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At least 469 records · Page 26Linked to original sources
PubMed2002

Impact of esophageal acid exposure on the endoscopic reversal of Barrett's esophagus.

OBJECTIVE: Normalization of esophageal acid exposure had been assumed to be necessary for the reversal of Barrett's esophagus with endoscopic therapy. This assumption is examined by evaluating the esophageal pH in a group of patients undergoing reversal therapy with fixed high-dose proton pump inhibitor therapy and endoscopic multipolar electrocoagulation (MPEC). METHODS: Patients with Barrett's esophagus of 2-6 cm in length were treated with omeprazole (40 mg b.i.d.). They underwent 24-h esophageal pH monitoring 5 cm above the upper margin of the lower esophageal sphincter determined by a water-perfused catheter. They then underwent MPEC therapy to an endpoint of elimination of Barrett's (reversal) by both endoscopy and biopsy 6 months after the last MPEC session or failure to achieve visual (endoscopic) reversal after six treatment sessions. RESULTS: Twenty patients had 24-h pH testing and reached a reversal endpoint. Three patients had abnormal pH tests, two total and supine and one supine only. These patients had documented reversal. The remaining 17 patients had normal pH testing but five failed reversal therapy. CONCLUSION: Barrett's esophagus can be completely reversed with endoscopic therapy despite abnormal esophageal acid exposure. Also, patients can fail reversal even with normal esophageal acid exposure. The necessary reduction of esophageal acid exposure for reversal therapy has yet to be defined.

Adult↗
PubMed1968

Gelatin-induced reversion of protoplasts of Bacillus subtilis to the bacillary form: electron-microscopic and physical study.

Protoplasts of Bacillus subtilis plated on SD medium form L colonies in quantitative yield and propagate in the L form indefinitely. L bodies or protoplasts placed in 25% gelatin medium form bacillary colonies. Details of the reversion of naked bodies to the walled form are reported. In 25% gelatin medium, reversion begins earlier (about 50% reversion in 4 hr) than the multiplication of bacilli. Thus, virtually all the observed bacillary forms are themselves revertants and not the offspring of a few growing clones. The optimal temperature for reversion is 26 C in 25% gelatin. When cells reverting at 26 C are warmed to 40 C for 3 min, reversion is delayed markedly, whereas viability is unaffected. For electron microscopy, a dense protoplast inoculum was placed on a gelatin surface, incubated, and then fixed in situ. There was no multiplication, but crowding delayed reversion markedly. Successive events of reversion are as follows. The loose nucleoid of the protoplasts condenses in response to the gelatin medium and condenses further and further as reversion proceeds. A thin coat of wall develops around the bodies of various sizes and shapes and then increases uniformly in thickness until a wall of normal aspect is formed. Rod-shaped cells grow out from these bodies-sometimes in several directions at once. A few mesosomes begin to appear only after a thin coat of wall has been formed. These are dense, atypical structures compartmented by membranes. They are located at the cell periphery and do not seem to be in contact with the nucleoids. Quantitative estimates showed that only 20 to 25% of revertant cells or cells grown on gelatin contain even a single mesosome. The others have no mesosome at all. Mesosomes thus do not appear to play a significant role in reversion, and normal mesosome functions must presumably be performed elsewhere in the cell in gelatin-grown bacilli. The role of cell wall, its synthesis, and its chemical nature in successive steps in reversion are discussed.

Bacillus subtilis↗
PubMed2002

The tRNA primer activation signal in the human immunodeficiency virus type 1 genome is important for initiation and processive elongation of reverse transcription.

