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R D Johnson

Publications and source records attributed to R D Johnson.

At least 55 records · Page 3Linked to original sources
PubMed1999

Mammalian XRCC2 promotes the repair of DNA double-strand breaks by homologous recombination.

The repair of DNA double-strand breaks is essential for cells to maintain their genomic integrity. Two major mechanisms are responsible for repairing these breaks in mammalian cells, non-homologous end-joining (NHEJ) and homologous recombination (HR): the importance of the former in mammalian cells is well established, whereas the role of the latter is just emerging. Homologous recombination is presumably promoted by an evolutionarily conserved group of genes termed the Rad52 epistasis group. An essential component of the HR pathway is the strand-exchange protein, known as RecA in bacteria or Rad51 in yeast. Several mammalian genes have been implicated in repair by homologous recombination on the basis of their sequence homology to yeast Rad51: one of these is human XRCC2. Here we show that XRCC2 is essential for the efficient repair of DNA double-strand breaks by homologous recombination between sister chromatids. We find that hamster cells deficient in XRCC2 show more than a 100-fold decrease in HR induced by double-strand breaks compared with the parental cell line. This defect is corrected to almost wild-type levels by transient transfection with a plasmid expressing XRCC2. The repair defect in XRCC2 mutant cells appears to be restricted to recombinational repair because NHEJ is normal. We conclude that XRCC2 is involved in the repair of DNA double-strand breaks by homologous recombination.

Animals↗
PubMed1999

Neurotrophin modulation of the monosynaptic reflex after peripheral nerve transection.

The effects of neurotrophin-3 (NT-3) and NT-4/5 on the function of axotomized group Ia afferents and motoneurons comprising the monosynaptic reflex pathway were investigated. The axotomized medial gastrocnemius (MG) nerve was provided with NT-3 or NT-4/5 for 8-35 d via an osmotic minipump attached to its central end at the time of axotomy. After this treatment, monosynaptic EPSPs were recorded intracellularly from MG or lateral gastrocnemius soleus (LGS) motoneurons in response to stimulation of the heteronymous nerve under pentobarbital anesthesia. Controls were preparations with axotomized nerves treated directly with vehicle; other axotomized controls were administered subcutaneous NT-3. Direct NT-3 administration (60 microgram/d) not only prevented the decline in EPSP amplitude from axotomized afferents (stimulate MG, record LGS) observed in axotomy controls but, after 5 weeks, led to EPSPs larger than those from intact afferents. These central changes were paralleled by recovery of group I afferent conduction velocity. Removal of NT-3 4-5 weeks after beginning treatment resulted in a decline of conduction velocity and EPSP amplitude within 1 week to values characteristic of axotomy. The increased synaptic efficacy after NT-3 treatment was associated with enhanced connectivity of single afferents to motoneurons. NT-4/5 induced modest recovery in group I afferent conduction velocity but not of the EPSPs they elicited. NT-3 or NT-4/5 had no effect on the properties of treated motoneurons or their monosynaptic EPSPs. We conclude that NT-3, and to a limited extent NT-4/5, promotes recovery of axotomized group Ia afferents but not axotomized motoneurons or the synapses on them.

Afferent Pathways↗
PubMed1999

Hypoxia limits differentiation and up-regulates expression and activity of prostaglandin H synthase 2 in cultured trophoblast from term human placenta.

OBJECTIVE: We determined the effect of hypoxic conditions on cellular differentiation and prostaglandin H synthase expression in cultured human term trophoblast. STUDY DESIGN: Cytotrophoblasts isolated from term placentas were cultured for 24-72 hours in a standard (20% oxygen) or hypoxic (1% to 2% oxygen) atmosphere. Trophoblast biochemical differentiation was determined by release of human chorionic gonadotropin. Morphologic differentiation was evaluated by epifluorescent and confocal microscopic examination of cultures after dual cytochemical staining for surface membrane desmosomes and intracytoplasmic nuclei. Prostaglandin H synthase 2 expression was determined by Western blot analysis and correlated with enzyme activity by immunoassay of 2 prostaglandin H synthase 2 products, prostaglandin E2 and thromboxane. RESULTS: Human chorionic gonadotropin levels in media and syncytiotrophoblast formation were markedly diminished in trophoblast cultured in hypoxia, compared with trophoblast in control cultures, whereas viability was unchanged. Hypoxia up-regulated expression of trophoblast prostaglandin H synthase 2 but not of prostaglandin H synthase 1 and increased prostaglandin E2 and thromboxane release. CONCLUSIONS: Hypoxia limits differentiation and enhances prostaglandin H synthase 2 expression in cultured villous trophoblast. These responses may account for the cytotrophoblast prominence characteristic of placentas exposed to attenuated oxygen delivery.

