Inactive Preparation Research Articles

Myths and Facts about Placebo

Many patients, physicians, and sometimes even academics have questionable perceptions of placebo and the so-called placebo effect, Many believe that placebo have its own effects. Although psychological aspects, namely expectations of patients or the persuasive power of the physicians, might sometimes be substantial, such aspects may have little or even no relevance in other situation where placebo control is essential nevertheless. Even in settings where psychological effects should be envisaged, their extent is usually highly variable, indicating that other factors might still exceed the importance of psychological effects. Placebo is defined in US regulations as an inactive preparation designed to resemble the test drug as far as possible. This means that placebo itself cannot be effective. If it would, its correctness is challenged and it should be replaced if still possible. And as placebo is not effective, it can also not have secondary effects, vulgo side effects. Placebo is always used for two reasons: To control bias and to provide the reasonably largest delta, i.e. the difference between two treatments. Placebo should never be interpreted as being able to cause effects.

Climatic design and assimilation of activity spaces (case study on Soheil residential complex in 19th district of Tehran)

Using the principles of climatic design, the most basic inactive energy preparation, would serve as a principled strategy to minimise energy cost and attain thermal comfort in urban spaces. In this study, Soheil residential complex on a plot of 9207 square metres of land in 19th district of Tehran has been studied as a case study to access thermal comfort in an urban space. Also, the main objective of this research is finding a framework and model that can provide all inactive energy preparations serving the goal. So, the climatic data were collected from Mehrabad weather station. Then, the Eco-tect and Climate Consultant 5.2 softwares were used firstly to draw up a general framework of space design or intended sphere in order to analyse the way that the climatic elements affected site design and determine the areas in which masses were located. Finally, the quality of the performance of climatic factors in the intended spheres was tested and assessed in the ENVI-met.4 software through assimilation with a...

Stages of health behavior change and factors associated with physical activity in patients with intermittent claudication

To analyze, in people with intermittent claudication, the frequency of individuals who are in each of stages of health behavior change to practice physical activity, and analyze the association of these stages with the walking capacity. We recruited 150 patients with intermittent claudication treated at a tertiary center, being included those > 30-year-old-individuals and who had ankle-arm index < 0.90. We obtained socio-demographic information, presence of comorbidities and cardiovascular risk factors and stages of health behavior change to practice physical activity through a questionnaire, they being pre-contemplation, contemplation, preparation, action and maintenance. Moreover, the walking capacity was measured in a treadmill test (Gardner protocol). Most individuals were in the maintenance stage (42.7%), however, when the stages of health behavior change were categorized into active (action and maintenance) and inactive (pre-contemplation, contemplation and preparation), 51.3% of the individuals were classified as inactive behavior. There was no association between stages of health behavior change, sociodemographic factors and cardiovascular risk factors. However, patients with intermittent claudication who had lower total walking distance were three times more likely to have inactive behavior. Most patients with intermittent claudication showed an inactive behavior and, in this population, lower walking capacity was associated with this behavior.

The role of Cys108 in Trigonopsis variabilisd-amino acid oxidase examined through chemical oxidation studies and point mutations C108S and C108D

Oxidative modification of Trigonopsis variabilis d-amino acid oxidase in vivo is traceable as the conversion of Cys108 into a stable cysteine sulfinic acid, causing substantial loss of activity and thermostability of the enzyme. To simulate native and modified oxidase each as a microheterogeneity-resistant entity, we replaced Cys108 individually by a serine (C108S) and an aspartate (C108D), and characterized the purified variants with regard to their biochemical and kinetic properties, thermostability, and reactivity towards oxidation by hypochlorite. Tandem MS analysis of tryptic peptides derived from a hypochlorite-treated inactive preparation of recombinant wild-type oxidase showed that Cys108 was converted into cysteine sulfonic acid, mimicking the oxidative modification of native enzyme as isolated. Colorimetric titration of protein thiol groups revealed that in the presence of ammonium benzoate (0.12 mM), the two muteins were not oxidized at cysteines whereas in the wild-type enzyme, one thiol group was derivatized. Each site-directed replacement caused a conformational change in d-amino acid oxidase, detected with an assortment of probes, and resulted in a turnover number for the O 2-dependent reaction with D-Met which in comparison with the corresponding wild-type value was decreased two- and threefold for C108S and C108D, respectively. Kinetic analysis of thermal denaturation at 50 °C was used to measure the relative contributions of partial unfolding and cofactor dissociation to the overall inactivation rate in each of the three enzymes. Unlike wild-type, C108S and C108D released the cofactor in a quasi-irreversible manner and were therefore not stabilized by external FAD against loss of activity. The results support a role of the anionic side chain of Cys108 in the fine-tuning of activity and stability of d-amino acid oxidase, explaining why C108S was a surprisingly poor mimic of the native enzyme.

