flavanone, 2'-hydroxyflavanone or 7-hydroxyflavanone.

Calcium-Induced Mitochondrial Permeability Transitions: Parameters of Ca2+ Ion Interactions with Mitochondria and Effects of Oxidative Agents.

We evaluated the parameters of Ca2+-induced mitochondrial permeability transition (MPT) pore formations, Ca2+ binding constants, stoichiometry, energy of activation, and the effect of oxidative agents, tert-butyl hydroperoxide (tBHP), and hypochlorous acid (HOCl), on Ca2+ -mediated process in rat liver mitochondria. From the Hill plot of the dependence of MPT rate on Ca2+ concentration, we determined the order of interaction of Ca2+ ions with the mitochondrial sites, n = 3, and the apparent Kd = 60 ± 12 µM. We also found the apparent Michaelis-Menten constant, Km, for Ca2+ interactions with mitochondria to be equal to 75 ± 20 µM, whereas that in the presence of 300 µM tBHP was 120 ± 20 µM. Using the Arrhenius plots of the temperature dependences of apparent mitochondrial swelling rate at various Ca2+ concentrations, we calculated the activation energy of the MPT process. ΔEa was 130 ± 20 kJ/mol at temperatures below the break point of the Arrhenius plot (30-34 °C) and 50 ± 9 kJ/mol at higher temperatures. Ca2+ ions induced rapid mitochondrial NADH depletion and membrane depolarization. Prevention of the pore formation by cyclosporin A inhibited Ca2+-dependent mitochondrial depolarization and Mg2+ ions attenuated the potential dissipation. tBHP (10-150 µM) dose-dependently enhanced the rate of MPT opening, whereas the effect of HOCl on MPT depended on the ratio of HOCl/Ca2+. The apparent Km of tBHP interaction with mitochondria in the swelling reaction was found to be Km = 11 ± 3 µM. The present study provides evidence that three calcium ions interact with mitochondrial site with high affinity during MPT. Ca2+-induced MPT pore formations due to mitochondrial membrane protein denaturation resulted in membrane potential dissipation. Oxidants with different mechanisms, tBHP and HOCl, reduced mitochondrial membrane potential and oxidized mitochondrial NADH in EDTA-free medium and had an effect on Ca2+-induced MPT onset.

Cranberry flavonoids prevent toxic rat liver mitochondrial damage in vivo and scavenge free radicals in vitro.

The present study was undertaken for further elucidation of the mechanisms of flavonoid biological activity, focusing on the antioxidative and protective effects of cranberry flavonoids in free radical-generating systems and those on mitochondrial ultrastructure during carbon tetrachloride-induced rat intoxication. Treatment of rats with cranberry flavonoids (7 mg/kg) during chronic carbon tetrachloride-induced intoxication led to prevention of mitochondrial damage, including fragmentation, rupture and local loss of the outer mitochondrial membrane. In radical-generating systems, cranberry flavonoids effectively scavenged nitric oxide (IC50 = 4.4 ± 0.4 µg/ml), superoxide anion radicals (IC50 = 2.8 ± 0.3 µg/ml) and hydroxyl radicals (IC50 = 53 ± 4 µg/ml). The IC50 for reduction of 1,1-diphenyl-2-picrylhydrazyl radicals (DPPH) was 2.2 ± 0.3 µg/ml. Flavonoids prevented to some extent lipid peroxidation in liposomal membranes and glutathione oxidation in erythrocytes treated with UV irradiation or organic hydroperoxides as well as decreased the rigidity of the outer leaflet of the liposomal membranes. The hepatoprotective potential of cranberry flavonoids could be due to specific prevention of rat liver mitochondrial damage. The mitochondria-addressed effects of flavonoids might be related both to radical-scavenging properties and modulation of various mitochondrial events.

Oxidative damage of rat liver mitochondria during exposure to t-butyl hydroperoxide. Role of Ca²âº ions in oxidative processes.

