Impaired innate interferon induction in severe therapy resistant atopic asthmatic children.

Deficient type I interferon-ß and type III interferon-λ induction by rhinoviruses has previously been reported in mild/moderate atopic asthmatic adults. No studies have yet investigated if this occurs in severe therapy resistant asthma (STRA). Here, we show that compared with non-allergic healthy control children, bronchial epithelial cells cultured ex vivo from severe therapy resistant atopic asthmatic children have profoundly impaired interferon-ß and interferon-λ mRNA and protein in response to rhinovirus (RV) and polyIC stimulation. Severe treatment resistant asthmatics also exhibited increased virus load, which negatively correlated with interferon mRNA levels. Furthermore, uninfected cells from severe therapy resistant asthmatic children showed lower levels of Toll-like receptor-3 mRNA and reduced retinoic acid inducible gene and melanoma differentiation-associated gene 5 mRNA after RV stimulation. These data expand on the original work, suggesting that the innate anti-viral response to RVs is impaired in asthmatic tissues and demonstrate that this is a feature of STRA.

A point prevalence survey of antibiotic use in four acute-care teaching hospitals utilizing the European Surveillance of Antimicrobial Consumption (ESAC) audit tool.

The objective of this research was to assess current patterns of hospital antibiotic prescribing in Northern Ireland and to determine targets for improving the quality of antibiotic prescribing. A point prevalence survey was conducted in four acute teaching hospitals. The most commonly used antibiotics were combinations of penicillins including ß-lactamase inhibitors (33·6%), metronidazole (9·1%), and macrolides (8·1%). The indication for treatment was recorded in 84·3% of the prescribing episodes. A small fraction (3·9%) of the surgical prophylactic antibiotic prescriptions was for >24 h. The results showed that overall 52·4% of the prescribed antibiotics were in compliance with the hospital antibiotic guidelines. The findings identified the following indicators as targets for quality improvement: indication recorded in patient notes, the duration of surgical prophylaxis and compliance with hospital antibiotic guidelines. The results strongly suggest that antibiotic use could be improved by taking steps to address the identified targets for quality improvement.

A prospective randomised controlled trial and economic modelling of antimicrobial silver dressings versus non-adherent control dressings for venous leg ulcers: the VULCAN trial.

OBJECTIVE: To examine the effectiveness and cost-effectiveness of antimicrobial silver-donating dressings for venous leg ulcers compared with simple non-adherent (also known as low-adherent) dressings. DESIGN: A pragmatic, prospective randomised controlled trial (RCT) and cost-effectiveness analysis of silver-donating versus low-adherent dressings in the treatment of venous leg ulcers. A non-randomised observational group was also recruited. SETTING: Primary and secondary care services in the north and south of England (Sheffield and Exeter). PARTICIPANTS: Consenting patients with active ulceration of the lower leg that had been present for a period of greater than 6 weeks. INTERVENTIONS: Patients were randomised to receive either a silver-donating or non-silver low-adherent dressing applied beneath compression bandages or hosiery. The choice of dressing within these groups was left to clinician preference. Evaluation was by clinical assessment, supplemented by evaluation of quality of life and cost-effectiveness. MAIN OUTCOME MEASURES: The primary outcome measure was complete ulcer healing at 12 weeks in the index limb. Secondary measures were costs and quality-adjusted life-years (QALYs), cost-effectiveness, time to healing, and recurrence rate at 6 months and 1 year. RESULTS: In total, 304 participants were recruited to the clinical trial: 213 to the RCT and 91 to the observational arm. Within the RCT 107 were randomised to antimicrobial dressings and 106 to the control dressings. There were no significant differences (p > 0.05) between the two groups for the primary outcome measure of proportion of ulcers healed at 12 weeks (59.6% for silver and 56.7% for control dressings). The overall median time to healing was also not significantly different between the two groups (p = 0.408). A total of 24 patients had recurrent ulcers within 1 year; the recurrence rates of 11.6% (n = 11) for the antimicrobial and 14.4% (n = 13) for the control dressings were not significant. Mean utility valuations for both the EuroQol 5 dimensions (EQ-5D) quality of life questionnaire and Short Form 6 dimensions (SF-6D) utility index showed no differences for either group at 1, 3, 6 or 12 months. Compared with the control group, the antimicrobial group had an incremental cost of 97.85 pounds and an incremental QALY gain of 0.0002, giving an incremental cost-effectiveness ratio for the antimicrobial dressings of 489,250 pounds. Cost-effectiveness modelling of the results of the RCT showed that antimicrobial dressings were not cost-effective. CONCLUSIONS: No significant differences in either primary or secondary end points were found between the use of antimicrobial silver-donating dressings and the control group of low-adherent dressings. Modelling showed that antimicrobial silver dressings were not cost-effective. TRIAL REGISTRATION: Current Controlled Trials ISRCTN72485131.

