New quinoxalin-1,3,4-oxadiazole derivatives: Synthesis, characterization, in vitro biological evaluations, and molecular modeling studies.

A new series of quinoxalin-1,3,4-oxadiazole (10a-l) derivatives was synthesized and evaluated against some metabolic enzymes including human carbonic anhydrase (hCA) isoenzymes I and II (carbonic anhydrases I and II), cholinesterase (acetylcholinesterase and butyrylcholinesterase), and α-glucosidase. Obtained data revealed that all the synthesized compounds were more potent as compared with the used standard inhibitors against studied target enzymes. Among the synthesized compounds, 4-fluoro derivative (10f) against hCA I, 4-chloro derivative (10i) against hCA II, 3-fluoro derivative (10e) against acetylcholinesterase and butyrylcholinesterase, and 3-bromo derivative (10k) against α-glucosidase were the most potent compounds with inhibitory activity around 1.8- to 7.37-fold better than standard inhibitors. Furthermore, docking studies of these compounds were performed at the active site of their target enzymes.

Evaluation of antiulcer activity of indole-3-carbinol and/or omeprazole on aspirin-induced gastric ulcer in rats.

The present work is an attempt to elucidate the antiulcer activity of indole-3-carbinol (I3C), which is one of the anticarcinogenic phytochemicals found in the vegetables of Cruciferae family such as broccoli and cauliflower, alone or in combination with omeprazole (OMP), a proton pump inhibitor, to diminish the effects of induced acute gastric ulcer by aspirin (ASA) in male albino rats. A total of 48 adult male albino rats were used in the present study. Animals were divided into eight experimental groups (six animals each group). They were given different experimental inductions of ASA at a dose of 500 mg/kg/body weight, OMP at a dose of 20 mg/kg/body weight and I3C at a dose of 20 mg/kg/body weight either alone or in combination with each other orally for a duration of 7 days. Inner stomach features, ulcer index, pH activity, body weight, stomach weight, hematological investigations, serum total protein albumin and reduced glutathione activity were investigated in addition to the histological, histochemical and immunohistochemical stain of cyclooxygenase-2 to the stomach tissue of normal control, ulcerated and treated ulcerated rats. The results of this study revealed that oral administration of ASA to rats produced the expected characteristic mucosal lesions. OMP accelerated ulcer healing but the administration of I3C either alone or in combination with OMP to ASA-ulcerated rats produced a profound protection to the gastric mucosa from injury induced by ASA. Our results suggested that administration of antiulcer natural substances such as I3C in combination with the perused treatment such as OMP is a very important initiative in the development of new strategies in ulcer healing.

Dynamic Psychotherapy: The Therapeutic Process in the Treatment of Obsessive-Compulsive Disorder.

Dynamic Psychotherapy (DP) was developed to overcome the limitations of traditional psychoanalysis, responding to a broader demand of patients who seek help to cope with specific problems in the short term, such as Obsessive-Compulsive Disorder (OCD). OCD is a chronic disabling mental disorder that leads to substantial distress, functional disability and severe occupational and social impairments. Recognizing the literature gap in this field, and the improvements reported by dynamic therapists who have dealt with patients suffering from OCD, a study on the treatment of these patients was conducted in order to discuss the effects of this technique. The method involved a narrative literature review and the analysis of two clinical cases to discuss therapeutic processes, which include the specificities of OCD patients and the mechanisms adopted in the treatment through DP. The therapist's active stance seemed to be essential to encourage the patient to face feared situations and identify the core conflict. Both patients who were treated through DP presented similarities, such as high anxiety, feelings of guilt and inhibition of aggressive and sexual impulses. Through emotional exploration, confrontation of defensive functioning and interpretative interventions of inner conflicts, patients had reached awareness of their hidden feelings and experiences, and their symptoms and feelings of guilt decreased. They also showed significant improvements in their interpersonal relationships. Although both treatments do not fit into short-term therapies, this technique has led to long-term results, providing evidence that DP may produce favorable outcomes in the treatment of OCD.

Essential parameters and risk factors of the patients for diabetes care and treatment.