Modified pyrophosphate-99mTc kit for application in nuclear cardiology

A modified99mTc(Sn)-pyrophosphate (PyP) kit for the application in nuclear cardiology (radioventriculography, angiocardiography, scintigraphy of blood pool) was developed. Each vial contains 12 mg PyP (Na4P2O7), 4 mg SnCl2·2H2O, 2.5 mg gentisic acid and 10 mg NaCl. The reconstitution is performed by dissolving the lyophilized kit in 3 ml 0.9% NaCl. In comparison with the standard pyrophosphate kit for bone scanning and detection of miocardial infarction, it contains an increased amount of Sn(II) so that the molar ratio ligand/reductant is lowered from 25 to 2.5. The radiochemical analyses showed that the radiochemical purity of the labeled kit is high (>90%) during three hours after addition of99mTc-activity. The shelf-life of the inactive freeze-dried preparation is up to four months providing that it is kept in vacuum and at appropriate temperature (2–8°C). The biodistribution studies revealed increased accumulation in blood and low uptake by liver and kidneys. It was concluded that the modified kit performs stable and reproducible properties.

Modulatory role for the serotonergic cerebral giant cells in the feeding system of the snail, Lymnaea. I. Fine wire recording in the intact animal and pharmacology.

1. The role of the paired serotonergic cerebral giant cells (CGCs) in the feeding system of Lymnaea was examined by electrophysiological and pharmacological techniques. 2. The firing characteristics of the CGCs were recorded by fine wires attached to their cell bodies in freely moving intact snails (in vivo recording) and their "physiological" rates of firing determined during feeding and other behaviors. 3. The mean CGC firing rates recorded in vivo varied between 1 and 20 spikes/min but never reached the average rates seen in the isolated CNS (60-120 spikes/min). Maximum rates of firing were seen during bouts of radula biting/rasping movements characteristic of the consummatory phase of feeding (15 +/- 1.69 spikes/min, mean +/- SE, range 7-20 spikes/min), with lower rates seen during locomotion (6.7 +/- 0.75 spikes/min; range 5-9 spikes/min. The cells were rarely active when the animal was quiescent (1.45 +/- 0.91 spikes/min; range 0-2 spikes/min). 4. In vivo recorded CGC firing was phase locked to the feeding movements of the animal, with spikes occurring just before the opening of the mouth, during the protraction phase of the feeding cycle. 5. Evoking firing rates on the CGCs in the isolated preparation similar to those seen in vivo during rasping movements (7-20 spikes/min) did not elicit a fictive feeding pattern in an inactive preparation. Neither did bath application of 10(-9) M serotonin (5-HT; the transmitter of the CGCs). 6. To allow the modulatory role of the CGCs to be examined during patterned activity, the fictive feeding pattern was evoked in the isolated preparation by injecting depolarizing current into a modulatory neuron, the slow oscillator (SO). 7. The tonic firing activity of the CGCs was accurately maintained by current injection in the isolated preparation at rates equivalent to that occurring during feeding, locomotion, and quiescence in the intact snail. This was possible where the CGCs became silent after 1-2 h. Only when the CGCs activity was maintained at a rate (approximately 15 spikes/min) similar to that occurring during rasping, was the SO able to drive a full, high-frequency fictive feeding pattern (15-20 cycles/min). At lower rates of CGC firing, the SO-driven rhythm was either of lower frequency or no rhythm occurred at all (CGCs silent). 8. In many isolated preparations (80%) the CGCs remained active, and it was difficult to maintain specific levels of tonic activity by current injection.(ABSTRACT TRUNCATED AT 400 WORDS)

