Twice-daily Subcutaneous Injections Research Articles

Prolactin Suppresses GnRH but Not TSH Secretion

Background/Aims: In animal models, prolactin increases tuberoinfundibular dopamine turnover, which has been demonstrated to suppress both hypothalamic GnRH and pituitary TSH secretion. To test the hypothesis that prolactin suppresses GnRH and TSH secretion in women, as preliminary evidence that a short-feedback dopamine loop also operates in the human, the effect of hyperprolactinemia on GnRH and TSH secretion was examined. Methods: Subjects (n = 6) underwent blood sampling every 10 min in the follicular phase of a control cycle and during a 12-hour recombinant human prolactin (r-hPRL) infusion preceded by 7 days of twice-daily subcutaneous r-hPRL injections. LH and TSH pulse patterns and menstrual cycle parameters were measured. Results: During the 7 days of r-hPRL administration, baseline prolactin increased from 16.0 ± 3.0 to 101.6 ± 11.6 µg/l, with a further increase to 253.7 ± 27.7 µg/l during the 12-hour infusion. LH pulse frequency decreased (8.7 ± 1.0 to 6.0 ± 1.0 pulses/12 h; p < 0.05) with r-hPRL administration, but there were no changes in LH pulse amplitude or mean LH levels. There were also no changes in TSH pulse frequency, mean or peak TSH. The decreased LH pulse frequency did not affect estradiol, inhibin A or B concentrations, or menstrual cycle length. Conclusion: These studies demonstrate that hyperprolactinemia suppresses pulsatile LH secretion but not TSH secretion and suggest that GnRH secretion is sensitive to hyperprolactinemia, but that TSH secretion is not. These data further suggest that the degree of GnRH disruption after 7 days of hyperprolactinemia is insufficient to disrupt menstrual cyclicity.

Experimental study of bone formation around a titanium rod with β- tricalcium phosphate and prostaglandin E2 receptor agonists

ß-Tricalcium phosphate (ß-TCP) is an excellent bone-filling material that is completely absorbed by the body and replaced by autologous bone. Unfortunately, its mechanical strength is low, rendering its application at loaded regions difficult. The purpose of this study is to evaluate the histological and mechanical effects of single and combined use of ß-TCP and EP4 agonist on bone formation around a titanium rod. ß-TCP was loaded into the femoral bone marrow from the distal end of the femur, where the titanium implants were inserted, and the animals received twice-daily subcutaneous injections of EP4 agonist. Group I received the rod only and was designated the control group; group II received EP4 agonist only; group Ill received ß-TCP only; and group IV received both ß-TCP and EP4 agonist. Examination of decalcified specimens revealed favorable bone formation in all treatment groups compared with that in group I, with the most active bone formation seen in group IV. Mechanical evaluation revealed significant differences in maximum pull-out force compared with group I at weeks 4 and 8. There were no differences between groups II and Ill at either week 4 or 8, but the values seen in group IV at weeks 4 and 8 were significantly higher compared with the other groups. Combined use of ß- TCP and EP4 agonist is expected to compensate for bone defects resulting from revision total joint arthroplasty and to achieve stability at an early stage.

Continuous periaortic infusion improves doxycycline efficacy in experimental aortic aneurysms