The symptoms of varicose veins: difficult to determine and difficult to study.

OBJECTIVES: To investigate the activities which may exacerbate symptoms in patients with varicose veins. METHODS: Questionnaires sent to patients before clinics and at least six months later. RESULTS: Both questionnaires were returned by 149 of 203 patients (74%) but only 124 contained adequate data for comparison--55 from patients who had surgical treatment and 69 who had no surgery. At initial presentation, worsening of discomfort attributed to varicose veins was common during (58%) or after (48%) standing and in hot weather (44%), but less when sitting with the feet down (31%), and after (31%) or when walking (19%). Surgery significantly reduced the total number of symptoms reported by patients at follow-up (p<0.02). However, none of the symptoms reported during specific activities was significantly lessened by surgery compared with no treatment--possibly because the attrition of patients during the study resulted in small numbers for analysis. CONCLUSIONS: Symptoms are a common indication for treating varicose veins and it is therefore important to be sure that they are due to the veins, rather than other causes. This report highlights traditional and logical questions which may help to identify symptoms caused by varicose veins but illustrates the difficulty of validating them.

Randomised clinical trial, observational study and assessment of cost-effectiveness of the treatment of varicose veins (REACTIV trial).

OBJECTIVES: To establish the cost-effectiveness of surgery and sclerotherapy for the treatment of varicose veins. DESIGN: Randomised controlled trials (RCTs) were carried out for conservative treatment, sclerotherapy and surgery for varicose veins. An economic analysis was carried out alongside the randomised trial. Economic modelling was undertaken based on the primary data collection and a literature review (database searches undertaken in April 2000 and updated in March 2001). SETTING: Primary data collection was from a large district general hospital and a teaching hospital both in England over a 2-year period from January 1999. Cost-effectiveness analysis and economic modelling were carried out using an NHS perspective. PARTICIPANTS: A total of 1009 patients were recruited. INTERVENTIONS: Thirty-four patents were randomised in Group 1 (minor varicose veins with no reflux, randomised between conservative treatment and sclerotherapy), 77 in Group 2 (moderate varicose veins with reflux, randomised between surgery and sclerotherapy) and 246 in Group 3 (severe varicose veins with reflux, randomised between conservative treatment and surgery). The remaining 652 patients formed the observational part of the study. MAIN OUTCOME MEASURES: The cost-effectiveness analysis was based on NHS treatment costs for the 2002--3 financial year, and utilities based on the Short Form 6D (SF-6D) preference-based health measure. For the clinical trial, the outcome measures were health-related quality of life (HRQoL) [Short Form with 36 Items (SF-36), EuroQol quality of life questionnaire (EQ-5D), visual analogue scale (VAS) and standard gamble], symptomatic relief, anatomical extent (for which a new classification was developed and validated), patient satisfaction and the incidence of complications. RESULTS: Of the RCTs, only the Group 3 trial was large enough to provide clear results. This showed that surgical treatment produced better results than conservative treatment in terms of HRQoL, symptomatic relief, anatomical extent and patient satisfaction. Clinical outcomes of surgery and sclerotherapy showed significant improvement in the extent of varicose veins, symptomatic and HRQoL parameters. Cost-effectiveness analysis based on the Group 3 trial showed that the surgery produced an estimated discounted benefit of 0.054 quality-adjusted life-year (QALY) over a 2-year period, with an additional discounted cost of pound 387.45, giving an incremental cost-effectiveness ratio (ICER) of pound 7175 per QALY. Economic modelling suggested that surgery produced a still greater benefit when considered with a 10-year time horizon, with an ICER of pound 1936 per QALY. Injection sclerotherapy produced an incremental benefit of approximately 0.044 QALY at a cost of pound 155 when compared with conservative treatment, giving an ICER of pound 3500 per QALY. When surgery was compared with sclerotherapy, surgery produced greater benefit with a lower ICER (showing extended dominance). CONCLUSIONS: Standard surgical treatment of varicose veins by saphenofemoral ligation, stripping and multiple phlebectomies is a clinically effective and cost-effective treatment for varicose veins, with an ICER well below the threshold normally considered appropriate for the funding of treatments within the NHS. Injection sclerotherapy also appears to be cost-effective, but produces less overall benefit, with a higher ICER than surgery for patients with superficial venous reflux. In minor varicose veins without reflux, sclerotherapy is likely to provide a small average benefit with acceptable cost-effectiveness. Research is needed into methods for accurate and acceptable utility evaluations for conditions with relatively minor effect on HRQoL and also for a validated and standardised method of classification for varicose veins.