AIMS: The present study was designed for further evaluation of the biochemical mechanism of hepatic mitochondrial dysfunction under oxidative damages induced by organic hydroperoxide, tert-butyl hydroperoxide (tBHP), for estimation of the molecular targets impaired during oxidative stress, and for investigation of the role of Ca(2+) ions in mitochondrial oxidative reactions and of the protective effect of melatonin during mitochondrial peroxidative damage. MAIN METHODS: Mitochondria were isolated by differential centrifugation from the rat liver. The effects of tBHP exposure, EDTA, Ca(2+) ions and melatonin on mitochondrial respiratory activity, mitochondrial enzyme activities and redox status were measured. KEY FINDINGS: The present study provides evidence that tBHP (at low concentrations of 0.02-0.065mM, in EDTA-free medium) induced uncoupling of the oxidation and phosphorylation processes and decreased the efficiency of the phosphorylation reaction. This effect depended on the respiratory substrate used. The presence of EDTA prevented oxidative impairment of mitochondrial respiration, but Ca(2+) ions in the medium enhanced oxidant-induced mitochondrial damage considerably. In the presence of 0.5mM EDTA, tBHP (at high concentrations, 0.5-2mM) considerably oxidized mitochondrial reduced glutathione, enhanced accumulation of membrane lipid peroxidation products and mixed protein-glutathione disulfides and led to an inhibition of oxoglutarate dehydrogenase and succinate dehydrogenase. SIGNIFICANCE: Direct oxidative modification of enzymatic complexes of the respiratory chain and mitochondrial matrix, mitochondrial reduced glutathione depletion, protein glutathionylation, membrane lipid peroxidation and Ca(2+) overload are the main events of mitochondrial peroxidative damages. Experiments in vitro demonstrated that melatonin inhibited the mitochondrial peroxidative damage, preventing redox-balance changes and succinate dehydrogenase inactivation.

Synthesis of Block Copolymers of Varying Architecture Through Suppression of Transesterification during Coordinated Anionic Ring Opening Polymerization.

Well-defined di- and triblock copolymers consisting of ε-caprolactone (CL), L-lactide (LA), and trimethylene carbonate (TMC) were synthesized via "PLA first route" in coordinated anionic ring opening polymerization/copolymerization (CAROP) with tin (II) octoate as catalyst. The desired block structure was preserved by use of protective additive α-methylstyrene by preventing the transesterification side-reactions. MALDI-TOF analysis revealed that the protection mechanism is associated with α-methylstyrene and tin (II) octoate complexation. Additionally, it was shown that use of α-methylstyrene in ring opening polymerization allowed the formation of polyesters with high molar mass.

Triblock copolymers of ε-caprolactone, trimethylene carbonate, and L-lactide: effects of using random copolymer as hard-block.

A series of triblock copolymers comprising end block of PLLA modified with PCL, and random copolymer of PCL and PTMC as soft segment were synthesized. DSC data show that PCL disrupted the crystallinity of PLLA, making the hard block to be completely amorphous when the PCL content is 50%. Correspondingly, the addition of PCL into PLLA block enhances the elongation of the triblock considerably. With regards to the elasticity, however, creep test results show that adding PCL to PLLA block seems to reduce the "equilibrium" recovery, while cyclic test results shows that the instantaneous recovery increased significantly with more PCL inside PLLA block. It was also observed that the degradation rate of triblock with added PCL inside the PLLA was slower compared to triblock with pure PLLA hard block. Compared to biodegradable polyurethane, these polymers are expected to yield less harmful degradation products, and offer more variables for the manipulation of properties. These polymers are also processable from the melt at temperatures exceeding about 130 °C. We expect to use these polymers in a variety of applications, including stent coatings, fully-degradable stents and atrial septal defect occluders.