Objective We created a novel continuous infusion system to evaluate the efficacy of juxta-aortic doxycycline delivery as a transitional step toward developing hybrid drug/device treatment strategies for abdominal aortic aneurysm (AAA) disease. Methods Controlled comparison of treatment outcomes was studied in animal models with molecular and morphologic tissue analysis in a collaboration between university and corporate research laboratories. Rat AAAs were created via porcine pancreatic elastase (PPE) infusion and grouped and analyzed by subsequent treatment status (either doxycycline in vehicle or vehicle alone) and drug delivery method (continuous infusion via periaortic delivery system [PDS] or twice-daily subcutaneous injection). The main outcome measures were AAA diameter via direct measurement, medial elastin lamellar preservation via light microscopy, mural smooth muscle cell (SMC) proliferation and SMC and macrophage density via immunostaining and counting, expression of matrix metalloproteinases 2, 9, and 14 and tissue inhibitors of metalloproteinases 1 and 2 via real-time reverse transcriptase–polymerase chain reaction, and enzymatic activity via substrate zymography. Serum drug levels were analyzed via liquid chromatography/mass spectroscopy. Results PDS (1.5 mg/kg/day) and subcutaneous (60 mg/kg/day) delivery methods caused comparable reductions in AAA diameter during the period of 14 days after PPE infusion. PDS rats gained more weight during the postoperative period ( P < .001), possibly as a result of reduced serum drug levels and systemic toxicity. Doxycycline treatment reduced AAA macrophage infiltration and SMC proliferation significantly. Despite reduced diameter, circumferential elastic lamellar preservation was not apparent in doxycycline-treated AAAs. Conclusions Continuous periaortic infusion lowers the effective doxycycline dose for experimental AAA limitation. Alternative biologic inhibition strategies might also be amenable to direct intra-aortic or juxta-aortic delivery. Periaortic infusion might improve the clinical outcome of minimally invasive AAA treatment strategies. Clinical relevance Aneurysm remodeling may continue after successful endovascular AAA exclusion. Continued proteolytic activity within the aneurysm wall potentiates late graft migration and failure. The doxycycline infusion system developed in these experiments may serve as a prototype for adjuvant treatment modalities that complement endovascular AAA exclusion. Local delivery of doxycycline or other agents active in AAA disease, either continuously or at selected intervals after graft implantation, may stabilize the wall and aid in maintaining aneurysm exclusion. Alternative delivery methods could include passive diffusion from either the graft material itself or treatment reservoirs incorporated into endografts. Given the recognized limitations of current technologies, adjuvant biologic therapies have the potential to improve long-term patient outcome significantly after endovascular exclusion.

Short-term adolescent nicotine exposure in rats elicits immediate and delayed deficits in T-lymphocyte function: critical periods, patterns of exposure, dose thresholds.

Prenatal nicotine exposure elicits lasting deficiencies in T-lymphocyte mitogenesis, and the period of vulnerability extends into adolescence, the stage at which smoking typically commences. We explored the importance of nicotine exposure patterns (continuous infusion vs. repeated subcutaneous injections), dose-effect relationships, and specificity of the effects. Adolescent rats were given nicotine infusions for 1 week beginning on postnatal day (PN) 30, using a regimen (6 mg/kg/day) that produces plasma nicotine levels (25 ng/ml) similar to those in smokers; another group received 2 mg/kg/day. At the end of the infusion period (PN37), T-lymphocyte mitogenic responses to concanavalin A were deficient in the group receiving 6 mg/kg/day; values for the 2 mg/kg/day group were intermediate between controls and the 6 mg/kg/day group. One week after the termination of nicotine treatment, responses returned to normal, only to reemerge in young adulthood (PN65), at which stage adverse effects were significant even for the group that received 2 mg/kg/day. In contrast to the T-cell alterations, B-lymphocyte responses were unaffected. Administering the same total doses of nicotine by twice-daily subcutaneous injections over the 1-week treatment period (1 or 3 mg/kg per injection) did not evoke deficits in responses of either T-cells or B-cells, even though the high dose produced overt systemic toxicity and persistent weight loss. Our results indicate that adolescent nicotine exposure, even at levels below those associated with active smoking, elicits selective deficits in T-lymphocyte function. Although short-term adaptations may correct the effects, deficits reemerge in young adulthood despite cessation of nicotine exposure.