Cost-effectiveness analysis of surgery versus conservative treatment for uncomplicated varicose veins in a randomized clinical trial.

BACKGROUND: Despite being a common procedure, the cost effectiveness of surgery for varicose veins has not been established. METHODS: Cost-effectiveness analysis was carried out alongside a randomized clinical trial at two vascular units within National Health Service (NHS) hospitals. Some 246 patients with uncomplicated varicose veins and evidence of saphenofemoral or saphenopopliteal reflux were allocated randomly to receive either conservative management or surgical treatment. Incremental cost per quality-adjusted life year (QALY) gained at 24 months following randomization was calculated. RESULTS: Total NHS costs during the 2-year study period were higher for the surgically treated group (733 UK pounds) than for those who had conservative treatment (345 UK pounds). The difference in costs was statistically significant. The mean incremental health gain from surgical treatment at 24 months was 0.083 QALYs, leading to a base-case estimate of 4682 UK pounds per QALY gained. Assuming an implicit threshold maximum willingness-to-pay value of 20 000 UK pounds for a QALY, the probability of surgical treatment for varicose veins falling below this threshold value was 70 per cent. This result was found to be robust to sensitivity analysis. CONCLUSION: For patients with uncomplicated varicose veins and evidence of saphenofemoral or saphenopopliteal reflux, surgical treatment for varicose veins offers a modest health benefit for relatively little additional NHS cost relative to conservative treatment.

Randomized clinical trial comparing surgery with conservative treatment for uncomplicated varicose veins.

BACKGROUND: Surgical treatment of medically uncomplicated varicose veins is common, but its clinical effectiveness remains uncertain. METHODS: A randomized clinical trial was carried out at two large acute National Health Service hospitals in different parts of the UK (Sheffield and Exeter). Some 246 patients were recruited from 536 consecutive referrals to vascular outpatient clinics with uncomplicated varicose veins suitable for surgical treatment. Conservative management, consisting of lifestyle advice, was compared with surgical treatment (flush ligation of sites of reflux, stripping of the long saphenous vein and multiple phlebectomies, as appropriate). Changes in health status were measured using the Short Form (SF) 6D and EuroQol (EQ) 5D, quality of life instruments based on SF-36 and EuroQol, complications of treatment, symptomatic measures, anatomical extent of varicose veins and patient satisfaction. RESULTS: In the first 2 years after treatment there was a significant quality of life benefit for surgery of 0.083 (95 per cent confidence interval (c.i.) 0.005 to 0.16) quality-adjusted life years (QALYs) based on the SF-6D score and 0.13 (95 per cent c.i. 0.016 to 0.25) based on the EQ-5D score. Significant benefits were also seen in symptomatic and anatomical measures. CONCLUSION: Surgical treatment provides symptomatic relief and significant improvements in quality of life in patients referred to secondary care with uncomplicated varicose veins.

Most patients with varicose veins have fears or concerns about the future, in addition to their presenting symptoms.