Human immunodeficiency virus type 1 (HIV-1) reverse transcription is primed by the cellular tRNA(3)(Lys) molecule, which binds, with its 3"-terminal 18 nucleotides (nt), to a complementary sequence in the viral genome, the primer-binding site (PBS). Besides PBS-anti-PBS pairing, additional interactions between viral RNA sequences and the tRNA primer are thought to regulate the process of reverse transcription. We previously identified a novel 8-nt sequence motif in the U5 region of the HIV-1 RNA genome that is critical for tRNA(3)(Lys)-mediated initiation of reverse transcription in vitro. This motif activates initiation from the natural tRNA(3)(Lys) primer but is not involved in tRNA placement and was therefore termed primer activation signal (PAS). It was proposed that the PAS interacts with the anti-PAS motif in the TphiC arm of tRNA(3)(Lys). In this study, we analyzed several PAS-mutated viruses and performed reverse transcription assays with virion-extracted RNA-tRNA complexes. Mutation of the PAS reduced the efficiency of tRNA-primed reverse transcription. In contrast, mutations in the opposing leader sequence that trigger release of the PAS from base pairing stimulated reverse transcription. These results are similar to the reverse transcription effects observed in vitro. We also selected revertant viruses that partially overcome the reverse transcription defect of the PAS deletion mutant. Remarkably, all revertants acquired a single nucleotide substitution that does not restore the PAS sequence but that stimulates elongation of reverse transcription. These combined results indicate that the additional PAS-anti-PAS interaction is needed to assemble an initiation-competent and processive reverse transcription complex.

5' Untranslated Regions↗
PubMed2003

Human immunodeficiency virus type 1 protease regulation of tat activity is essential for efficient reverse transcription and replication.

The human immunodeficiency virus type 1 (HIV-1) Tat protein enhances reverse transcription, but it is not known whether Tat acts directly on the reverse transcription complex or through indirect mechanisms. Since processing of Tat by HIV protease (PR) might mask its presence and, at least in part, explain this lack of data, we asked whether Tat can be cleaved by PR. We used a rabbit reticulocyte lysate (RRL) system to make Tat and PR. HIV-1 PR is expressed as a Gag-Pol fusion protein, and a PR-inactivated Gag-Pol is also expressed as a control. We showed that Tat is specifically cleaved in the presence of PR, producing a protein of approximately 5 kDa. This result suggested that the cleavage site was located in or near the Tat basic domain (amino acids 49 to 57), which we have previously shown to be important in reverse transcription. We created a panel of alanine-scanning mutations from amino acids 45 to 54 in Tat and evaluated functional parameters, including transactivation, reverse transcription, and cleavage by HIV-1 PR. We showed that amino acids 49 to 52 (RKKR) are absolutely required for Tat function in reverse transcription, that mutation of this domain blocks cleavage by HIV-1 PR, and that other pairwise mutations in this region modulate reverse transcription and proteolysis in strikingly similar degrees. Mutation of Tat Y47G48 to AA also down-regulated Tat-stimulated reverse transcription but had little effect on transactivation or proteolysis by HIV PR, suggesting that Y47 is critical for reverse transcription. We altered the tat gene of the laboratory strain NL4-3 to Y47D and Y47N so that overlapping reading frames were not affected and showed that Y47D greatly diminished virus replication and conveyed a reverse transcription defect. We hypothesize that a novel, cleaved form of Tat is present in the virion and that it requires Y47 for its role in support of efficient reverse transcription.

Animals↗
PubMed1975

Determinants of reversible asynergy. The native coronary circulation.