Long-Term Treatment with Subcutaneous T-20, a Fusion Inhibitor, in HIV-Infected Patients: Patient Satisfaction and Impact on Activities of Daily Living

T-20 is a novel antiretroviral agent that inhibits the fusion of human immunodeficiency virus (HIV) with target cell membranes. It is delivered by self-administered, twice-daily, subcutaneous injections. The impact of this mode of administration on patients' ability to conduct normal activities of daily living (ADL) and comply with a T-20 treatment regimen was assessed as part of a 48-week, phase 2 trial (T20-205). Patients' opinions on the impact of T-20 on ADL, ease of use of T-20, and choice to continue with T-20 were assessed by two questionnaires completed at baseline and week 48 (or study withdrawal). ADL were measured using a Likert-type scale based on established instruments with questions added to assess HIV-specific issues. Seventy previously treated patients received T-20 in combination with an average of five oral antiretroviral agents. Relative to other HIV/AIDS drugs, T-20 had little impact on ADL, with the majority of patients (54%-96%) agreeing (somewhat or strongly) that subcutaneous injections had not limited ADL. Patients found the injections relatively easy to perform with more than 47% of patients stating that each aspect of the injections (ease of injection, storage, reconstitution, and disposal of sharps) were very easy or easy. If medically indicated, 98% of patients stated that they would choose to continue with T-20. The most common reasons for this were the perceived effectiveness of T-20 and lack of side effects. In conclusion, the need to deliver T-20 via twice-daily subcutaneous injections was not considered an important barrier by HIV-positive patients seeking improvement or stabilization of their condition.

Enoxaparin: an update of its clinical use in the management of acute coronary syndromes.

Enoxaparin (enoxaparin sodium) is a low molecular weight heparin (LMWH) indicated for use in the treatment of ischaemic complications of unstable angina and non-Q wave myocardial infarction (MI). Unfractionated heparin (UFH) has for many years represented the standard in anticoagulant therapy for patients with acute coronary syndromes; however, recent studies suggest that enoxaparin is also a viable option for anticoagulant therapy in these patients. The ESSENCE (Efficacy and Safety of Subcutaneous Enoxaparin in Non-Q wave Coronary Events) and the TIMI 11B (Thrombolysis in Myocardial Infarction) studies reported that twice daily enoxaparin was significantly more effective than a continuous infusion of UFH in reducing the composite triple endpoint of death, MI, or recurrent angina or urgent revascularisation. Follow-up of both patient populations showed continued benefit associated with enoxaparin. Enoxaparin has been compared with tinzaparin in the treatment of unstable coronary artery disease using a nonblind study design. There was no difference between treatment groups in the therapeutic endpoints. Three nonblind studies have also compared the effects of enoxaparin and UFH in patients receiving thrombolytic therapy following acute MI. The HART II (Heparin and Aspirin Reperfusion Therapy), the ASSENT 3 (Assessment of the Safety and Efficacy of a New Thrombolytic Regimen) and the ENTIRE-TIMI 23 (Enoxaparin and Tenecteplase with or without glycoprotein IIb/IIIa Inhibitor as Reperfusion strategy in ST Elevation MI - Thrombolysis in Myocardial Infarction) studies have revealed that enoxaparin in combination with alteplase or tenecteplase is at least equivalent (HART II and ENTIRE-TIMI 23), and possibly superior (ASSENT 3) to UFH. Enoxaparin is administered as a twice-daily subcutaneous injection. In contrast, UFH is administered as an intravenous infusion which requires routine monitoring of the activated partial thromboplastin time to ensure adequate levels of anticoagulation are maintained. During the acute phase of the the ESSENCE and TIMI 11B studies, the incidence of major bleeding was similar in patients receiving enoxaparin to that in patients receiving UFH. In contrast, the rates of minor bleeding were higher in patients receiving enoxaparin than in those receiving UFH throughout these studies. Data from the ESSENCE, TIMI 11B and ASSENT 3 studies have prompted calls for those LMWHs which have been shown to be superior to UFH, to be considered as first choice treatment for anticoagulation in unstable coronary syndromes. To date, these suggestions are not reflected in current guidelines which consider UFH and LMWHs equally. Irrespective, the clinical data reported in this review support the use of enoxaparin in the treatment of acute coronary syndromes. These data suggest that enoxaparin shows certain clinical and practical advantages over standard treatment with UFH and represents an important development in the treatment of acute coronary syndromes.