AIM: The aim was to explore the association between Type 2 Diabetes Mellitus [T2DM] treatment satisfactions of patients regarding their socio-economic, life-style, history medication and clinical outcome in a Turkish population SUBJECT AND METHODS: A cross-sectional study conducted from February 2016 to September 2016. Of the total 1500 diabetic patients approached, 1094 (72.9%) gave their consent. Data analysis included, sociodemographic, serum lipid profiles (LDL, HDL), calcium, uric acid, blood pressure and glycated hemoglobin (HbA1c) at baseline and after six months. The Diabetes Treatment Satisfaction Questionnaire (DTSQ) tools were used to measure the patient satisfaction. RESULTS: The mean and standard deviation of age among gender, there were significant differences between males (51.81±14.40) and females (49.76±13.99) (p=0.024). There were statistically significant differences between males and females regarding place of living (city 76% vs town 26%) p<0.001) and consanguinity (p=0.040). Almost of the patients with diabetes were overweight (males 44.5% vs females 41.8%) while more than a quarter (31.2%) males were obese. Among patients with diabetes, significantly larger proportion were treated for DM with insulin (females 28.8% vs males 22.5%) and 'insulin & oral anti diabetic drugs' (females 21.6% vs males 18.4%%; p<0.001) in comparison. Reported average sleeping haours was significantly more among males (6.5±1.1 vs. 6.1±1.2; p<0.001) than females. Males and females reported significantly greater improvements in mean values of blood glucose (-2.07 p<0.001; vs. -2.36; p=0.007), HbA1c (-1.72 p<0.001; vs. -1.47 p=0.038), potassium (+0.98 p<0.001; vs. +0.93 p<0.005); albumin (-3.38 p<0.001; vs. -3.60; p<0.001); billirubin (-0.69 p=0.049; vs. -0.98; p<0.001); uric acid (+11.9 p=0.017; vs. +14.3; p<0.001); systolic blood pressure (-3.86 p<0.001; vs. -3.2 p<0.001) and diastolic blood pressure (-3.17 p<0.001; vs. -3.2 p<0.001) in comparison to 6 months before. Multivariate stepwise regression analysis showed that the satisfaction DTSQ scores for HbA1c (p<0.001), h of sleep (p<0.001), neuropathy (p=0.007), diabetic education (p=0.014), SBP (mmHg) (p=0.021) DBP (mmHg) (p=0.028) were identified as significantly associated with higher treatment satisfaction. CONCLUSION: The study confirms a positive correlation between diabetes patient's satisfaction with care and treatment. Females DM patients compared to males had a better satisfaction score with current treatment, unacceptably low blood glucose level, flexibility in treatment and understanding of DM.

Novel potassium-channel openers: preparation and pharmacological evaluation of racemic and optically active N-(6-amino-3-pyridyl)-N'-bicycloalkyl-N"-cyanoguanidine derivatives.

The previous paper reported on the synthesis and pharmacological evaluation of N-(6-amino-3-pyridyl)-N'-bicycloalkyl-N"-cyanoguanidine derivatives, from among which three compounds were selected as potent potassium-channel openers. In the present study, selected compounds were tested for antagonism of potassium-induced contraction of rat aorta, hypotensive activity in normotensive rats, and diuretic activity in spontaneously hypertensive rats. This led to further evaluation of compound (+/-)-10 and selection of (+)-N-(6-amino-3-pyridyl)-N'- [(1S,2R,4R)-bicyclo- [2.2.1]hept-2-yl]-N"-cyanoguanidine ((+)-10) (AL0670) for development as an antihypertensive agent. Although AL0670 is regarded as a pinacidil-type K(+)-channel opener, it showed different pharmacological and conformational profiles from pinacidil.

Novel potassium channel openers: synthesis and pharmacological evaluation of new N-(substituted-3-pyridyl)-N'-alkylthioureas and related compounds.

This report describes the synthesis and pharmacological evaluation of a series of novel potassium channel openers related to the pinacidil-type compounds. Thioureas, cyanoguanidines, and pyridine N-oxides were systematically evaluated for their effects on both the inhibition of spontaneous mechanical activity in rat portal vein (in vitro) and their antihypertensive activity (in vivo), and the structure-activity relationship for this series of compounds was discussed. Good correlation between in vitro and iv antihypertensive activity was observed for these compounds. Among them, cyanoguanidines bearing a conformationally rigid unit such as a norbornyl group generally possessed potent activity in both in vitro and in vivo studies. Especially, N-(6-amino-3-pyridyl)-N'-cyano-N"-(1-methyl-2-norbornyl)guanidine (23d) was identified as a more potent potassium channel opener in vitro (EC100 = 3 x 10(-8) M) than pinacidil (EC100 = 10(-7) M).

Synthesis, structure-activity relationships, and pharmacokinetic profiles of nonpeptidic difluoromethylene ketones as novel inhibitors of human chymase.