To evaluate the influence of the native coronary circulation on the reversibility of asynergy, ventriculograms before and after sublingual nitroglycerin were performed in 51 patients with coronary artery disease and asynergy. The severity of stenotic lesions and caliber of the distal coronary vessels were determined by comparison with external catheter tip diameter corrected for magnification. Of 42 asynergic zones associated with larger than or equal to 90% proximal coronary occlusion, 27 (64%) were akinetic or dyskinetic while only 11 of 38 zones (29%) with less than 90% occlusion showed akinesis (P less than 0.005). Twenty-six of the 38 asynergic zones (69%) with less than 90% occlusion were reversible in contrast to 19 of the 42 zones (45%) with larger than or equal to 90% occlusion (P less than 0.05). Coronary collaterals were observed in 23 of 42 (55%) zones with larger than or equal to 90% occlusion in contrast to only 11 of 38 zones (29%) with less than 90% occlusion (P less than 0.05). Of the zones with both greater than or equal to 90% occlusion and collaterals, 74% were reversible, in contrast to only 11% without collaterals (P less than 0.001). Of the asynergic zones without collaterals, 63% with less than 90% occlusion were reversible in contrast to only 11% with larger than or equal to 90% occlusion (P less than 0.001). Pathologic Q waves were associated with 24 of 42 zones (57%) with larger than or equal to 90% occlusion compared to only nine of the 38 zones (24%) with less than 90% occlusion (P less than 0.01). The presence of Q waves was associated with a significant decrease in the incidence of reversibility regardless of the degree of coronary occlusion. Excluding the asynergic zones with either collaterals or Q waves, 79% with less than 90% occlusion were reversible in contrast to only 37% with larger than or equal to 90% coronary occlusion (P less than 0.05). In contrast, the caliber of the distal vessel could not be correlated with either the severity of asynergy or the presence of collaterals and was similar in both reversible and irreversible asynergic zones. In summary, larger than or equal to 90% proximal stenosis is associated with severe asynergy which is less likely to be reversible compared to asynergy associated with less than 90% coronary occlusion. In the presence of larger than or equal to 90% occlusion, coronary collaterals are associated with a significantly higher incidence of reversible asynergy and thus appear to serve a protective function. However, the caliber of the distal vessel per se does not effect the severity or reversibility of asynergy.

Cardiac Output↗
PubMed2003

Modeling reverse-phi motion-selective neurons in cortex: double synaptic-veto mechanism.

Reverse-phi motion is the illusory reversal of perceived direction of movement when the stimulus contrast is reversed in successive frames. Livingstone, Tsao, and Conway (2000) showed that direction-selective cells in striate cortex of the alert macaque monkey showed reversed excitatory and inhibitory regions when two different contrast bars were flashed sequentially during a two-bar interaction analysis. While correlation or motion energy models predict the reverse-phi response, it is unclear how neurons can accomplish this. We carried out detailed biophysical simulations of a direction-selective cell model implementing a synaptic shunting scheme. Our results suggest that a simple synaptic-veto mechanism with strong direction selectivity for normal motion cannot account for the observed reverse-phi motion effect. Given the nature of reverse-phi motion, a direct interaction between the ON and OFF pathway, missing in the original shunting-inhibition model, it is essential to account for the reversal of response. We here propose a double synaptic-veto mechanism in which ON excitatory synapses are gated by both delayed ON inhibition at their null side and delayed OFF inhibition at their preferred side. The converse applies to OFF excitatory synapses. Mapping this scheme onto the dendrites of a direction-selective neuron permits the model to respond best to normal motion in its preferred direction and to reverse-phi motion in its null direction. Two-bar interaction maps showed reversed excitation and inhibition regions when two different contrast bars are presented.

Animals↗
PubMed2003

Pursuit of the ineffable: perceptual and motor reversals during the tracking of apparent motion.

Pursuit can be guided by perceived rather than physical motion, but the temporal relationship between motion perception and pursuit is unknown. We used an apparent motion stimulus consisting of a horizontal row of evenly spaced Kanizsa illusory squares (1.44 deg2): the illusory contours appeared at the midpoints of the illusory squares presented in the previous frame, producing bi-directional apparent motion of the illusory contours (21.5 deg/s) that could be reversed at will. We measured eye movements in five subjects asked to (1) track the motion of the illusory squares, and (2) reverse the perceived direction while continuing to track the squares. We measured the timing of the voluntary perceptual reversals and compared this to the time course of the reversal in tracking direction. We found that subjects could smoothly track the apparent motion of illusory squares and also produce saccade-free reversals in pursuit velocity. The time course of these motor reversals closely followed the measurements of the perceptual reversal and, on average, the perceptual reversals preceded the pursuit reversals by 53 ms, a delay shorter than when the perceptual reversal was visually guided. Smooth pursuit and the perception of motion direction were in temporal register and highly correlated, suggesting that pursuit can provide a real-time readout for the state of motion perception.