Pregnancy-Associated Plasma Protein-A-Induced Inhibition of Human Leukocyte Elastase: An Artifact

Pregnancy-associated plasma protein A (PAPP-A), was reported to be an inhibitor in many in vitro systems. Since it was shown that the inhibition of coagulation and complement activity attributed to PAPP-A was in fact due to a contamination by heparin occurring during the purification process, we undertook the present study to see whether the reported PAPP-A-induced inhibition of human leukocyte elastase (HLE) could also be attributed to heparin contamination. PAPP-A was purified from maternal pregnancy EDTA plasma by a method which was previously shown to eliminate contaminating heparin: this preparation was inactive in the HLE assay. But PAPP-A isolated by heparin-Sepharose chromatography, or a PAPP-A-free washing of the heparin-Sepharose column were both inhibitors of HLE. Furthermore the inactive PAPP-A preparation, when incubated with the PAPP-A-free washing of the heparin-Sepharose column, yielded a high molecular weight preparation which inhibited HLE. It is concluded that PAPP-A is not an inhibitor of HLE and that the inhibition of HLE previously attributed to PAPP-A was due to contaminating heparin.

African horse sickness: An overview

A short overview of African horse sickness disease was given. Results were described of a research project that was carried out in Egypt cooperatively between Egyptian and American scientists. The virus was isolated from the blood of street dogs and camels from southern Egypt close to the Sudan border. Camel tick Hyaloma dromadarrii and dog tick Rhipicephalus sanguineus sanguineus harbored and transmitted the virus. Optimal reaction times were determined for agar diffusion precipitin and direct fluorescent antibody tests. Complement fixation and agar precipitin diffusion do not differentiate virus serotypes. Formalin, acetylethyleneimine, and binary ethyleneimine respectively inactivated the virus. Inactive preparation with adjuvant are promising vaccines. The virus was inactivated by irradiation. Plant antiviral extracts were investigated. Latent AHS virus in dogs was activated, producing viremia incident to the dogs being bitten by Culicoides . An antimoulting insecticide was more effective on early developmental stages of Culicoides pipiens .

Effect of Anti-Thrombospondin Antibodies on the Hemagglutination Activities of the Endogenous Platelet Lectin and Thrombospondin

The proposal that thrombospondin is the endogenous platelet lectin was evaluated using antisera and monoclonal antibodies to thrombospondin. The platelet-bound hemagglutinin activity of human platelets stimulated with A23187 was inhibited by rabbit anti-thrombospondin sera and by a monoclonal anti-thrombospondin IgG. A second monoclonal IgG did not inhibit platelet-bound agglutinin activity. Preparations of purified platelet thrombospondin differed in their hemagglutination activities. The hemagglutination activity of an active preparation of thrombospondin was inhibited by the monoclonal antibody that inhibited platelet-bound lectin activity. The hemagglutination activity of an almost inactive preparation of thrombospondin was enhanced by the anti-thrombospondin monoclonal antibody that did not block platelet-bound lectin activity. The results demonstrate that expression of the platelet-bound form of the endogenous lectin is thrombospondin-dependent and suggest that thrombospondin must become part of a larger complex, either by binding to the platelet surface or by becoming aggregated in solution, before hemagglutination activity can be expressed.

Characterization studies of inactive renin from human kidney and from several plasma sources.

Plasma of normal pregnant women and plasma of some patients with diabetic nephropathy contain increased quantities of an inactive form of renin compared with normal plasma. In order to evaluate the significance of the increased levels of inactive renin in these conditions, we isolated and compared inactive renin from several plasma sources and from human kidney. Renal inactive renin and inactive renin from normal plasma and plasma of patients with diabetic nephropathy and plasma of pregnant women displayed reversible acid activation and similar binding characteristics to cibacron-blue. Using cibacron-blue affinity chromatography, we obtained a totally inactive renin preparation from renal cortex and from plasma of subjects from each of the clinical states. Gel filtration of these preparations and detection of inactive renin using a modified acid activation technique indicated that human inactive renin exists in at least two forms with the following apparent molecular weights: 56,300 +/- 1,500 and 49,200 +/- 1,000 for renal inactive renin, 58,000 and 49,000 for inactive renin in normal plasma, 58,500 and 52,000 for inactive renin in diabetic plasma, 57,000 and 49,000 for inactive renin in pregnancy plasma. These studies suggest that: 1) the kidney is a major source of circulating inactive renin in man, 2) inactive renin from normal, diabetic and pregnancy plasma is similar, and 3) human inactive renin is heterogeneous.