Blotting, Western↗
PubMed1999

Pharmacokinetics of sparfloxacin in patients with renal impairment.

Sparfloxacin is a fluoroquinolone antimicrobial agent with a broad spectrum of activity and long elimination half-life. Because its single-dose pharmacokinetics are altered by renal impairment, the present study was undertaken to determine the effects of moderate or severe renal insufficiency on the multidose pharmacokinetic characteristics of and tolerance to sparfloxacin. The pharmacokinetic characteristics of sparfloxacin were assessed in 32 subjects (15 men, 17 women) with (1) normal renal function (creatinine clearance [CLcr]> or = 250 mL/min per 1.73 m2) and a mean age of 52.6 years and mean weight of 70.4 kg; (2) moderate renal insufficiency (CLcr 30-49 mL/min per 1.73 m2) and a mean age of 54.4 years and mean weight of 67.8 kg; and (3) severe renal insufficiency (CLcr 10-29 mL/min per 1.73 m2) and a mean age of 50.8 years and mean weight of 73.1 kg. The first 2 groups received a 400-mg loading dose on day 1 followed by 200 mg once daily for 9 days; subjects with severe renal insufficiency received a 400-mg loading dose on day 1 followed by 200 mg every 48 hours on days 3, 5, 7, and 9. The plasma and urinary pharmacokinetics of sparfloxacin and its glucuronide metabolite were determined after the last dose. All subjects were monitored for changes in the corrected QT (QTc) interval and for adverse events. Renal insufficiency altered the steady-state pharmacokinetic variables of sparfloxacin and its glucuronide metabolite, reducing their renal clearances and increasing both maximum plasma concentration and area under the plasma concentration-time curve. Mean steady-state plasma sparfloxacin concentrations in subjects with severe renal insufficiency (48-hour dosing interval) were comparable to those in subjects with normal renal function (24-hour dosing interval). However, mean plasma sparfloxacin concentrations in patients with moderate renal insufficiency were 2 to 3 times greater than the corresponding concentrations in subjects with normal renal function receiving the same dosage regimen. The QTc interval was slightly increased in all groups (the greatest increases were 14, 14, and 6 milliseconds in the groups with normal renal function and moderately and severely impaired renal function, respectively, at 5.5 hours post-dose on day 9 or 10) but similar among subjects with normal renal function or with renal insufficiency. Sparfloxacin was well tolerated. Thus sparfloxacin clearance is reduced and plasma concentrations raised by moderate or severe renal insufficiency. These increases do not appear to augment drug effects on the QTc interval or enhance the risk for adverse events. These results suggest that alternate-day dosing (48-hour dosing interval) following a double loading dose on day 1 should be used in patients with severe renal insufficiency and may be appropriate for patients with moderate renal insufficiency.

Adult↗
PubMed1999

Pharmacokinetic interaction of sparfloxacin and digoxin.

Sparfloxacin, a broad-spectrum, oral fluoroquinolone antimicrobial agent, has a long elimination half-life that permits once-daily administration. Antibiotics may increase the oral bioavailability of digoxin, leading to increases in its plasma concentration. Since patients treated with sparfloxacin may be receiving concurrent treatment with digoxin, the possibility of an interaction between sparfloxacin and digoxin was examined in a double-masked, placebo-controlled, multiple-dose, two-way crossover study in 24 healthy male volunteers between 20 and 49 years of age. All subjects were given digoxin 0.3 mg once daily throughout the 20-day study. Sparfloxacin (or placebo) was given as a 400-mg loading dose on day 1, followed by single 200-mg daily doses for 9 days, with crossover to the alternate treatment on days 11 through 20. Plasma levels of digoxin were analyzed by validated radioimmunoassay, and plasma levels of sparfloxacin were analyzed by validated high-performance liquid chromatography. Concomitant administration of sparfloxacin and digoxin was generally well tolerated. Mean values for steady-state area under the concentration-time curve over 24 hours for the 2 treatments were virtually identical: 28.4 ng/h per mL(-1) for digoxin administered with placebo and 28.9 ng/h per mL(-1) for digoxin administered concomitantly with sparfloxacin. Mean steady-state maximum plasma concentrations were 3.91 and 3.59 ng/mL for digoxin with placebo and digoxin with sparfloxacin, respectively. Mean steady-state trough plasma digoxin concentrations for the 2 treatments were 0.87 and 0.89 ng/mL, respectively. Mean times to steady-state maximum plasma concentrations were identical at 0.89 hours for both treatments. Mean steady-state oral clearance was 10.6 L/h for digoxin alone and 10.4 L/h for digoxin with sparfloxacin. Thus administration of sparfloxacin in combination with digoxin did not alter the pharmacokinetics of digoxin in healthy male volunteers aged 20 to 49 years. Steady-state plasma sparfloxacin concentrations were consistent with those obtained in other multiple-dose phase I studies, suggesting that digoxin does not alter the steady-state pharmacokinetics of sparfloxacin.