INTRODUCTION: This study aimed to document fears and concerns of patients about their varicose veins. REPORT: Completed questionnaires about symptoms and concerns were brought to clinic by 62% (203/329) patients referred with uncomplicated primary varicose veins. 'Concerns, worries or fears' about their veins were reported by 79%. These were seldom mentioned in referral letters, and included fears about thrombosis (31%), trauma (16%), ulcers (15%) and general concerns about the future (57%). DISCUSSION: Many patients have unrecognised fears and concerns about their varicose veins. Specific enquiry and reassurance about these is part of good management and may avoid unnecessary treatment.

The clinical effectiveness of hand held Doppler examination for diagnosis of reflux in patients with varicose veins.

OBJECTIVE: To assess the accuracy of hand held Doppler (HHD) as a rapid screening test for selecting varicose vein patients for duplex imaging. DESIGN: Prospective single blind study of consecutive patients in a randomised trial. MATERIALS: Use of hand held Doppler and duplex ultrasound scanners. METHODS: One thousand two hundred and eighteen legs (943 patients) were examined by HHD and then duplex. HHD examiners recorded whether they would normally have requested duplex. RESULTS: HHD results of one Clinical Assistant (166 limbs) were significantly poorer than all others and his results were excluded from analysis. Duplex would not have been requested in 645 of 1052 (62%) limbs. Among these HHD missed significant reflux in the long saphenous vein in 18 (3%) and the short saphenous in 25 (4%). Reasons for requesting duplex were popliteal fossa reflux (202); recurrent (94) or atypical (86) varicose veins; and possible previous thrombosis (67). Differences were observed between staff and units in requests for duplex; and in thoroughness and style of duplex reporting. CONCLUSIONS: Selective use of HHD can avoid duplex imaging for many patients, with a low failure rate for detecting correctable venous reflux. Observed variations between individuals and units in results of HHD and duplex imaging have implications for the increasing use of duplex by clinicians.

Venous ulcer services in the United Kingdom.

OBJECTIVES: To obtain comprehensive information about venous ulcer services throughout the United Kingdom (UK). DESIGN: Questionnaire based survey. MATERIALS: Questionnaire. METHODS: Letters about venous ulcer services were sent to consultant vascular surgeons in all areas of the UK (total 181). Questionnaires were then directed to the appropriate clinician in each area. RESULTS: Responses were received from 177 (98%) areas. Fifteen (8%) had no dedicated service. Completed questionnaires were returned for 112 (63%) areas. Fifty-six (54%) services were managed by acute hospitals, 29 (28%) by primary care (community) and 19 (18%) jointly. Doctors supervised services in 65 (64%) (vascular surgeons 49, dermatologists 12, both 4) and nurses in 31 (33%). New referrals per week were 1-50 based on audit (33%) or estimates (67%). Sixty-three (58%) services had no database. Written guidelines existed for 76% services (90% nurse-supervised and 64% doctor-supervised services--p<0.02). CONCLUSIONS: Dedicated venous ulcer services have been shown to improve healing rates and quality of life. Although now present in most areas of the UK, their organisation varies considerably and many are not based in the community, near patients homes. This survey provides a benchmark for comparison with venous ulcer services in other countries.

Foot and ankle injuries in dance.