Potent human chymase inhibitors with high enzymatic selectivity and satisfactory metabolic stability were obtained by replacing the Val-Pro (P3-P2) dipeptide portion of the previously described inhibitor 1 with a nonpeptidic pyrimidinone skeleton. The potency of the novel compounds was further enhanced by the introduction of carbamoyl-substituted difluoromethylene ketone moieties. The most potent chymase inhibitor of the newly created series was 2u (Y-40018), which had a K(i) of 2.62 nM. Compound 2u possessed high selectivity for human chymase since it lacked significant activity toward other representative human proteolytic enzymes. Moreover its strict specificity for human chymase suggested that 2u strongly inhibited human and canine chymases but not rat and mouse ones. Pharmacokinetic studies in rats and dogs indicated that 2u was absorbed rapidly after oral administration and had satisfactory bioavailability in these experimental animal species (rat, 17%; dog, 32%). In conclusion, 2u is a novel, potent, and orally active chymase inhibitor which would prove very useful in revealing the precise roles of the latter in various pathophysiological processes.

Reduction of the leukemogenic potential of malignant murine leukemic cells by in vivo treatment with recombinant human granulocyte colony-stimulating factor.

We have investigated the effect of recombinant human granulocyte colony-stimulating factor (rhG-CSF) administration on the leukemogenic potential of L-103 murine leukemic cells. Leukemogenic potential was assessed by comparing the regression lines drawn between the number of inoculated leukemic spleen cells and the mean survival time (MST) of the syngeneic recipients. rhG-CSF injected 2.5 micrograms daily for 14 days reduced the leukemogenic potential of spleen cells of the leukemic mice to 1/200 of the control. This phenomenon was not observed with the leukemic spleen cells treated with r-murine granulocyte-macrophage (rmGM)-CSF in vivo. Cytochemical study indicated that morphologically identifiable blast cells were fewer in the rhG-CSF-treated leukemic spleen. Furthermore, leukemic cells in the rhG-CSF-treated spleen were less proliferative than the control in spite of having more clonogenic cells in the leukemic cell preparation. Cytogenetical analysis showed that chromosome abnormalities found in the original leukemic cells were not altered by rhG-CSF administrations. It also showed that the frequency of the abnormal karyotype was reduced in rhG-CSF-treated leukemic spleen (4/17) as compared with the control (8/8), indicating that the mitotic fraction was smaller in the rhG-CSF-treated leukemic cells. These findings indicate that in addition to the reduced number of leukemic cells in the spleen cell preparation, a reduction of the proliferative capacity of the original leukemic cells is involved in the reduction of leukemogenic potential of leukemic cells treated with rhG-CSF in vivo.

Energetic feasibility of hydrogen abstraction and recombination in coenzyme B(12)-dependent diol dehydratase reaction.

Coenzyme B(12) serves as a cofactor for enzymatic radical reactions. The essential steps in all the coenzyme B(12)-dependent rearrangements are two hydrogen abstraction steps: hydrogen abstraction of the adenosyl radical from substrates, and hydrogen back-abstraction (recombination) of a product-derived radical from 5'-deoxyadenosine. The energetic feasibility of these hydrogen abstraction steps in the diol dehyratase reaction was examined by theoretical calculations with a protein-free, simplified model at the B3LYP/6-311G* level of density functional theory. Activation energies for the hydrogen abstraction and recombination with 1,2-propanediol as substrate are 9.0 and 15.1 kcal/mol, respectively, and essentially not affected by coordination of the substrate and the radical intermediate to K+. Since these energies can be considered to be supplied by the substrate-binding energy, the computational results with this simplified model indicate that the hydrogen abstraction and recombination in the coenzyme B(12)-dependent diol dehydratase reaction are energetically feasible.

Direct participation of potassium ion in the catalysis of coenzyme B(12)-dependent diol dehydratase.

The direct ion-dipolar interactions between potassium ion (K(+)) and the two hydroxyl groups of the substrate are the most striking feature of the crystal structure of coenzyme B(12)-dependent diol dehydratase. We carried out density-functional-theory computations to determine whether K(+) can assist the 1,2-shift of the hydroxyl group in the substrate-derived radical. Between a stepwise abstraction/recombination reaction proceeding via a direct hydroxide abstraction by K(+) and a concerted hydroxyl group migration assisted by K(+), only a transition state for the latter concerted mechanism was found from our computations. The barrier height for the transition state from the complexed radical decreases by only 2.3 kcal/mol upon coordination of the migrating hydroxyl group to K(+), which corresponds to a 42-fold rate acceleration at 37 degrees C. The net binding energy upon replacement of the K(+)-bound water for substrate was calculated to be 10.7 kcal/mol. It can be considered that such a large binding energy is at least partly used for the substrate-induced conformational changes in the enzyme that trigger the homolytic cleavage of the Co-C bond of the coenzyme and the subsequent catalysis by a radical mechanism. We propose here a new mechanism for diol dehydratase in which K(+) plays a direct role in the catalysis.