Outpatient DVT Management by the Emergency Department

Outpatient management of proximal lower-extremity deep venous thrombosis (DVT) may be an economical alternative to traditional inpatient management. Outpatient management involves daily or twice-daily subcutaneous injections of low-molecular-weight heparin; inpatient management typically entails continuous infusion of unfractionated heparin. In both cases, satisfactory oral anticoagulation is achieved with warfarin. Patients diagnosed with DVT in the …

Safety and efficacy of subcutaneous bolus injection of deferoxamine in adult patients with iron overload

We compared 48-hour urinary iron excretion after a twice-daily subcutaneous bolus injection of deferoxamine and after 12 hours of subcutaneous continuous infusion of the drug in 27 patients with iron overload (mean age, 55.7 years). In most patients, the iron overload was due to multiple transfusions administered during chemotherapy or as part of supportive care for a hematologic or oncologic disorder. One patient had sickle cell anemia and 1 had hereditary hemochromatosis and spherocytosis. Similar urinary iron excretion was observed with the 2 methods of administration; mean ± SD values were 6935.3 ± 3832.3 μg/48 hours with subcutaneous bolus injection and 6630.4 ± 3606.9 μg/48 hours with subcutaneous continuous infusion (P = .3). Twenty-six patients (96.3%) chose to continue therapy with bolus injection. The long-term efficacy of bolus injection was evaluated by measuring the serum ferritin concentration at regular intervals for a follow-up time of 20.1 ± 4.5 months. Ferritin concentration decreased to below 1000 μg/L in 73% of the patients and to below 500 μg/L in 42% and became normal in 26%. Best results were obtained in patients who were no longer receiving blood transfusions when chelation therapy was initiated. Three of 26 patients (11.5%) had mild, transient side effects after bolus injection. Larger prospective, randomized studies must be conducted before deferoxamine bolus injection can be routinely recommended for patients with iron overload.

Synergistic effects of insulin-like growth factor-I and human chorionic gonadotropin in the rat ovary

Insulin and low doses of lutenizing hormone (LH) activity (human chorionic gonadotropin [hCG]) act synergistically in the rat to produce anovulation, large ovarian cysts, and elevated plasma androstenedione levels. Further, both insulin and insulin-like growth factor-I (IGF-I) affect the ability of gonadotropins to enhance both ovarian theca and granulosa cell function in vitro. The present series of experiments were performed to determine if recombinant human IGF-I (rhIGF-I) can act in a manner similar to insulin when combined with subovulatory doses of hCG in adult normally cycling rats. Fifty-four female Sprague-Dawley rats were randomly assigned to the following treatment groups at the age of 64 days: (A) vehicle alone (controls, phosphate-buffered saline containing 0.09% pig gelatin), (B) twice-daily subcutaneous injections of 0.5 to 3.0 U insulin, (C) twice-daily subcutaneous injections of 1.5 U hCG, (D) both insulin and hCG, (E) twice-daily subcutaneous injections of rhIGF-I (2.5 mg/kg/d), and (F) both hCG and rhIGF-I. After 22 days of treatment, the animals were killed on day 23, trunk blood was collected, and the ovaries were excised for histological study. Eight of 9 control rats and 5 or 6 of 9 rats treated with insulin, hCG, or rhIGF-I alone displayed normal estrus cycles throughout the in vivo treatment period as assessed by daily vaginal smears. In marked contrast, only 1 animal treated with hCG + insulin and 2 animals treated with hCG + rhIGF-I continued to display vaginal smears indicative of normal cycling. Multiple large ovarian follicular cysts were found only in these latter 2 groups (3 of 9 animals in each group). Mean serum testosterone levels were significantly elevated in animals receiving insulin + hCG (0.72 +/- 0.28 v 0.17 +/- 0.03 ng/mL in controls, P = .05). Mean serum androstenedione levels were significantly elevated in animals receiving hCG and animals receiving rhIGF-I + hCG (5.57 +/- 0.99 and 2.39 +/- 0.68 ng/mL, respectively, v0.14 +/- 0.14 ng/mL in controls, P< .01 and P< .05, respectively). We conclude that rhIGF-I and insulin act synergistically with subovulatory doses of hCG to disrupt normal reproductive cycling, elevate serum androgen concentrations, and induce large ovarian cysts in intact adult rats.