Adult↗
PubMed1999

Effect of single ascending, supratherapeutic doses of sparfloxacin on cardiac repolarization (QTc interval).

This double-masked, randomized, placebo-controlled study was conducted in healthy adult male and female volunteers with no clinically relevant baseline electrocardiographic (ECG) abnormalities to assess the cardiac tolerability margin of sparfloxacin (as measured by the effect on QTc interval) under conditions of potential overdose at up to 4 times the usual therapeutic loading dose. The 23 enrolled volunteers received a sequence of single doses of sparfloxacin (400, 800, 1200, and 1600 mg), 1 dose in each of 4 study periods. Six volunteers received placebo during each period. A 14-day washout separated the periods. Serial blood samples and ECG measurements were collected in each period to determine the pharmacokinetic and pharmacodynamic characteristics of sparfloxacin. The area under the concentration-time curve from time zero to infinity (AUC0-infinity) exhibited dose proportionality. The maximum plasma concentration (Cmax) after the 1200- and 1600-mg doses was lower than would be expected for a linear dose relationship. This was also the case with the mean increase and mean maximum increase in QTc interval. Increases in the QTc interval correlated well with Cmax but not with AUC0-infinity. The time to reach Cmax showed a slight tendency to increase with dose, as did the terminal elimination half-life. Changes in QTc-interval dispersion were similar for both placebo recipients and sparfloxacin-treated volunteers and were of no clinical consequence. At supratherapeutic doses, the extent of sparfloxacin's absorption (AUC0-infinity) was dose independent; however, the rate of absorption was dose dependent, with Cmax increasing substantially less than proportionally to the administered dose. This limited the Cmax of sparfloxacin at supratherapeutic doses and thus the increase in QTc interval. Rechallenge demonstrated that only 2 of 8 subjects had the same degree of QTc-interval prolongation, emphasizing the marked variability in the QTc interval.

Adult↗
PubMed1999

Effects of food on the pharmacokinetics of sparfloxacin.

Sparfloxacin, a fluoroquinolone with a broad antimicrobial spectrum and long elimination half-life, is indicated for the treatment of community-acquired pneumonia and acute bacterial exacerbations of chronic bronchitis in adult patients. The present study was undertaken to determine the effects of skim milk and a high-fat breakfast without milk on the single-dose pharmacokinetic characteristics of this antibiotic. The pharmacokinetics of a single 200-mg dose of sparfloxacin were assessed in a 3-way crossover study that included 23 healthy male volunteers who had fasted, had ingested 240 mL of skim milk, or had consumed a standard high-fat breakfast. The subjects' mean age and weight were 26.5 years and 73.2 kg, respectively; 17 were white, 5 Hispanic, and 1 black. Neither skim milk nor the high-fat breakfast had a statistically significant effect on sparfloxacin absorption, as reflected in the maximum plasma concentration (Cmax) and area under the plasma concentration-time curve (AUC). Ninety percent confidence limits for logarithmically transformed AUC from time zero to infinity and Cmax were within the 80% to 125% range, demonstrating that the rate and extent of sparfloxacin absorption with skim milk or a high-fat breakfast were not different from those under fasted conditions. As indicated by an increase in the time to Cmax from 3.6 to 5.4 hours, the high-fat breakfast slightly delayed the onset of sparfloxacin absorption. Skim milk and the high-fat breakfast did not significantly affect the elimination kinetics of sparfloxacin. Sparfloxacin was well tolerated in all 3 treatment groups. Despite the apparent delay in the onset of absorption, the bioavailability of sparfloxacin in the healthy male subjects in this study population was not affected by concomitant administration with skim milk or a high-fat meal. Accordingly, the results suggest that sparfloxacin can be administered without regard to the ingestion of milk or meals.