Adult↗
PubMed1974

Reversal of ouabain and acetyl strophanthidin effects in normal and failing cardiac muscle by specific antibody.

Isolated cat right ventricular papillary muscles were used to study the effects of antibodies with high affinity for ouabain and acetyl strophanthidin on myocardium exposed to these cardioactive steroids. Antibodies with average intrinsic affinity constants for ouabain and acetyl strophanthidin of the order of 10(8) M(-1) were raised in rabbits challenged by repeated injection of a conjugate of ouabain covalently linked to a poly D,L-alanyl derivative of human serum albumin. Effects were assessed in terms of time-course and extent of inotropy reversal, influence of experimentally induced ventricular failure, digitalis-antibody concentration relations, influence of digitalis-antibody complex on response to additionally added digitalis, and relation of antibody effects on digitalis-induced automaticity and contracture to reversal of inotropy. Specific antibody (but not control antibody) in 1.1-1.5-fold molar excess over cardioactive steroid concentrations blocked positive inotropic effects of ouabain and acetyl strophanthidin, and gradually reversed established contractile effects of these agents with a mean time for half-reversal of ouabain-induced inotropy of 124+/-6 (SEM) min and 37+/-3 min for half-reversal of acetyl strophanthidin-induced inotropy. Papillary muscles from cats with right ventricular failure induced by chronic pulmonary artery constriction responded similarly. Both normal and failing muscles returned to but not below levels of contractility existing before cardioactive steroid exposure, and time for half-reversal of inotropy by antibody was significantly shorter than time for half-reversal after removal of ouabain or acetyl strophanthidin by muscle bath washout alone. Presence of ouabain- or acetyl strophanthidin-antibody complex did not alter the myocardial contractile response to subsequently added cardioactive steroids. Spontaneous automaticity occurring as a toxic response to ouabain or acetyl strophanthidin in eight muscles was rapidly reversed by specific antibody at a time when positive inotropic effects were still fully manifest. Early contracture was also reversed by specific antibody. These studies provide further support for the concept that cardiac glycoside-specific antibodies are capable of reversing established cellular effects of cardioactive steroids.

Animals↗
PubMedDate not supplied

Evidence of lower oxidative stress in the air spaces of patients with reversible COPD.

The mechanism responsible for the reversibility of airflow limitation in stable chronic obstructive pulmonary disease (COPD) patients is unknown. The aim of this study is to assess the relationship between the reversibility of airflow limitation, the redox balance and the inflammatory cells in the sputum of patients with stable COPD. For this purpose we examined 15 normal healthy control subjects and 20 nonatopic COPD patients. The COPD patients were divided into two groups: reversible COPD (increase in FEV1> 200 ml and/or > or =12> or = after 200 microg of inhaled salbutamol) or non-reversible COPD. GSH, GSSG were measured in induced sputum and blood. Protein carbonyls were evaluated by WB in sputum and IL-4 and IL-6 and TNF-alpha in plasma and sputum. GSH oxidation and protein oxidation were lower in reversible COPD patients than in those with no reversibility. The sputum eosinophil count was significantly higher in the reversible group than in the non-reversible group, and IL-4 concentration was higher in the same patients both in sputum and in plasma. In contrast, IL-6 and TNF-alpha were increased in non-reversible COPD patients in both biological samples. We conclude that airflow reversibility in COPD patients is associated with airway oxidative stress and activation of eosinophil inflammatory pattern in sputum and blood, suggesting that these patients could respond to specific pharmacological treatment.