Evidence for a completely inactive renin zymogen in the kidney by affinity chromatographic isolation

Summary A complete inactive renin was isolated from hog kidney extract by hydrophobic interaction chromatography on Octyl-Sepharose followed by affinity chromatography on pestatin-aminohexyl Sepharose. The inactive renin was activated by trypsin. It has a molecular weight of 53,000 ± 2,000 by gel filtration on Sephadex G-100. The active renin generated from this inactive renin by trypsin treatment has the molecular weight of 41,000 ± 1,000. Treatments of the inactive renin with dithiothreitol, NaCl, SDS or an acidic buffer did not cause activation indicating that the inactive preparation is not a renin-inhibitor complex.

The distribution of biologically active and inactive radioiodinated human calcitonin in the rat.

The anatomical distribution of a biologically active and biologically inactive preparation of radioiodinated human calcitonin (hCT) was studied in rats. The former was prepared by electrolysis and the latter by chloramine-T oxidation. When administered iv, the distribution of the two species of [125I]hCT was nearly identical. The kidney was the principle organ of distribution for the hormone while less was present in liver, intestine, or bone. In vitro, kidney homogenates rapidly degraded [125I]hCT. This degradation was partially blocked by trypsin inhibitor and fully blocked by ACTH at both 4 and 37 C. These studies demonstrate the importance of kidney in the metabolism of calcitonin and suggest that the in vivo distribution of biologically active and inert species of the hormone may be indistinguishable.

The role of renal adenosine 3′,5′-monophosphate in the control of erythropoietin production

A regulatory role for adenosine 3′,5′-monophosphate (cyclic AMP) in the production of the renal hormone erythropoietin following erythropoietic stimulation with cobaltous chloride hexahydrate is proposed. Studies in rats reveal a temporal relationship between renal cyclic AMP levels and plasma titers of erythropoietin. In addition, cobalt increases the activity of an erythropoietin-generating enzyme (renal erythropoietic factor) with maximal enzyme activity occurring after the rise in cyclic AMP levels but before the increase in erythropoietin titers.This increase in renal cyclic AMP is localized to the renal cortex. Cobalt stimulates renal cortical adenylate cyclase but has no effect on renal cyclic nucleotide phosphodiesterase. The addition of cyclic AMP (3 × 10−6M) and a partially purified cyclic AMP-dependent protein kinase from rat kidney to an inactive preparation of renal erythropoietic factor increases the ability of renal erythropoietic factor to generate erythropoietin.Data from the polycythemic mouse assay, a bioassay used to quantitate erythropoietic activity of test substances, indicate that dibutyryl cyclic AMP is erythropoietically active with respect to its ability to increase radioactive-labelled iron (59Fe) incorporation into heme of newly formed red blood cells. Theophylline, which by itself is erythropoietically inactive, potentiated the erythropoietic effect of cobalt in polycythemic mice.These results suggest that cyclic AMP plays a significant role in the renal production of erythropoietin following cobalt administration. It is postulated that cobalt stimulates renal cortical adenylate cyclase, thus increasing renal cyclic AMP levels. Cyclic AMP then activates a protein kinase which subsequently stimulates renal erythropoietic factor to generate erythropoietin. A similar cyclic AMP mechanism may be operative after erythropoietic stimulation by exposure to hypoxia or prostaglandin treatment.