Acute traumatic injuries are common in ballet dancers. A careful history, thorough examination, and appropriate imaging should allow for the diagnosis of most problems. The clinician must have a high index of suspicion for occult bony injuries, especially if the patient fails to recover as expected. Aggressive treatment of the sprained ankle is essential to maintain foot and ankle mobility and prevent prolonged disability and subsequent overuse injuries. Kinetic chain dysfunctions are common in ballet dancers with overuse injuries and commonly follow ankle sprains. They may represent a secondary phenomenon that developed in response to the compensatory movement changes caused by the initial injury. It is important to remember, however, that these dysfunctions may have been long standing and a causative factor in the injury. Regardless of the time of onset of the dysfunction, residual kinetic chain dysfunction associated with incomplete rehabilitation of an injury may predispose the dancer to further injuries. Untreated dysfunctions at one site in the kinetic chain may predispose to compensatory dysfunction at other sites in the chain. Accordingly, it is essential to thoroughly examine the entire chain for functional movements when dealing with an injury, because identification and treatment of the kinetic chain dysfunction is important in the rehabilitation of the dancing athlete. Kinetic chain dysfunctions are common in injured ballet dancers and may be a cause of repeated injury. Why then are these dysfunctions left untreated? Medical personnel caring for dancers are sometimes guilty of tunnel vision, and focus solely on the injured site without considering what is happening at other sites in the kinetic chain. This oversight is compounded when the physicians or therapists are satisfied with discovering simply what injury has occurred rather than asking why the injury has occurred. The significance of kinetic chain dysfunctions is only just beginning to be recognized, and many examiners are not aware of the relationship between abnormal motion and injury. Generally, people see only what they look for, and they look only for what they know. Kinetic chain dysfunctions can easily be detected with simple tests of functional movement if the examiners include these tests in their assessment of the injured dancer. As long as clinicians are either unaware of or unwilling to perform these tests, these dysfunctions will remain untreated and may put the dancer at risk of failed rehabilitation or predispose them to further injury.

Tepoxalin enhances the activity of an antioxidant, pyrrolidine dithiocarbamate, in attenuating tumor necrosis factor alpha-induced apoptosis in WEHI 164 cells.

The nuclear transcription factor-kappaB (NF-kappaB) and free radicals are known to be involved in apoptosis. We studied the effects of a series of di-aryl-substituted pyrazole NF-kappaB inhibitors including tepoxalin on tumor necrosis factor alpha (TNFalpha)-induced apoptosis in murine fibrosarcoma WEHI 164 cells. We found that potent inhibitors of NF-kappaB were also effective in attenuating apoptosis. WEHI 164 cells that had been dually treated with tepoxalin and the antioxidant pyrrolidine dithiocarbamate (PDTC) were significantly protected from TNFalpha-induced killing. To study the role of free radicals in mediating TNFalpha-induced apoptosis, stable WEHI 164 cells overexpressing Bcl-2, an antioxidant protein, were generated. These cells were protected from TNFalpha-induced apoptosis and neither tepoxalin nor PDTC provided further significant protection. These results suggest that Bcl-2, PDTC, and tepoxalin may attenuate apoptosis in this system by affecting the same signaling pathway or converging pathways. Because tepoxalin suppresses the release of free radicals, PDTC scavenges free radicals and Bcl-2 is an antioxidant protein, free radicals are among the key mediators of this TNF-induced killing event. Tepoxalin and antioxidants may be useful in developing new therapeutics for treating neurodegenerative diseases, autoimmune deficiency syndrome, and ischemia-reperfusion injuries.

Trauma recurrence in the pediatric emergency population.

BACKGROUND: A link between ambulatory trauma and trauma recurrence may be hypothesized: (a) children who present for trauma may be those with characteristics that place them at particular risk; (b) children who frequently present for trauma may be those who present because of parental anxiety; and (c) children may learn from traumatic episodes and therefore decrease their trauma risk. METHODS: A retrospective chart review of billing data was conducted to determine the value of the emergency department trauma visit count as a predictor of future trauma visits and to validate one of the models described above. RESULTS: Records for 16,994 patients were obtained for the period January 1 through December 31, 1997, and these patients were followed as a cohort through the first 6 months of 1998. There were 9,236 males and 7,757 females. Of these, 5,413 patients had a trauma visit count in 1997 (TC97) of at least 1; 120 patients had a TC97 of 3 or more. The relationship between TC97 and trauma visit count in 1998 was linear, present in both males and females and across all age categories, with odds ratios for the group with TC97 > or =3 ranging from 3.8 to 6.4. Conversely, the risk of presentation with respiratory tract infection during the study period in 1998 was not increased with higher TC97 values. CONCLUSION: Our data support the theory that children who present with trauma tend to be those with extrinsic or intrinsic factors that increase trauma risk. We have demonstrated a direct relationship between emergency department visit counts for trauma with future trauma occurrence risk. This relationship is present at all ages and in both males and females. The emergency trauma visit count is an easily obtained measure that has merit as an index to identify higher-risk children for possible intervention strategies. Further work is needed.

Minimal interaction between fluoxetine and multiple-dose zolpidem in healthy women.