Adult↗
PubMed1999

[Reverse redistribution of technetium-99m-tetrofosmin after acute coronary revascularization in patients with myocardial infarction].

This study qualified the severity and localization of reverse redistribution of technetium-99 m (Tc)-tetrofosmin rest imaging. Both Tc-tetrofosmin and thallium-201 (Tl) rest imaging with early images and delayed images were obtained in the subacute phase of myocardial infarction in 21 patients with first anterior myocardial infarction and with successful transluminal angioplasty (including stenting). Relative myocardial uptake (%uptake), degree of reverse redistribution (%), and washout rate (%) were evaluated quantitatively in 6 left ventricular segments (inferoseptal, anteroseptal, anterior, anterolateral, lateral, inferolateral and inferior) by circumferential profile analysis. The percentage reverse redistribution in the infarct area was larger in Tc-tetrofosmin imaging than in Tl imaging (p = 0.013). Reverse redistribution was most prominent in the anterior wall (anterior > anteroseptal > inferoseptal, p = 0.020). This suggests that infradiaphragmatic scatter is unlikely as the mechanism of reverse redistribution. The washout rate of Tc-tetrofosmin in the infarct area (reverse redistribution area) was higher than that in the normal area (non-reverse redistribution area), and was also higher than the washout rate of Tl imaging in the infarct area. The %uptake of delayed images in the infarct area was larger in Tc-tetrofosmin than that in Tl imaging, whereas %uptake of early images did not differ. The percentage reverse redistribution did not correlate with the degree of collateral circulation and the residual stenosis. In conclusion, reverse redistribution of Tc-tetrofosmin was more prominent in the infarct area, and this was due to the relatively lower uptake of reverse redistribution of Tc-tetrofosmin than delayed Tl images.

Angioplasty, Balloon, Coronary↗
PubMedDate not supplied

[Long-term patency of reversed and in situ femoro-popliteal bypasses].

INTRODUCTION: The small choice of graft materials is one of the greatest problems in femoro-popliteal (F-P) bypass reconstructions. Besides all biosynthetics(2-5) and synthetics(6) graft materials, there is no right alternative for autologous saphenous vein graft in F-P reconstructions. There are two main techniques for F-P reconstructions: "reversed" and "in situ". The aim of this study is the comparison of the long-term patency between "reversed" and "in situ" F-P bypasses. PATIENTS AND METHODS: In the study were included 191 patients with "reversed" and 99 patients with "in situ" F-P bypass grafts operated on between 1988 and 1994. There were 153 (80.10%) male and 38 (19.90%) female patients in the group with "reversed" bypass, and 78 (78.78%) male and 21 (21.22%) female patients in the group with "in situ" bypass. The average age of all patients was 59.04 (27-80) years. Eighty five (44.5%) patients in the group with "reversed" F-P bypass had diabetes mellitus and 43 (43.43%) in the group with "in situ" bypass. One hundred and fifty two (79.68%) patients in the group with "reversed" bypass were cigarette smokers and as 80 (80.8%) in the group with "in situ" bypass. In Table 1 the Fontain classification of occlusive diseases in operated patients is presented. The early proximal reconstructions were performed in 49 patients with "reversed" and 16 patients with "in situ" bypasses (Table 2). The associated proximal reconstructions were performed in 21 patients with "reversed" and in 14 patients with "in situ" bypasses (Table 3). All patients were controlled by physical and Doppler ultrasonographic examination immediately after the operation, after 1, 3, 6 months, and then every year postoperativelly. In cases with suspected graft occlusion or any other complication, control angiographic examinations was also performed. The statistical analysis of the results was done using "Life table" analysis. RESULTS: The patients were followed-up from 3 to 10 years. The results of "life-table" analysis are presented in Tables 4-8 and Graph 1. The "in situ" technique showed statistically significant better long-term patency compared to "reversed" technique, after 2 and 10 years (p < 0.05). The immediate patency in cases with "reversed" bypass was 98.96%, while limb salvage was 97.91%. In the same group long-term patency was 72.8% and limb salvage 73.9%. In the group with "in situ" bypasses the immediate patency as well as limb salvage were 96.97%. In the same group long-term patency was 73.8% and limb salvage 77.2%. In Table 5 potential advantages of the "in situ" F-P bypass technique are shown (16-21). However, there are controversial data on clinical results of both bypasses. Some authors described better long-term results of the "in situ" F-P bypass technique (28-30), while according to other data there are no significant differences between these two bypass groups (31-33). Most authors emphasized the two advantages of "in situ" bypasses in F-P reconstructions: a small diameter of the saphenous vein; in cases with pure run off (34-36).