Quantitation of acetylcholinesterase and acetylcholine- binding sites in excitable membrane fragments from electric eel

1. 1. Titration of membrane-bound acetylcholinesterase with a specific active-site reversible inhibitor 3-hydroxyphenyltrimethylammonium iodide allows a quantitative determination of active site concentration of the enzyme in the membrane fragments from electroplax of the electric eel. 2. 2. Extraction of acetylcholinesterase activity from membrane fragments by I M NaCl results in an enzymatically inactive preparation which is still capable of binding acetylcholine. 3. 3. The quantity of acetylcholine binding sites is about 16–20% of the catalytic sites of the enzyme in the electroplax membrane fragments, in agreement with results obtained with monocellular electroplax preparation. The K D for the acetylcholine-receptor complex is 1.8· 10 −7 M.

Prostaglandin-like substances: initiation and maintenance of rabbit testicular contraction in vitro.

SummaryThe spontaneous contractions that occur in rabbit testes in vitro were abolished by changing the bathing medium. Adding the initial medium from an active to an inactive preparation induced activity in the inactive testis. Extraction of the initial medium at pH 3 into ethyl acetate, a system used for recovering prostagandins, yielded a residue that restored the activity. Acetylcholine and serotonin had little effect on the inactive preparation nor could a smooth-muscle active substance be extracted from these preparations. Oxytocin stimulated neither the active nor the inactive testis appreciably. Epinephrine increased tonus in both inactive or active preparations, but did not induce rhythmic contractions. Prostaglandins of the E-type were stimulatory at low, but inhibitory at higher concentrations. A myogenic agent with prostaglandin-like properties was present only in the bathing media from active preparations and appeared to determine the response that was obtained by treatment with exogenous pro...

Multicentre, Short-term Therapeutic Trial of Disodium Cromoglycate, With and Without Prednisone, in Adults with Asthma

A total of 117 asthmatic patients were studied in a double-blind, cross-over trial with half-periods of three weeks in which disodium cromoglycate (D.S.C.G.) was compared with an inactive preparation. D.S.C.G. improved spirometric measurements, at a very high level of statistical significance, more than did placebo. The mean advantage of D.S.C.G. was less than 10% after three weeks. Symptomatic improvement by D.S.C.G. was small though significant; it was not correlated with spirometric improvement. A history of chronic cough was the only clinical characteristic associated with a better-than-average spirometric response to D.S.C.G.

Constitutive Inorganic Pyrophosphatase of Escherichia coli: V. RECONSTITUTION OF NATIVE ENZYME PARTICLES FROM SUBUNIT POLYPEPTIDE CHAINS

Abstract In concentrated guanidine hydrochloride solutions Escherichia coli pyrophosphatase dissociated to 6 randomly coiled polypeptide chains of molecular weight 20,000. When the guanidine salt was removed by dialysis, enzymatic activity reappeared, provided that a sulfhydryl compound was present during the renaturation process. The reconstituted enzyme had a molecular weight and sedimentation-diffusion properties which were indistinguishable from those of native protein. Optical, electrophoretic, and catalytic properties of reconstituted and native enzymes were also identical. If the protein cysteine groups (2 per polypeptide subunit) were allowed to form disulfide bonds, inactive, compact globular structures, about half the size of native protein, were formed upon removal of guanidine hydrochloride. Alkylation of the cysteines with N-ethylmaleimide and subsequent removal of the guanidine salt also yielded an inactive preparation; this appeared to consist of large, asymmetric aggregates of partially unfolded chains.

Preparation and In Vitro Evaluation of Antibiotic Drug Immunoconjugates

The aim of the present study was to develop drug immunoconjugates (DIC) of antibiotics like Ampicillin trihydrate, Amoxycillin trihydrate, Streptomycin sulphate, Rifampicin and doxorubicin with Anti-D serum using different coupling methods. Protein content in Anti- D serum was estimated by Lowry’s method. The conjugates were purified from unbound drug by dialysis. The drugs to antibody ratios were estimated by determining the quantity of antibacterial drugs coupled during coupling reactions by turbidimetric or colorimetric assay. The conjugates were tested for both antibody and antibacterial activities by simple in vitro methods. Slide agglutination test was used to know the retention of activity of the antibody qualitatively. The antibacterial activity was observed by reduction in turbidity due to growth inhibition. When glutaraldehyde method was used to couple the drugs the resulted DICs of Ampicillin trihydrate, Amoxycillin trihydrate and Streptomycin sulphate retained both antibody and antibacterial activities, while a direct linkage of Streptomycin sulphate to antibody resulted in the loss of antibacterial activity. Rifampicin was also linked to antibody by two methods. Upon direct linkage of Rifampicin to antibody retained both activities, but when linked by dicyclohexyl carbodiimide method, an antibody inactive preparation was resulted. Doxorubicin an anticancer antibiotic was also linked to antibody by glutaraldehyde method which retained antibody activity. Taken together, the results of our studies would be useful in the design of drug immunoconjugates with antibiotic drugs to treat specific infections in chemotherapy.