Adolescent↗
PubMed1999

Changes in neuronal receptive field characteristics in caudal brain stem following chronic spinal cord injury.

Chronic spinal cord injury pain is poorly understood and, thus, not effectively relieved by traditional treatments. In the present study, a variety of partial, severe and sham chronic spinal lesions were made in 31 male rats at spinal level T8. During routine care/handling and brief behavioral testing of the animals throughout the 30-day recovery period, the majority of those with severe contusion injuries (verified histologically) showed signs of mechanical hypersensitivity on the dorsolateral trunk just rostral to the level of injury (i.e., upper thoracic territory). Terminal electrophysiological experiments were performed on all rats (urethane anesthesia). Single unit recordings were made at two supraspinal locations within the caudal brainstem, the nucleus reticularis gigantocellularis and nucleus reticularis gigantocellularis pars alpha. Neurons in these areas normally receive bilateral nociceptive somatovisceral inputs from many parts of the body. Seventy-three percent of the animals with severe contusion injuries developed novel low-threshold neuronal responses to stimulation of the dorsolateral trunk (upper thoracic territory). This amount was significantly greater than for animals with more moderate spinal lesions (dorsal or lateral hemisection; 29% and 25%, respectively) or sham controls (0%). These data suggest (1) that the spinal contusion is a reliable model for studies of the neural mechanisms that underly central spinal cord injury-related pain and (2) that the caudal brainstem is one supraspinal location where neurons undergo significant changes in responsiveness following severe chronic spinal cord injury. The observed plasticity is likely part of the central reorganization producing the multitude of sensory disturbances that surface following spinal cord injury.

Afferent Pathways↗
PubMed1999

Effects of acute and chronic midthoracic spinal cord injury on neural circuits for male sexual function. I. Ascending pathways.

Normal male reproductive function, particularly ejaculation, requires the integrity of urogenital sensory input and its ascending spinal projections. After midthoracic chronic spinal cord injury, sexual dysfunction occurs in both rats and humans. Neurons in the medullary reticular formation (MRF) are involved in the processing of bilaterally convergent sensory inputs from multiple cutaneous, mucocutaneous, and visceral regions of the body, including the penis and male urogenital tract. A variety of acute and chronic lesions were used to determine the midthoracic location of ascending spinal pathways conveying sensory input from the penis and male urogenital tract to MRF. A total of 371 single neurons were recorded in the MRF of 34 urethan-anesthetized mature male rats. Twenty-seven rats received a chronic T8 dorsal (DHx) or lateral (LHx) hemisection or contusion (Cx) injury 30 days before the terminal electrophysiological experiments. In addition, nine dorsal nerve of the penis (DNP)-responsive MRF neurons in seven intact control animals were tested completely both before and after various select acute spinal cord lesions. The chronic lesion data indicate that low and high threshold input from the penis (mucocutaneous) and male urogenital tract (visceral) ascend bilaterally within the dorsal quadrant at T8 as opposed to high threshold input from the hindpaws (cutaneous), which ascends unilaterally in the ventrolateral quadrant (VLQ). The acute lesion data indicate that the low-threshold information conveyed from the penis and male urogenital tract ascends in the dorsal columns, as opposed to the high-threshold nociceptive inputs that ascend bilaterally in the dorsolateral quadrant (DLQ). These results, as well as previous data on ascending projections from female reproductive organs within the dorsal columns and DLQ to other caudal brain stem nuclei, provide evidence for ascending pathways conveying nociceptive information centrally via the DLQ. This spinal gray-DLQ pathway(s) conveying information from mucocutaneous/pelvic/visceral territories therefore differs from the traditionally recognized spinal gray-VLQ pathway(s), which is known to convey nociceptive information from cutaneous regions of the body.

Acute Disease↗
PubMed1999

Clinical and pharmacokinetic phase I study of multitargeted antifolate (LY231514) in combination with cisplatin.