Adult↗
PubMed1987

Human immunodeficiency virus reverse transcriptase. General properties and its interactions with nucleoside triphosphate analogs.

Using affinity purified human immunodeficiency virus (HIV) reverse transcriptase the reaction assay conditions were determined. The optimum incorporation of dTMP into the (rA)n(dT)10 template with HIV reverse transcriptase required 6 mM MgCl2 and 80 mM KCl. The template specificity of HIV reverse transcriptase is quite different from those of the human gamma-polymerase-associated reverse transcriptase or avian virus reverse transcriptase. The preferential inhibition of HIV reverse transcriptase as compared to human gamma-reverse transcriptase was observed with several nucleoside analog triphosphates. The Ki values for thymidine triphosphate analogs with HIV reverse transcriptase ranged from 5 to 13 nM with decreasing effectiveness for 3'-fluoro greater than 3'-amino greater than 2',3'-dideoxy greater than 3'-azido groups. This study provides information on the structure activity relationships of the triphosphate analogs inhibitory effects on HIV reverse transcriptase versus human gamma-polymerase-associated reverse transcriptase, and the possible mechanisms of action of 3' azido thymidine and the 2',3'-dideoxynucleosides, and also identifies other nucleoside analogs for possible development as inhibitors of HIV.

Deoxyguanine Nucleotides↗
PubMed1989

[Changes of the chewing movements following the correction of anterior reversed occlusion in children].

The purpose of this study was to investigate the functional changes following the orthodontic treatment of anterior reversed occlusion in children. Samples consist of 94 children with anterior reversed occlusion as the experimental group and 34 children with normal incisor occlusion as the control group. Experimental samples were classified into 3 groups according to the erupting stages of incisors. Among them, a group of the permanent incisor dentition was divided into 2 groups, one was a "Shift group" showing functional mandibular movement from the centric relation to intercuspal position and the other was a "non-shift group" showing no such functional movement. In the experimental group, 28 patients showed the correction of anterior reversed occlusion orthodontically. Mandibular movements were recorded during gum chewing on all subjects by using Sirognatho Analysing System II. Mandibular movement was examined as to the cycle time and the chewing movement on the horizontal plane. The results were as follows: 1. The chewing cycle was irregular in samples of reversed occlusion. 2. Samples of reversed occlusion with deciduous incisors and of normal occlusion were equivalent in the cycle time. 3. The chewing cycle was more longer in samples of reversed occlusion with maxillary deciduous and mandibular permanent incisors than those of normal occlusion. 4. The chewing cycle was longer in the Shift group than in the normal group. But it was not longer in the Non-shift group than in the normal group. 5. The chewing movement of the Shift group showed type-1, 2, 3 rather than type 4. But the non-shift group showed type-4 more than type-1, 2, 3. 6. The chewing cycle became more regularized after the correction of anterior reversed occlusion. 7. The chewing cycle was shortened in the Shift group after the correction of anterior reversed occlusion. 8. The chewing movement of type-1 and type-4 were decreased after the correction of anterior reversed occlusion. In conclusion, the correction of anterior reversed occlusion by orthodontic method provides improvements of not only the morphological incisor occlusion but also the functional mandibular movements.