The objective was to evaluate possible pharmacokinetic and pharmacodynamic interactions for repeated nightly zolpidem dosing with fluoxetine. Twenty-nine healthy female volunteers (mean age, 25. 6 years) received zolpidem (10 mg) and fluoxetine (20 mg) in the following open design: zolpidem on night 1 followed by 1 washout day, a daily morning dose of fluoxetine on days 3 through 27, and a morning dose of fluoxetine plus an evening dose of zolpidem on days 28 through 32. Plasma levels of zolpidem, fluoxetine, and norfluoxetine were determined at the transitions from one regimen to the next. Morning psychomotor tests were performed on days 1, 2, 28, 29, and 33. Steady-state plasma concentrations of fluoxetine/norfluoxetine were reached by day 24 of fluoxetine dosing. No significant differences in any pharmacokinetic parameters for fluoxetine and norfluoxetine were observed between day 27 and day 32. There were no significant differences in AUC, maximal plasma concentration, or time to maximal concentration parameters for zolpidem plasma concentrations among nights 1, 28, and 32. There was a statistically significantly increased t1/2 for zolpidem on night 32, compared with night 28 (3.64 and 3.29 hr, respectively). There were no significant differences in the next-morning Digit Symbol Substitution Test performance at any time in the study. Both zolpidem and fluoxetine were well tolerated alone or during coadministration. These findings indicate the absence of clinically significant pharmacokinetic or pharmacodynamic interactions between fluoxetine and zolpidem (five consecutive doses) when the drugs are coadministered to healthy women. Therefore, based on these observations, short-term cotherapy with fluoxetine (20 mg) and zolpidem (10 mg) appears safe.

An alternative method of treatment for metacarpal fractures in athletes.

OBJECTIVE: To determine whether closed metacarpal fractures (MCFs) without evidence of shortening, displacement, rotational deformity, or unacceptable angulation could be successfully treated with a modified functional casting technique (glove cast). DESIGN: Retrospective chart review with a follow-up telephone survey to determine long-term outcome. SETTING: Outpatient sport medicine clinic. PATIENTS: 24 patients (22 male, 2 female, average age 15 +/- 1.7 years) with 25 MCFs resulting from sports participation. INTERVENTION: The casting technique consisted of either a fiberglass cast (22 patients) or orthoplast splint (2 patients), allowing full range of motion of the wrist. Patients were allowed to return to modified activity after initial evaluation and immobilization and full athletic activity upon radiographic demonstration of circumferential callus at the fracture site. MAIN OUTCOME MEASURES: Fracture healing, return to participation, complications, reinjury rates, residual symptoms, functional status, and patient satisfaction. RESULTS: Duration of immobilization was 4-5 weeks. All patients demonstrated radiographic evidence of union of the fracture site. All patients had returned to limited sports participation by 2 weeks and full participation by 4 weeks, with no complications, and no reinjuries occurred during the treatment period. Long-term follow-up at an average of 17.2 months showed no functional restrictions and good overall patient satisfaction. CONCLUSIONS: This method of immobilization demonstrates an acceptable alternative for specific, stable MCFs in athletes. Advantages include maintenance of wrist and forearm function during the period of immobilization, protection from reinjury, maximization of sports participation, and a high patient satisfaction rate.

Nocturnal dosing of a novel delivery system of verapamil for systemic hypertension. Verapamil Study Group.

To evaluate the efficacy and safety of a novel delivery system of physiologic pattern release (PPR)-verapamil administered nocturnally to patients with stages I and II hypertension using ambulatory blood pressure (BP) monitoring, we performed a multicenter (17 centers), double-blind, randomized, placebo-controlled, parallel-group trial with placebo and 120, 180, 360, and 540 mg of verapamil in 287 randomized patients. The delivery system has a delay in the release of verapamil for 4 to 6 hours, and then delivers the drug from an osmotic pumping system for approximately 12 hours. Patients were dosed at 10 P.M. The primary end point was change from baseline in trough diastolic BP assessed by ambulatory BP monitoring from 6 to 10 P.M. after 8 weeks of therapy, whereas secondary measures included changes from baseline in peak, early morning (6 to 10 A.M.) systolic and diastolic BP, trough clinic BP, and 24-hour average daytime (8 A.M. to 8 P.M.) and nighttime (8 P.M. to 8 A.M.) BP. The 180, 360, and 540 mg verapamil doses achieved statistically significant reductions in trough (6 to 10 P.M.) diastolic BP (-3.9 +/- 1.0, -7.8 +/- 1.2, and -10.6 +/- 1.1 mm Hg, respectively). Reductions in peak early morning (6 to 10 A.M.) diastolic BP were greater (-4.6 +/- 0.9, -13.3 +/- 1.2, and -19.0 +/- 1.2, for 180, 360, and 540 mg, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)