Structural characterization of natural and recombinant human granulocyte colony-stimulating factors.

Granulocyte colony-stimulating factor (G-CSF) is a glycoprotein which stimulates predominantly neutrophilic granulocyte colony formation in mammals. Natural human G-CSF (hG-CSF) and recombinant hG-CSF produced in Chinese hamster ovary (CHO) cells transfected with the cDNA clone for hG-CSF have been purified to apparent homogeneity for structural and biological comparison. The amino acid sequence of recombinant hG-CSF, composed of 174 amino acid residues, was identical with that of natural hG-CSF and also with the sequence predicted from the cDNA. Both forms of hG-CSF have a free Cys-17 and two intramolecular disulfide linkages, between Cys-36 and Cys-42, and between Cys-64 and Cys-74. The O-glycosylation occurred at Thr-133 in both hG-CSFs. Similar CD spectra were obtained for both hG-CSFs. Additionally, both forms showed almost the same biological activities determined by in vitro colony-forming assay and in vivo assay. It is thus concluded that the recombinant hG-CSF is indistinguishable from its natural counterpart and that the former is valuable for more detailed characterization and clinical use.

Polypeptide and carbohydrate structure of recombinant human interleukin-6 produced in Chinese hamster ovary cells.

A glycosylated form of human interleukin 6 (hIL-6) has been produced in Chinese hamster ovary (CHO) cells transfected with a cDNA clone for human IL-6. Recombinant hIL-6 was purified from a culture supernatant of the transfected CHO cells, and used for structural characterization. The complete amino acid sequence, composed of 185 amino acid residues, was determined and is identical to that predicted from the cDNA sequence. However, a recombinant hIL-6 species lacking two amino acid residues (Ala-Pro) from the N-terminus was also found. Two disulfide bonds are formed, between Cys45 and Cys51 and between Cys74 and Cys84. Recombinant hIL-6 carries one O-linked carbohydrate chain, and Thr139 is fully O-glycosylated. A portion of recombinant hIL-6 protein carries one N-linked sialooligosaccharide chain, and the N-glycosylation occurs at Asn46. The structure of the N-linked sugar chains was estimated by a combination of sugar mapping and glycosidase digestion. The major structure of the N-linked sugar chain predicted was of a fucosylated biantennary or triantennary complex type. Fucosylated triantennary sugar chains with one or two N-acetyllactosaminyl repeats were also found. The structure of the O-linked sugar chain was determined by 500 mHz 1H-NMR to be NeuAc alpha 2-3Gal beta 1-3(NeuAc alpha 2-6) Gal-NAcol.

Physicochemical and biological comparison of recombinant human erythropoietin with human urinary erythropoietin.

Physicochemical and biological properties of recombinant human erythropoietin (rhEPO) were compared with human urinary erythropoietin (uEPO). uEPO and rhEPO were purified to apparent homogeneity from the urine of patients with aplastic anemia and from the conditioned medium of Chinese hamster ovary (CHO) cells transfected with a cDNA clone for human EPO, respectively. The microheterogeneous nature of both factors, observed on isoelectric focusing, is derived from the difference of the number of terminal sialic acid residues bound to the carbohydrate chains of the EPO molecule. The primary structure of rhEPO, consisting of 165 amino acid residues, was determined, and the C-terminal arginine predicted from the cDNA sequence was confirmed to be missing, as described previously (Recny et al. (1987) J. Biol. Chem. 262, 17156). Three N-glycosylation and one O-glycosylation sites of both factors were determined as Asn24, Asn38, and Asn83 and Ser126, respectively. Two disulfide linkages are located between Cys7 and Cys161, and between Cys29 and Cys33, in both EPOs. Hematogenic potencies of rhEPO and uEPO compared in normal and in partially nephrectomized rats were approximately the same. Both factors also stimulated the colony formation of CFU-E, BFU-E, and CFU-Meg in a dose-dependent manner. From these results, it is concluded that rhEPO produced in CHO cells transfected with cDNA clone for human EPO is indistinguishable from uEPO physicochemically and biologically, and is valuable for further research and for clinical use.