Asociación entre niveles de TNF-a en fluido crevicular gingival de dientes con periodontitis apical asintomática / Levels of TNF-a increase in gingival crevicular fluid of teeth with asymptomatic apical periodontitis

Introducción: Como consecuencia de necrosis pulpar séptica, el ingreso de productos bacterianos en el periápice induce la producción de citoquinas pro-inflamatorias como el Factor de Necrosis Tumoral (TNF)- a, que entre otras funciones participa en la diferenciación y activación de los osteoclastos para inducir reabsorción ósea, fenómeno característico de la periodontitis apical asintomática (PAA). El fluido crevicular gingival (FCG) ofrece un gran potencial como fuente de factores asociados con la actividad osteoclástica. El objetivo del presente estudio fue determinar los niveles de TNF- a en FCG de dientes con PAA y controles sanos contralaterales. Métodos: Se incluyeron 14 pacientes en la Clínica de Endodoncia de la Facultad de Odontología de la Universidad de Chile con diagnóstico clínico de PAA y se obtuvieron muestras de FCG con tiras de papel a partir de los dientes afectados y sanos contralaterales por un periodo estandarizado de 30 segundos. Se determinaron las concentraciones de proteínas totales mediante el método del ácido bisciconitico y los niveles de TNF- a, mediante ensayo ELISA. Los datos se analizaron con Test-t pareado utilizando el programa StataV11. Resultados: Se detectaron niveles de TNF- a significativamente mayores en el FCG de dientes con PAA estandarizados, tanto por 30 segundos de toma de muestra como por mg de proteínas totales. Conclusiones: Este estudio provee evidencia preliminar de que los niveles de TNF- a en el FCG reflejan la presencia de PAA y podría ser de utilidad como complemento al diagnóstico clínico y monitoreo del estado de salud o enfermedad de los tejidos perirradiculares.

Introduction: As a consequence of septic pulp necrosis, the entry of bacterial products into periapical tissues induces the release of pro-inflammatory cytokines, such as the tumor necrosis factor (TNF)- a. This pleiotropic cytokine is involved in the differentiation and activation of osteoclasts to induce bone resorption, a hallmark of asymptomatic apical periodontitis (AAP). Gingival crevicular fluid (GCF) has a great potential as a source of factors associated with osteoclastic activity. The aim of this study was to determine the levels of TNF- a in GCF of teeth with PAA and contralateral healthy controls. Methods: A total of 14 patients with clinical diagnosis of AAP were enrolled from the Clinic of Endodontics, Faculty of Dentistry, Universidad de Chile during 2009 and 30-second GCF samples were obtained with paper strips from AAP teeth and contralateral healthy controls. Total protein concentration and TNF- a levels were determined through bisciconitic acid method and ELISA assay, respectively. Paired t test and StataV11 software were used for statistical analysis. Results: Levels of TNF- a were significantly higher in GCF from teeth with AAP than controls when standardized by either 30s of sampling and total protein content. Conclusions: The present study provides preliminary data supporting that TNF- a levels in GCF reflect periapical status. Screening of TNF- a levels in GCF might represent a useful side-diagnostic tool to the monitoring of the apical status.

Corrections by melatonin of liver mitochondrial disorders under diabetes and acute intoxication in rats.

The aim of the present work was to investigate the mechanisms of oxidative damage of the liver mitochondria under diabetes and intoxication in rats as well as to evaluate the possibility of corrections of mitochondrial disorders by pharmacological doses of melatonin. The experimental (30 days) streptozotocin-induced diabetes mellitus caused a significant damage of the respiratory activity in rat liver mitochondria. In the case of succinate as a respiratory substrate, the ADP-stimulated respiration rate V3 considerably decreased (by 25%, p < 0·05) as well as the acceptor control ratio (ACR) V3/V2 markedly diminished (by 25%, p < 0·01). We observed a decrease of the ADP-stimulated respiration rate V3 by 35% (p < 0·05), with glutamate as substrate. In this case, ACR also decreased (by 20%, p < 0·05). Surprisingly, the phosphorylation coefficient ADP/O did not change under diabetic liver damage. Acute rat carbon tetrachloride-induced intoxication resulted in considerable decrease of the phosphorylation coefficient because of uncoupling of the oxidation and phosphorylation processes in the liver mitochondria. The melatonin administration during diabetes (10 mg·kg⁻¹ body weight, 30 days, daily) showed a considerable protective effect on the liver mitochondrial function, reversing the decreased respiration rate V3 and the diminished ACR to the control values both for succinate-dependent respiration and for glutamate-dependent respiration. The melatonin administration to intoxicated animals (10 mg·kg⁻¹ body weight, three times) partially increased the rate of succinate-dependent respiration coupled with phosphorylation. The impairment of mitochondrial respiratory plays a key role in the development of liver injury under diabetes and intoxication. Melatonin might be considered as an effector that regulates the mitochondrial function under diabetes.