Cytochrome oxidase apoprotein in "petite" mutant yeast mitochondria. Reconstitution of cytochrome oxidase by combining apoprotein with cytohemin.

1 Cytochrome oxidase extracted from mitochondria of wild-type baker's yeast by means of bile salts and fractionated with ammonium sulfate had a specific activity about 18 times that of the mitochondria. When mitochondria of a cytoplasmic “petite” mutant were subjected to the same procedure, the resulting preparation exhibited virtually no cytochrome oxidase activity. 2 Gel filtration resolved the active enzyme preparation into five protein peaks, the first of which contained copper and cytohemin and showed cytochrome oxidase activity. Bio-Gel P-300 filtration of the inactive preparation from mitochondria of the “petite” mutant likewise gave five peaks. The first peak, however, while containing copper, was free from cytohemin and devoid of cytochrome oxidase activity. 3 When the material recovered from the first gel filtration peak of the active cytochrome oxidase preparation was subjected to disc electrophoresis on polyacrylamide, there appeared two protein bands. Two bands characterized by the same mobilities were also observed upon disc electrophoresis of the first peak material derived from mutant mitochondria. 4 The evidence for the presence in “petite” mutant mitochondria of a protein which is identical with or closely similar to the apoprotein of wild-type yeast cytochrome oxidase was confirmed by the finding that the incubation of submitochondrial particles of the “petite” mutant with cytohemin produced cytochrome oxidase activity. The enzymatic activity obtained was about 30% of that encountered in submitochondrial particles of wild-type yeast.

Application of radioisotopes in wear and lubrication studies. (in German) : K. Wagner, Isotopenpraxis, 4 (1968) 85–94; 4 figs., 1 table, 341 refs.

A regulatory role for adenosine 3′,5′-monophosphate (cyclic AMP) in the production of the renal hormone erythropoietin following erythropoietic stimulation with cobaltous chloride hexahydrate is proposed. Studies in rats reveal a temporal relationship between renal cyclic AMP levels and plasma titers of erythropoietin. In addition, cobalt increases the activity of an erythropoietin-generating enzyme (renal erythropoietic factor) with maximal enzyme activity occurring after the rise in cyclic AMP levels but before the increase in erythropoietin titers.This increase in renal cyclic AMP is localized to the renal cortex. Cobalt stimulates renal cortical adenylate cyclase but has no effect on renal cyclic nucleotide phosphodiesterase. The addition of cyclic AMP (3 × 10−6M) and a partially purified cyclic AMP-dependent protein kinase from rat kidney to an inactive preparation of renal erythropoietic factor increases the ability of renal erythropoietic factor to generate erythropoietin.Data from the polycythemic mouse assay, a bioassay used to quantitate erythropoietic activity of test substances, indicate that dibutyryl cyclic AMP is erythropoietically active with respect to its ability to increase radioactive-labelled iron (59Fe) incorporation into heme of newly formed red blood cells. Theophylline, which by itself is erythropoietically inactive, potentiated the erythropoietic effect of cobalt in polycythemic mice.These results suggest that cyclic AMP plays a significant role in the renal production of erythropoietin following cobalt administration. It is postulated that cobalt stimulates renal cortical adenylate cyclase, thus increasing renal cyclic AMP levels. Cyclic AMP then activates a protein kinase which subsequently stimulates renal erythropoietic factor to generate erythropoietin. A similar cyclic AMP mechanism may be operative after erythropoietic stimulation by exposure to hypoxia or prostaglandin treatment.