Child↗
PubMed1995

Characteristics of a reversed circadian blood pressure rhythm in pregnant women with hypertension.

The objective of the study was to describe the 24h blood pressure (BP) pattern in pregnant women with a reversed circadian BP rhythm. The 24h BP monitorings (24BPM) were performed in 73 pregnant women hospitalised for hypertension. Daytime (6.00 am to 10.00 pm) and night-time (10.00 pm to 6.00 am) BP variables were calculated. A reversed circadian BP rhythm was defined as a day/night mean BP ratio < 1.0. Fourteen women had a reversed circadian BP rhythm. Their diurnal mean BP was equal to women with a normal rhythm, but their nocturnal mean BP was significantly higher and their BP oscillation amplitudes tended to be lower. The day/night mean SBP and DBP ratios correlated significantly, with the DBP ratios slightly higher than corresponding SBP ratios. At repeated 24BPM (n = 14), the day/night ratio did not change but a reversed rhythm occurred during anti-hypertensive medication in one case, and a reversed rhythm switched spontaneously to normal in one case. The diurnal BP pattern did not differ between women with a reversed and a normal circadian BP rhythm. Women with a reversed rhythm had a higher nocturnal BP, but this was part of the definition. The BP variations were generally smaller in the group with a reversed rhythm, although the variations were still large. The definition with a day/night ratio cut-off at 1.0 was rather a convenient mathematical choice than a biologically-determined limit. The day/night BP ratio remained unchanged at repeated measurements, but a reversed rhythm was not a constant phenomenon in occasional cases. Women with a reversed circadian BP rhythm were therefore not a well-defined group and, with exception for a tendency to smaller BP fluctuations, no specific behavioural BP pattern could be described. Until the normal circadian BP rhythm in normal pregnancy has been described, there is no possibility to identify women with an abnormal circadian BP rhythm. Ideally, any definition of an abnormal rhythm should be related to the clinical relevance.

Adolescent↗
PubMed1995

Secondary structure in the 3' UTR of EGF and the choice of reverse transcriptases affect the detection of message diversity by RT-PCR.

The secondary structure in mRNA is essential for many processes, but it can present a technical problem in making full-length cDNA with reverse transcriptases. Furthermore, different reverse transcriptases have differing abilities to transcribe through regions with secondary structure, which can alter the products obtained by reverse-transcribing RNA and then PCR-amplifying the product (RT-PCR). We have been interested in studying the posttranscriptional regulation of epidermal growth factor by RT-PCR and have tested the ability of several reverse transcriptases to reverse transcribe the 3'-untranslated region (3'UTR), a region that contains substantial secondary structure. When low levels of either total RNA or poly(A)+ mRNA were used, we found avian myeloblastosis virus reverse transcriptase (AMV-RT) to be the most robust of all the enzymes tested. Furthermore, contrary to reports that AMV-RT is inhibited by tRNA--which should make it less effective than Moloney murine leukemia virus reverse transcriptase (MMLV-RT) at reverse-transcribing total RNA--adding tRNA to poly(A)+ RNA actually increased the amount of specific RT-PCR product obtained with AMV-RT while it decreased the amount of product and enhanced mispriming with MMLV-RT. We found that pre-incubation of the oligo(dT) primer with total RNA at elevated temperature prior to reverse transcription improved the efficiency of both native and modified MMLV-RTs. These findings support the concept that secondary structures in RNA differentially affect the abilities of different reverse transcriptases to detect transcript diversity and raise the possibility that such structures could affect quantitation using RT-PCR with internal mRNA standards.

Avian Myeloblastosis Virus↗
PubMed1997

Reverse-micelle model: pH, electromagnetic field and inhibitor enzyme interaction.