A randomized trial of the effect of low molecular weight heparin vs. warfarin on mortality in the long-term treatment of proximal vein thrombosis

It has been shown that several different low molecular weight (LMW) heparins, given by a once- or twice-daily subcutaneous injection without laboratory monitoring, are as effective or even more effective than continuous intravenous, unfractionated heparin monitored by the activated partial thromboplastin time (APTT). One such study demonstrated a statistically significant decrease in mortality over the 3 month follow-up study. This was particularly striking in patients with cancer. Based on these findings, a multicentre, randomized clinical trial has been designed to compare the effect of long-term, once-daily LMW heparin with standard treatment using heparin and warfarin to ensure that equal numbers of cancer patients are in both groups. This will demonstrate whether or not the long-term use of LMW heparin can have a significant impact on mortality in patients who have proximal venous thrombosis with or without cancer.

Distribution of cocaine and metabolites in the pregnant rat and fetus in a chronic subcutaneous injection model.

We examined the distribution of cocaine and its metabolites benzoylecgonine (BE) and norcocaine (NOR) in pregnant Sprague-Dawley rats and fetuses following twice-daily subcutaneous (s.c.) injections of 20 mg/kg cocaine HCl from gestational day (GD) 8 through GD 20. On GD 21, the animals received a single injection and maternal trunk blood, fetal blood, fetal brains, and amniotic fluid were collected 5 min, 15 min, 30 min, 1 h, 2 h, 4 h, 8 h, or 12 h later for cocaine and metabolite analyses by high-performance liquid chromatography (HPLC) with UV detection. The highest concentrations of cocaine and BE were detected in maternal plasma at 1 h and 4 h respectively. Cocaine peaked at 2 h and BE at 4 h in both fetal plasma and brain. In amniotic fluid, cocaine levels peaked at 2 h, but the highest BE levels were found at 8 h postinjection. An additional group of chronically treated dams was given both cocaine injections on GD 21 and sacrificed 2 h later. Benzoylecgonine concentrations were increased in fetal plasma, fetal brain, and amniotic fluid when compared with the 2-h results following a single cocaine treatment. Moreover, NOR, which had not been previously detected, was now measurable in the amniotic fluid.

Placental development and fetal growth in growth hormone-treated ewes.

The effects of recombinant bovine growth hormone (bGH) treatment of pregnant ewes on maternal metabolism, placental development and fetal growth were examined in two studies. In a preliminary study (experiment one), single-bearing ewes were treated by twice-daily subcutaneous injection for 7 days with bGH (n = 8) at a dose of 0.15 mg/kg LW/day or with saline (n = 8) between days 101 and 107 of gestation inclusive. In experiment two, single- and twin-bearing ewes were treated for 14 days with bGH (0.15 mg/kg L W/day) (n = 10) or saline (n = 10) between days 70 and 83 or days 98 and 111 of gestation inclusive. Ewes were killed on the day following termination of bGH treatment and fetal and placental measurements recorded. Maternal plasma concentrations of GH, IGF-I and insulin were higher (P < 0.001) in bGH-treated ewes relative to saline-treated ewes in both experiments. Consistent across experiments was an increase (P < 0.05) in the weight of the myoendometrium in bGH-treated ewes. Treatment with bGH also increased the total weight of the gravid uterus (P < 0.05) in both experiments. Weights of the uterine fluids were increased by bGH in experiment one (P < 0.05), but an effect of the same magnitude could not be repeated in experiment two. In experiment one, there was a tendency towards increased mean fetal body weights after growth hormone treatment, although the effect was non-significant. In experiment two, treatment with bGH was associated with significantly (P < 0.05) higher fetal weights, but only at the later stage of gestation (day 112). This effect was additive with that of fetal rank. Exogenous bGH treatment had little discernible effect on measures of placental size. It is concluded that administration of exogenous bGH to pregnant ewes can stimulate fetal growth, but only after about day 100 of gestation. This response seems most likely to reflect changes in maternal nutrient partitioning or placental function, rather than placental size. These studies suggest a role for growth hormone of maternal or placental origin in the regulation of fetal growth.