Triblock copolymers of ε-caprolactone, L-lactide, and trimethylene carbonate: biodegradability and elastomeric behavior.

For the triblock copolymer of ε-caprolactone, trimethylene carbonate, and L-lactide, where L-lactide blocks form the two ends, there is a range of compositions over which elastomeric behavior is obtained. Within this composition range, these polymers show good creep and recovery at ambient temperature, and exhibit high elongations to break. Additionally, we demonstrate that the recovery is independent of stress and strain for the elastomer compositions. The range of compositions that yield elastomeric character is rationalized based on the structure; specifically, there must be a minimum crystallinity of the end blocks and no crystallinity in the midblock, in addition to molar mass requirements. These polymers degrade by simple hydrolysis, and the rate of degradation is potentially programmable by manipulation of the molar ratio of hard segment to soft segment. Compared to biodegradable polyurethane, these polymers are expected to yield less harmful degradation products, and offer more variables for manipulation of properties. These polymers are also processable from the melt at temperatures exceeding about 130 °C. We expect to use these polymers in a variety of applications, including stent coatings, fully-degradable stents, and atrial septal defect occluders.

Thermoplastic biodegradable elastomers based on ε-caprolactone and L-lactide block co-polymers: a new synthetic approach.

Although biodegradable polymers have found extensive application in medical devices, there are very few commercially available elastomeric biodegradable polymers. In this work, starting with the well-known monomers L-lactide and ε-caprolactone, we developed elastomers using a multiblock co-polymer approach. This ensures that the degradation products of such elastomers are also acceptable from a cytotoxicity standpoint. A series of polymers with various structures was synthesized utilizing a design of experiment approach. The basic structure is that of a diblock, with each block being modified by the addition of co-monomer. The synthesized polymers exhibited a range of mechanical properties from a typical thermoplastic polymer to that approaching a good thermoplastic elastomer. 13C nuclear magnetic resonance analysis, size exclusion chromatography and differential scanning calorimetry measurements have been utilized to relate the observed range of mechanical properties to the structure. In addition, the elastomeric nature has been established with the use of creep and recovery measurements. Such elastomers may find a variety of biomedical applications, ranging from stent coatings to atrial septal defect occluders.

Premature termination mutations in exon 3 of the SMN1 gene are associated with exon skipping and a relatively mild SMA phenotype.

Autosomal recessive spinal muscular atrophy (SMA) is a common motor neuron disease caused by absence or mutation in the survival motor neuron (SMN1) gene. SNM1 and a nearly identical copy, SMN2, encode identical proteins, but SMN2 only produces a little full length protein due to alternative splicing. The level of functional SMN protein and the number of SMN2 genes correlate with the clinical phenotype ranging from severe to very mild. Here, we report on premature termination mutations in SMN1 exon 3 (425del5 and W102X) which induce skipping of the mutated exon. The novel nonsense mutation W102X was detected in two patients with a relatively mild phenotype who had only two copies of the SMN2 gene, a number that has previously been found associated with the severe form of SMA. We show that the shortened transcripts are translated into predicted in frame protein isoforms. Aminoglycoside treatment suppressed the nonsense mutation in cultured cells and abolished exon skipping. Fibroblasts from both patients show a high number of nuclear structures containing SMN protein (gems). These findings suggest that the protein isoform lacking the exon 3 encoded region contributes to the formation of the nuclear protein complex which may account for the milder clinical phenotype.