The immunostimulatory compound 7-allyl-8-oxoguanosine (loxoribine) induces a distinct subset of murine cytokines.

C8- and N7, C8-substituted guanine ribonucleosides comprise a class of molecules with potent immunostimulatory activity for a variety of humoral and cellular immune responses. Although it has been suggested that the immunostimulatory activity may be partially mediated by cytokine production, to date there has been no systematic evaluation of the spectrum of cytokines elicited by these nucleosides. In this study, we examine the cytokines produced by murine spleen cells in response to the di-substituted guanosine analog loxoribine (7-allyl-8-oxoguanosine). First, the levels of cytokine mRNA in spleens from vehicle- or loxoribine-treated mice were compared using PCR analysis with a panel of cytokine-specific primers. Enhancement of IL-1 alpha, TNF-alpha, TNF-beta, IL-6, IFN-alpha, and IFN-gamma mRNA was seen in the spleens of loxoribine-treated mice. IL-12 mRNA responses were more complex, with an increase in the p40 chain and a decrease in the p35 chain. In contrast, no increase was seen for mRNA levels of IL-2, IL-3, IL-4, IL-5, IL-7, or GM-CSF. ELISA assays on the supernatants of loxoribine-treated spleen cells demonstrated that IL-1 alpha, IL-6, TNF-alpha, and IFN-gamma were all produced in a dose-dependent fashion with TNF-alpha produced first, followed by IL-6 and IFN-gamma, and last by IL-1 alpha. IFN-alpha beta activity rose as quickly as TNF-alpha, leveling off at 8 to 12 hr, and was supplanted by a later-occurring surge of IFN-gamma production. IL-1 alpha, IL-6, TNF-alpha, and IFN-gamma were also detected in the sera of mice injected with loxoribine. When antibodies against the relevant cytokines were tested, only anti-IFN-alpha beta inhibited NK activity or lymphocyte proliferation and, in both cases, activity was partially restored by the addition of exogenous IFN-alpha beta. Taken together, these data indicate that loxoribine induces the production of a selective cohort of cytokines all of which have been shown to have immunostimulatory activity. However, only IFN-alpha beta appears to play a role in the enhancement of NK activity and lymphocyte proliferation.

Novel intracellular signaling function of prostaglandin H synthase-1 in NF-kappa B activation.

Many potent nonsteroidal antiinflammatory drugs (NSAIDs) exert their effects by inhibiting the cyclooxygenase activity of prostaglandin H synthase-1 (PGHS1, thus disrupting prostaglandin biosynthesis. However, these drugs do not block the activation of NF-kappa B, an inducible transcription factor which regulates numerous inflammation-related genes. Here we demonstrate that PGHS1 peroxidase, a NSAID-insensitive activity of PGHS1, mediates NF-kappa B activation through an intracellular reactive oxygen signaling pathway. Overexpression of PGHS1 strongly potentiated NF-kappa B activation by phorbol esters and dramatically elevated the generation of intracellular reactive oxygen species (ROS) in response to low concentrations of t-butyl peroxide. Both functions were dependent on PGHS1 peroxidase activity and could be suppressed by the potent antioxidant pyrrolidine dithiocarbamate. In contrast, elimination of PGHS1 cyclooxygenase activity by NSAIDs or site-directed mutagenesis failed to block ROS production or NF-kappa B activation. Thus, PGHS1 peroxidase serves an intracellular signaling function leading to NF-kappa B activation, separable from its role in prostaglandin synthesis.