Does desensitization to hexarelin occur?

Hexarelin, a potent growth hormone (GH)-releasing peptide, is capable of causing profound GH release in normal individuals. If the GH response to hexarelin in humans becomes appreciably attenuated following long-term administration, this would seriously limit the potential therapeutic use of hexarelin and similar agents. The effect of twice-daily subcutaneous injections of hexarelin on GH release was therefore investigated over a period of 16 weeks in 12 healthy elderly individuals. The mean (+/- SEM) areas under the GH curve (AUCGH) at weeks 0, 1, 4 and 16 were 19.1 +/- 2.4, 13.1 +/- 2.3, 12.3 +/- 2.4 and 10.5 +/- 1.8 microg/l/hour, respectively. There was a significant change in AUCGH over the study period (P = 0.0003). Further analysis showed that the decreases in AUCGH at weeks 4 and 16 were significant (P < 0.05 and P < 0.01, respectively) compared with baseline values. Four weeks after completion of the 16-week study period, hexarelin was again administered. On this occasion, AUCGH increased significantly compared with that at week 16 (from 10.5 +/- 1.8 to 19.4 +/- 3.7 microg/l/hour; P < 0.05) and was not significantly different compared with that at week 0. These results show that attenuation of the GH response after long-term hexarelin therapy is partial and reversible.

Evaluation of a new method of administration of the iron chelating agent deferoxamine

We compared the effectiveness of deferoxamine administered by twice-daily subcutaneous injections with conventional administration by prolonged subcutaneous infusion in 20 patients with thalassemia. Urinary iron excretion was comparable with the two methods but decreased significantly when the total daily dose was administered as a single injection. Local reactions were similar with infusion and injection. Subcutaneous injections of deferoxamine may be considered as an alternative to conventional infusions. (J Pediatr 1997;130:86-8)

Prolonged Antithrombin Activity of Low-Molecular-Weight Heparins

The mechanism for the efficacy of once- or twice-daily subcutaneous injections of low-molecular-weight heparins (LMWHs) for the treatment of venous thromboembolism has been difficult to explain. The confusion exists because the observation from experimental studies that the antithrombin activity of LMWHs is necessary for their antithrombotic effect is inconsistent with the reported short half-life of the antithrombin activity of LMWHs. Previous pharmacokinetic studies were performed with lower doses of LMWHs than have been used in contemporary trials, and antithrombin activity was assessed with the barely sensitive chromogenic assay. We performed a pharmacokinetic study to compare the relative half-lives of prophylactic and therapeutic doses of LMWHs assessing antithrombin activity with both the chromogenic and a more sensitive assay (plasma thrombin neutralization assay). An eight-way cross-over randomized study in healthy volunteers was performed. Enoxaparin (20 and 40 mg and 1 and 2 mg/kg) and nadroparin (7500 and 10,000 ICU and 225 and 450 ICU/kg) were administered subcutaneously. The maximal peak activity for aPTT ratio was 1.7. A dose-dependent peak activity was found for both antifactor Xa and antithrombin activities. Disappearance time of these activities after the highest dose of both LMWHs was longer than 16 hours. Overall mean antifactor Xa activity half-life was 4.6 hours. Overall mean antithrombin activity half-life was longer than 4 hours. Our results provide an explanation for the effectiveness of LMWHs administered either once or twice daily. High and sustained plasma antithrombin activity is achieved when LMWHs are administered in therapeutic doses used in contemporary trials with only a moderate prolongation of the aPTT.