Detection of the survival motor neuron (SMN) genes by FISH: further evidence for a role for SMN2 in the modulation of disease severity in SMA patients.

Spinal muscular atrophy (SMA) is a common autosomal recessive neuromuscular disorder which presents with various clinical phenotypes ranging from severe to very mild. All forms are caused by the homozygous absence of the survival motor neuron ( SMN1 ) gene. SMN1 and a nearly identical copy ( SMN2 ) are located in a duplicated region at 5q13 and encode identical proteins. The genetic basis for the clinical variability of SMA remains unclear, but it has been suggested that the copy number of SMN2 could influence the disease severity. We have assessed the number of SMN2 genes in patients with different clinical phenotypes by fluorescence in situ hybridization (FISH) using as SMN probe a mixture of small specific DNA fragments. Gene copy number was established by FISH on interphase nuclei, but the presence of two SMN2 genes on the same chromosome could also be revealed by FISH on metaphase spreads. All patients had at least two SMN2 genes. We found two or three copies of SMN2 in severely affected type I patients, three copies in intermediately affected type II patients, generally four copies in mildly affected type III patients and four or eight copies in patients with very mild adult-onset SMA. No alterations of the genes were detected by Southern blot and sequence analysis, suggesting that all gene copies of SMN2 were intact. These data provide additional evidence that the SMN2 genes modulate the disease severity and suggest that knowledge of the gene copy number could be of some prognostic value.

Construction of a 2.5-Mb integrated physical and gene map of distal 21q22.3.

The gene-rich telomeric region of 21q harbors several loci relevant to human diseases including autoimmune polyglandular disease type I, nonsyndromic deafness, Knobloch syndrome, holoprosencephaly, and bipolar affective disorder. A contig of genomic clones in this region would facilitate the isolation of these genes. However, distal 21q22.3 has yet been poorly mapped, presumably due to the presence of sequences that are underrepresented in yeast artificial chromosome (YAC) libraries. We generated a framework of YACs and used these clones as starting points for the isolation of a combination of bacterial artificial chromosome clones, P1-derived artificial chromosome clones, and cosmid clones by chromosome walking procedures. These studies resulted in the construction of a high-resolution contig map spanning the 2.5-Mb region from PFKL to the telomere, approximately 2 Mb of which are covered by ready-to-sequence contigs. Within this map we determined the location and relative distance of 21 markers. These include 9 established genetic markers, the order of which is cen-PFKL-D21S154-D21S170-D21S171-D21S1903- D21S1897- D21S112-D21S1446-D21S1575-tel. Moreover, we established the precise map position of 13 genes and 4 ESTs including the recently isolated genes C21ORF2, SMT3H1, RNA editing deaminase 1 (ADARB1), folate transporter (SLC19A1), COL18A1, lanosterol synthase (LSS-PEN), pericentrin (PCNT), and arginine methyltransferase (HRMT1L1). This integrated map provides a useful resource for the mapping and isolation of disease genes and for the construction of a complete transcription map of distal 21q as well as for large-scale sequencing efforts.

Ligands of the histamine H3-receptor: new potent antagonists of the 2-thioimidazole type.

An overview of H3-receptor ligands is presented, with particular attention to antagonists. The protein binding of the classical H3-receptor antagonist thioperamide and its effect on in vivo distribution are discussed. A series of H3-receptor antagonists characterised by the presence of an imidazole ring, a spacer (ethylthio-, ethylamino-, propylthio- or propylamino-chain), a second heterocycle nucleus and a lipophilic group is described. Their H3-receptor antagonist potency has been measured on electrically stimulated guinea-pig intestine, and their affinity for central H3-receptor has been determined by competitive inhibition of [3H]N alpha-methylhistamine binding to rat cortex. Biphasic inhibition curves have been observed in some cases. Compounds endowed with interesting activity belong mostly to the class of 2-[[2-[4(5)-imidazolyl]ethyl]thio]imidazole, having a phenyl or a cyclohexyl group.