PURPOSE: Multitargeted antifolate (MTA; LY231514) has broad preclinical antitumor activity and inhibits a variety of intracellular enzymes involved in the folate pathways. This study was designed to (1) determine the maximum-tolerated dose (MTD), dose-limiting toxicities (DLT), and pharmacokinetics of MTA combined with cisplatin; (2) determine a recommended dose for phase II studies; and (3) collect anecdotal information on the antitumor activity of MTA combined with cisplatin. PATIENTS AND METHODS: Patients with solid tumors received MTA intravenously over 10 minutes and cisplatin over 2 hours once every 21 days. In cohort 1, both agents were administered on day 1 starting with MTA 300 mg/m(2) and cisplatin 60 mg/m(2). In cohort 2, MTA (500 or 600 mg/m(2)) was administered on day 1, followed by cisplatin (75 mg/m(2)) on day 2. RESULTS: In cohort 1, 40 assessable patients received 159 courses of treatment. The MTD was MTA 600 mg/m(2)/cisplatin 100 mg/m(2). DLTs were reversible leukopenia/neutropenia and delayed fatigue. Hydration before cisplatin therapy did not influence MTA pharmacokinetics. Eleven objective remissions included one complete response in a patient with relapsed squamous cell head and neck carcinoma, and partial responses in four of ten patients with epithelial pleural mesothelioma. In cohort 2, 11 assessable patients received 23 courses of treatment. The MTD was MTA 600 mg/m(2) and cisplatin 75 mg/m(2). DLTs were neutropenic sepsis, diarrhea, and skin toxicity. Two patients died of treatment-related complications during the study. Two patients had objective remissions (one mesothelioma patient, one colon cancer patient). CONCLUSION: The combination of MTA and cisplatin is clinically active, and administering both agents on day 1 is superior to a split schedule. Further development of this combination for mesothelioma is warranted.

Adult↗
PubMed1999

Preliminary results of a phase I study with MTA (LY231514) in combination with cisplatin in patients with solid tumors.

MTA (multitargeted antifolate, LY231514) is a novel antimetabolite resulting from structure/activity studies of the lometrexol-type antifolates. It has been shown to inhibit various enzymes of folate pathways and has broad antitumor activity in a variety of in vitro and in vivo tumor models. Clinical phase 1 studies have been performed using different administration schedules, and subsequently the every-21-days schedule has been selected for further development. We report the preliminary findings from a combination phase I study of MTA and cisplatin administered every 21 days. In the first cohort (34 patients), both agents were administered on day 1 with a starting dose of 300 mg/m2 MTA and 60 mg/m2 cisplatin. In a second cohort (10 patients), MTA (500 or 600 mg/m2) was administered on day 1 followed by cisplatin (75 mg/m2) on day 2. The maximum tolerated doses were reached at 600 mg/m2 MTA/100 mg/m2 cisplatin (cohort 1) and 600 mg/m2 MTA/75 mg/m2 cisplatin (cohort 2). In cohort 1, dose-limiting toxicities consisted of reversible myelosuppression with leukopenia and neutropenia. In addition, delayed fatigue also was of clinical significance. Pharmacokinetic analyses indicated that hydration administered before the administration of cisplatin did not influence the major pharmacokinetic parameters of MTA. Eleven objective remissions were observed, including one complete response in a patient with relapsed squamous cell carcinoma of the head and neck and partial responses in four of seven patients with mesothelioma In contrast, the dose-limiting toxicities in patient cohort 2 consisted of neutropenic sepsis, diarrhea, and skin toxicity with two possibly treatment-related deaths on study. No objective remissions are presently observed in cohort 2. We conclude that the combination of MTA and cisplatin is feasible and clinically active when both agents are administered on day 1 and that it should be pursued for further clinical development.

Adult↗
PubMed1998

Brainstem microstimulation differentially inhibits pudendal motoneuron reflex inputs.

Sensory information from the genitourinary tract is transmitted to perineal muscle reflex circuits in the spinal cord and to the medullary reticular formation (MRF). This is the first report of a functional descending connection between neurons in the MRF and sensory inputs of the pudendal motoneuron reflex circuitry. In anesthetized rats, microstimulation of the MRF produced a decrease in amplitude and increase in latency of pudendal motoneuron reflex discharges (PMRD) elicited by stimulation of the dorsal nerve of the penis. No effects on pelvic nerve-elicited PMRD were found. The most robust reflex depression was observed when the lateral paragigantocellular reticular nucleus was microstimulated bilaterally. Reproductive abnormalities that occur following severe spinal cord injury probably result from disruption of these descending connections.