Citron, a Rho target that affects contractility during cytokinesis.

The small GTPase Rho, which regulates cell shape, is thought to contribute to cytokinesis. Recently, Citron was characterized as a Rho target. This large protein contains a Ser/Thr kinase domain related to that of ROCK, another Rho effector. Both endogenous Citron and recombinant Citron localize to the cleavage furrow in dividing cells and to the midbody in post-mitotic cells. Moreover, overexpression of Citron deleted from its C-terminal sequence caused abnormal contractions specifically during cytokinesis, resulting in the formation of multinucleated cells. Cell shape, F-actin, intermediate filaments, and microtubules appeared essentially normal in these cells during interphase. Thus, Citron is a Rho effector that appears to function during cytokinesis, modulating its contractile process. In brain, however, Citron is highly expressed in a subset of neurons as a brain-specific isoform that lacks a kinase domain, Citron-N. This protein accumulates in synapses and associates to the NMDA receptor via interaction with the adaptor protein PSD95, suggesting that the function of Citron is specialized in the neurons.

Effects of repeated hand instrumentation on the marginal portion of a cast gold crown.

A study was conducted to evaluate the effects of repeated hand instrumentation on the marginal portion of a cast gold crown. Seven extracted periodontally diseased premolars were used. The finishing line of the preparation was placed on the root surface and then the crown was cast and cemented in the usual manner. One proximal surface of each sample was divided into 2 areas: root planing (RP) area and RP plus polishing (RPP) area. The marginal portion of the crown was measured to give a baseline value using a surface roughness- and profile-analyzing system. Then, the marginal portion was painted with a waterproof pen. RP was performed to remove paint in the RP area with the curets. In the RPP area, RP followed by polishing was done by silicone polishing points and a rubber cup with polishing paste. The relevant procedures and measurements were repeated 3 times in each area. Changes in the sample roughness and profile were evaluated and compared between the 2 techniques. The results showed that repeated instrumentation altered the surface of the marginal portion of the cast gold crown, resulting in increased roughness in both areas (P < 0.01). However, the roughness of the RPP area was considerably restored to the baseline value by polishing after RP. Therefore, it is suggested that polishing after RP smoothes the marginal portion of the cast gold crowns and appears to be an efficient prophylactic system.

Structure of G-CSF: significance of the sugar chain.

1. The carbohydrate chain protects rhG-CSF from polymerization and/or conformational alterations associated with physicochemical changes, elevation of pH or temperature fluctuations. 2. The carbohydrate chain of rhG-CSF prevents loss of its biological activity in normal human serum by inhibiting proteinase activity. 3. These facts indicate that the carbohydrate chain of rhG-CSF has a markedly important role in maintaining the stability of the protein itself as well as in effecting the exertion of its biological activity.

Patients' preference for olanzapine orodispersible tablet compared with conventional oral tablet in a multinational, randomized, crossover study.

OBJECTIVES: The aim of this study was to compare patients' preference for olanzapine orodispersible tablet (ODT) with oral conventional tablet (OCT). METHODS: A 12-week randomized, crossover, multinational, open-label study was conducted to estimate the proportion of patients preferring ODT or OCT. Outpatients with stable schizophrenia on OCT monotherapy were randomly assigned 1:1 to ODT or OCT. Compliance and drug attitude were measured using the Drug Attitude Inventory (DAI-10) and Medication Adherence Form (MAF) scales; tolerability and safety by Association for Methodology and Documentation in Psychiatry (AMDP-5) questionnaire and adverse event summary. RESULTS: A total of 175 patients answered a preference question: 106 (61%) preferred ODT and 48 (27%) preferred OCT (P<0.001 adjusted for treatment sequence); 21 (12%) expressed no preference. There was no significant change in DAI-10 with either formulation. MAF was above 75% in 94% vs. 93% of patients on ODC and OCT, respectively. Compliance as measured by tablet count was above 98% on both formulations. The adverse event profiles did not differ between formulations. Mean weight increase over 6 weeks on ODT was 0.8 kg and on OCT was 0.6 kg. CONCLUSIONS: Given the importance of patients' preference for treatment planning and success, the ODT formulation should be routinely considered as a treatment option.

Sporeless mutants of Bacillus thuringiensis. II. mutants derived from var. thuringiensis and var. sotto.

Three sporeless mutants of Bacillus thuringiensis, 2 derived from var. thuringiensis and 1 from var. sotto were selected after mutagenic treatment. They were completely lacking in ability to form spores, yet maintained intact insecticidal activity.