QSAR study on H3-receptor affinity of benzothiazole derivatives of thioperamide.

Starting from the structure of thioperamide, a known H3-antagonist, a new series of compounds with a benzothiazole nucleus instead of the cyclohexylcarbothioamide moiety was synthesized. Various substituents, selected by experimental design, were introduced in position 6 of the benzothiazole nucleus, in order to change its physico-chemical characteristics. The lipophilicity of the synthesized compounds was measured by means of RP-HPLC, and their H3-receptor affinity was evaluated by competitive binding assays on rat cortex synaptosomes, with the labelled ligand N alpha-[3H]methylhistamine. A QSAR analysis was performed on the experimental data, using also substituent constants taken from the literature. The newly synthesized compounds showed lower H3-affinities than thioperamide; quantitative structure-activity relationships, described by models obtained with PLS and MRA techniques, were observed among benzothiazole derivatives. According to these relationships, any attempt to improve the potency of these compounds should involve the substitution of the benzothiazole moiety with less bulky and/or more flexible structures, which should also be less lipophilic and allow better electronic interactions with the binding site. 1-(Benzothiazol-2-yl)-4-[(1H)-imidazol-4-yl]piperidine represents a limit structure for H3-activity, since it seems impossible to improve its affinity by means of substitution in the studied position of the benzothiazole nucleus, as shown by predictions performed by a PLS model.

Rotational disorders in intramedullary fixation of the lower limbs.

The routine employment of intramedullary fixation in fractures of the femur and tibia has led the authors to thoroughly evaluate malunions. With the aid of CT scan, precise multilevel quantification of residual rotational deformity was possible by comparison with the contralateral limb after fracture healing. The results of this study confirm the effectiveness of the method, filling an interpretative gap in the literature.

Anomalous responses to histamine stimulation of guinea-pig oesophageal muscularis mucosae.

The incomplete tachyphylaxis of the contractile response to the H1-stimulants observed on guinea-pig oesophageal muscularis mucosae seems to be H2- and H3-antagonist as well as atropine- and tetrodotoxin-resistant. Lidocaine and eserine partially prevented this process probably by a mechanism independent of their main activity. The dualistic antagonism exerted by mepyramine and methysergide on reproducible histamine responses could be explained by a kinetic condition of "hemi-equilibrium state" together with changes of drug-receptor interaction and by non-specific properties of methysergide. On the whole, the present data indicate that the role of histamine in this tissue has still to be defined.

Influence of urea-equivalent groups in position 5 of 2-amino, 2-(1-aminoethylidenamino) and 2-guanidino thiazole derivatives on H2-receptor antagonist activity in gastric fistula cat.

A series of thiazole derivatives, in which a side-chain with different urea-equivalent groups was introduced in position 5 of the heterocycle, have been tested as inhibitors of dimaprit-induced gastric acid and pepsin secretion in the gastric fistula cat. By comparing the in vivo and the previously reported in vitro activity of these compounds, we can note a very close parallelism not only in the quality of their action but also in the estimates of pA2 values. These data support an interdependence between the molecular substructures and the affinity for the H2-receptor.

H2-receptor antagonist activity of N-methylthiourea, N-cyano-N'-methylguanidine, N-cyanoamidine, N-carbamoylamidine and N-sulfamoylamidine 5-substituted thiazole derivatives on guinea-pig atria.

The H2-antagonist activity of thiazole derivatives, substituted on position 5 with urea-equivalent groups, has been tested on guinea-pig isolated atria stimulated by dimaprit. By comparing the activities of the 2,5-disubstituted thiazole derivatives with those of the corresponding 2,4-disubstituted derivatives it can be seen that the side-chain position is critical to activity and differently influences activity in the various series. The heteroaromatic ring atom sequence N-C-S-C-side chain is always associated with a low antagonist activity.