Animals↗
PubMed1998

Anodally focused polarization of peripheral nerve allows discrimination of myelinated and unmyelinated fiber input to brainstem nuclei.

We investigated the ability of a novel direct current (DC) polarization technique to block selectively the conduction in peripheral myelinated nerve fibers and allowing propagation in only unmyelinated fibers. In anesthetized adult rats, distal branches of the sciatic nerve (caudal cutaneous sural and tibial nerves) were exposed for electrical stimulation of A- and C-fibers. Two specially fabricated trough electrodes of different size and surface area were placed onto the sciatic nerve. Through these proximal electrodes a controlled ramped DC was timed to coincide with the arrival of A- and C-fiber action potentials, evoked electrically at the distal nerves or naturally from the foot or ankle, with the intent of blocking propagation in A-fibers while allowing C-fiber throughput. Neuronal recordings were made both peripherally (proximal sciatic nerve fascicles or L5 dorsal roots) and centrally (single cells in the nucleus gracilis or nucleus reticularis gigantocellularis). The DC polarization was shown to block conduction in myelinated A-fibers effectively, while allowing conduction in the unmyelinated C-fibers, without activation of fibers via the DC polarization itself. This was dependent upon the following factors: electrode polarity, onset rate of polarization, peak amplitude of polarization, distance between polarizing electrodes, size difference between polarizing electrodes, and gross nerve size. These experiments demonstrate that anodally focused DC polarization, applied utilizing two trough electrodes of different sizes, is capable of effectively, reversibly, and reproducibly blocking conduction in myelinated A-fibers evoked either electrically or naturally, while still allowing conduction to occur in the unmyelinated C-fiber population. In the context of experimental usage, we have demonstrated blocking of low-threshold A-fiber, but not C-fiber, mediated inputs to the caudal brainstem. This technique should find wide application in studies involving the processing of information conveyed centrally by the unmyelinated C-fiber afferent population, including discriminating afferent responses to peripheral stimuli, the role of C-fiber input in reflex activity, and the plasticity following injury or other manipulations.

Action Potentials↗
PubMed1998

Lipopolysaccharide enhances the transcription of prostaglandin H synthase-2 gene in primary human trophoblasts.

OBJECTIVE: Our purpose was to test the hypothesis that lipopolysaccharide potentiates the transcription of human placental prostaglandin H synthase-2 gene, a rate-limiting enzyme in prostaglandin synthesis. STUDY DESIGN: We transfected normal term cytotrophoblasts with a reporter vector containing either prostaglandin H synthase-1 or prostaglandin H synthase-2 promoters, cloned upstream of a luciferase. We correlated lipopolysaccharide effect on luciferase activity with enzyme expression and prostaglandin E2 production. RESULTS: Lipopolysaccharide (10 microg/ml) enhanced prostaglandin H synthase-2 but not prostaglandin H synthase-1 promoter activity. This effect was abolished by dexamethasone (10(-6) mol/L). Platelet-derived growth factor, a known stimulant of prostaglandin H synthase-2 in nonplacental tissue, had no effect on the activity of prostaglandin H synthase-2 promoter. Lipopolysaccharide induced the expression of prostaglandin H synthase-2 messenger ribonucleic acid fivefold to sixfold and caused a 3.5-fold increase in prostaglandin E2 production. CONCLUSION: Lipopolysaccharide enhances the transcription of prostaglandin H synthase-2 gene in human trophoblasts. Enhanced expression of prostaglandin H synthase-2 results in a higher production of prostaglandins, which may alter villous blood flow.

Anti-Inflammatory Agents↗
PubMed1998

The release of 15-hydroxyeicosatetraenoic acid by human placental trophoblast is increased in preeclampsia.