The reverse micelle is one of many models thought to have properties more nearly resembling the biological cellular environment, than does the traditional dilute-solution biochemical reaction system. In order to evaluate the results of EMF perturbation of enzyme-catalyzed reactions, the description of the AOT reverse-micelle model, with respect to its internal pH, effect of chemical inhibitors, temperature, and electromagnetic-field perturbation has herein been extended. Acetylcholinesterase and NADPH cytochrome-P450 reductase, reacting within the AOT reverse-micelle, exhibit a temperature vs. activity profile equivalent to the same reaction in a buffered dilute-solution environment. In reverse micelles, some inhibitors of AChE (propidium, and d-tubocurarine) have much less effect upon indophenol-acetate hydrolysis than they do in a dilute solution environment. Other inhibitors act in the same manner within the structured environment of the reverse micelle as in the conventional dilute solution reaction model. These differences are explicable in terms of mechanism of action of the individual inhibitors. Perturbation by low-intensity microwave fields has a similar inhibitory effect upon dilute-solution reactions, as those in the 'low-water-activity' environment of the reverse micelle. However, the interactions between physical and chemical perturbants are differently limited by the structure of the aqueous phase of the reverse micelle. pH of the 'internal' reverse-micelle environment is a function of the availability of H-ions supplied by system components. Use of indicator dyes show that the low-molarity buffers which are compatible with reverse-micelle stability, are often insufficient to maintain a constant pH. Too, in the reverse micelle, reaction rate, for proton yielding reactions, is dramatically greater than the rate of the same reaction in dilute solution at the same acidic pH.

Animals↗
PubMed1997

The endocrine mechanism of sex reversal in the protandrous black porgy, Acanthopagrus schlegeli: a review.

Black porgy, Acanthopagrus schlegeli Bleeker, a marine protandrous hermaphrodite, is a functional male for the first 2 years of life but begin to sexually reverse to female after the third year. This sex pattern provides a very good model to study the mechanism of sex reversal in fish. The gonad at 5 month of age consisted of testicular tissue with few primary oocytes at 5 month of age. The ovarian tissue became dominant at 18 months of age during the non-spawning season. Testicular and ovarian tissues were separated by connective tissue. Plasma estradiol-17 beta(E2), vitellogenin and 11-ketotestosterone (11-KT) profiles in males were significantly different from those in the 3-year-old reversing females. Peak levels of plasma E2 in the reversing females occurred during the early prespawning season (in October). Lower levels of plasma E2 were, however, observed in the males. Plasma 11-KT levels significant decreased but no changes of plasma testosterone were detected in the reversing females. Exogenous E2 suppressed the testicular development but induced the gonadal aromatase activity, ovarian development and sex reversal in 2-year-old black porgy. Exogenous T and LHRH analog did not have effects on the sex reversal. Higher concentrations of pituitary GtH II and mRNA of GtH II-beta subunit were detected in the reversed females. These data suggested that E2 and gonadal aromatase closely associated to the occurrence of sex reversal. A working model of the sex reversal in black porgy is proposed.

Animals↗
PubMed1999

Eukaryotic expression of enzymatically active human immunodeficiency virus type 1 reverse transcriptase.

Reverse transcriptase of human immunodeficiency virus type I is a vitalenzyme in the HIV-1 replication cycle and an attractive target of attempts to arrest a primary viral infection. We designed a vector for eukaryotic expression of the 66 kDa subunit of reverse transcriptase under the control of the immediate early cytomegalovirus promoter. Efficient transient expression of the 66 kDa subunit of reverse transcriptase was achieved in a variety of cells. Immunostaining of the transfected cells revealed the cytoplasmatic localization of reverse transcriptase. Reverse transcriptase activity was detected in all transfected cell lines. Injection of this plasmid encoding the 66 kDa subunit of reverse transcriptase into mice resulted in strong reverse transcriptase-specific immune responses indicating that the 66 kDa subunit of reverse transcriptase is expressed in vivo. Sera from DNA-immunized mice inhibited reverse transcription in vitro.

3T3 Cells↗