OBJECTIVE: We tested the hypothesis that trophoblast produces 15-hydroxyeicosatetraenoic acid, and its level is elevated in trophoblast from preeclamptic women compared with normal. We also used selective enzymatic inhibitors to determine the relative contributions of 15-lipoxygenase and the two isozymes of prostaglandin H synthase to 15-hydroxyeicosatetraenoic acid levels. STUDY DESIGN: Cytotrophoblasts isolated from placentas of normal or preeclamptic women were cultured in the presence or absence of enzyme inhibitors. Media levels of 15-hydroxyeicosatetraenoic acid were measured by radioimmunoassay. RESULTS: When compared with normal pregnancies, cytotrophoblasts from preeclamptic pregnancies released up to fivefold higher levels of 15-hydroxyeicosatetraenoic acid. Aspirin, an inhibitor of both the prostaglandin H synthase-1 and prostaglandin H synthase-2 isozymes, and nordihydroguaiaretic acid, a selective inhibitor of lipoxygenases, both significantly inhibited 15-hydroxyeicosatetraenoic acid production. In contrast, the selective prostaglandin H synthase-2 inhibitor NS-398 had no effect on 15-hydroxyeicosatetraenoic acid release in the absence of aspirin, but NS-398 reduced 15-hydroxyeicosatetraenoic acid levels in normal trophoblast pretreated with aspirin. CONCLUSIONS: The data indicate that 15-hydroxyeicosatetraenoic acid is produced in trophoblasts and its release by cytotrophoblasts is higher in preeclamptic pregnancies compared with normal controls. Both lipoxygenase and prostaglandin H synthase contribute to 15-hydroxyeicosatetraenoic acid production, and aspirin reduces 15-hydroxyeicosatetraenoic acid secretion. We suggest 15-hydroxyeicosatetraenoic acid plays a role in the oxidation of lipoproteins and the endothelial damage characteristic of preeclampsia.

Aspirin↗
PubMed1998

Gene expression of nitric oxide synthase in cultured human term placental trophoblast during in vitro differentiation.

The human placental syncytiotrophoblast is derived from differentiating cytotrophoblasts and is in contact with maternal blood. This endothelial function positions the trophoblast to regulate maternal-fetal exchange and to influence circulatory dynamics through paracrine interactions in the placenta. Two isoforms of nitric oxide synthase (NOS) are expressed in placenta, and northern analysis, reverse transcription-polymerase chain reaction (RT-PCR), and immunocytochemistry were used to correlate expression of the type II, inducible NOS (iNOS) and the type III, endothelial NOS (eNOS) with state of differentiation in cultured trophoblast from term placentae. It was also tested whether cytokines known to induce NOS in other cell systems would induce iNOS in human trophoblast. The mRNA for eNOS was detected by RT-PCR, but not by Northern analysis, in cultures grown for 24 h when cytotrophoblasts were dominant. In contrast, eNOS mRNA was abundant in cultures grown for 72 h when syncytiotrophoblast was present. Immunocytochemical staining for eNOS protein showed specific fluorescence in a few cells in cultures at 24 h, but the vast majority of cells expressed eNOS at 72 h. The iNOS isoform was expressed neither basally in any trophoblast culture nor was this isoform induced in cultures exposed to interleukin-1, tumour necrosis factor-alpha, interferon-gamma and lipopolysaccharide. The in vitro pattern of trophoblast eNOS expression models the in vivo pattern of eNOS expression described for villous trophoblast. The results suggest that eNOS plays a role in human trophoblast differentiation and function.

Base Sequence↗
PubMed1998

A comparison of transnasal and transoral oesophagogastroduodenoscopy.

Transnasal oesophagogastroduodenoscopy (OGD) with a narrow-bore endoscope has been demonstrated to be feasible in unsedated volunteers. The aim of the study was to compare efficacy, safety, patient tolerance and costs between this novel approach to OGD and standard sedated OGD. Sixty day patients were enrolled for either sedated transoral endoscopy with a standard calibre endoscope or for unsedated transnasal endoscopy with a narrow-bore (5.3 mm diameter) endoscope. Visualization was unsatisfactory in one examination in each group. The only complications were minor epistaxis in four of the transnasal group and oxygen desaturation below 90% in two of the sedated transoral group. On a 1 (very uncomfortable) to 5 (very comfortable) visual analogue scale, the mean transnasal group score was 3.09, compared with 3.86 in the transoral group (P = 0.013). In the transnasal group, mean procedure room time was 15 min compared with 20 min in the transoral group (P < 0.0003), and mean recovery room time was 7 min compared with 37 min (P < 0.0001). Consumable and pharmaceutical costs were reduced by 65 and 92%, respectively. This study demonstrates that unsedated transnasal OGD is a safe and effective route for OGD and has acceptable patient tolerance. The safety and decreased recovery times offer major cost savings and the potential for this method of OGD to become an office procedure for the investigation of the upper gastrointestinal tract.

Adult↗