Drug Treatment Associated with Heart Valve Replacement

This article reviews a number of specific pharmacological considerations for patients with prosthetic heart valves. All patients with mechanical heart valves should be anticoagulated. In the past, an International Normalised Ratio (INR) of 2.5 to 4.5 has been recommended. Recent nonrandomised studies have suggested that a patient with a prosthetic valve who is at low risk for thromboembolic events could have an INR ranging from 1.8 to 3.5. The lower end of this range should only be used for patients at higher than average risk of haemorrhage, until randomised data show that levels below 2.5 may be applied universally. In high-risk patients (particularly those with previous thromboembolic events) low dose aspirin should be added. During noncardiac surgery, a patient at low risk for thromboembolic events could be managed by discontinuing anticoagulation 3 days before the operation, with warfarin recommenced as soon as possible afterwards. Perioperative heparinisation would be appropriate in a higher risk patient. Women with prosthetic heart valves wishing to become pregnant should be converted to the use of twice-daily subcutaneous heparin injections. Patients with bioprosthetic valves can be managed without anticoagulation unless they have some other reason to require it. Patients at high risk should be treated with aspirin or warfarin. Thrombolytic therapy for acute valve thrombosis should be used for those who are haemodynamically compromised and therefore have a high risk of mortality from operative intervention. All patients with prosthetic heart valves undergoing invasive procedures potentially causing bacteraemia should receive antibiotic prophylaxis for endocarditis. The actual drugs used depend on the likely nature of the bacteraemia, and any possible patient hypersensitivity.

The Effects of Short-Term Treatment with the Prostaglandin E1 (PGE1) Analog Misoprostol on Inflammatory Mediator Production in Murine Lupus Nephritis

MRL-lpr/lpr mice spontaneously develop an autoimmune disease with nephritis similar to human systemic lupus erythematosus. In these animals, treatment with E-series prostaglandins ameliorates renal disease and prolongs survival, perhaps by modulating production of cytokines or eicosanoids. To further define the mechanisms of action of E-series prostaglandins in established murine lupus nephritis, we administered the prostaglandin E1 (PGE1) analog misoprostol to 20-week-old MRL-lpr/lpr mice by twice-daily subcutaneous injection. After 2 days of treatment, misoprostol reduced renal cortical interleukin-1β (IL-1β) mRNA levels compared to vehicle-treated controls (0.19 ± 0.06 (misoprostol) vs 0.50 ± 0.04 (vehicle) densitometry units; P < 0.005). A similar reduction in cortical IL-1β mRNA levels was found in left kidneys harvested from MRL-lpr/lpr mice following 2 days of treatment with misoprostol compared to right kidneys harvested from the same animal prior to the first dose of PGE1 analog (0.12 ± .05 (left) vs 0.39 ± 0.18 (right) densitometry units; P < 0.05). This reduction in cortical IL-1β mRNA levels was not associated with alterations in renal production of thromboxane B2, PGE2, or leukotriene B4 or with significant changes in the severity of renal inflammatory cell infiltrates. Timecourse studies indicated that IL-1β mRNA levels were decreased within 24 hr of initiating misoprostol therapy. This reduction in IL-1β mRNA levels was transient because levels were not reduced after 1 week of treatment with the PGE1 analog. Thus, while the beneficial effects of chronic treatment with E-series prostaglandins in MRL-lpr/lpr mice may be mediated by other mechanisms, these data suggest than an important short-term effect of misoprostol in this model may be suppression of renal cortical IL-1β mRNA levels.

Angiopeptin inhibits oncogene induction in rabbit aorta after balloon denudation.

Angiopeptin, a synthetic cyclic octapeptide analogue of somatostatin, reduces neointimal hyperplasia after balloon denudation of rabbit aorta if administered before injury. The aim of this study was to analyze the effect of angiopeptin pretreatment on the level of expression of the c-fos and c-jun protooncogenes, early markers of smooth muscle cell proliferation, after balloon denudation of rabbit aorta. For histological analysis of the effect of angiopeptin on neointimal thickening after aortic balloon denudation, rabbits were randomized into three groups: group 1 (controls), twice-daily injections of saline begun 24 hours before balloon denudation (n = 9); group 2, twice-daily injections of angiopeptin 10 micrograms/kg begun 24 hours before balloon denudation (n = 9); and group 3, twice-daily injections of angiopeptin 10 micrograms/kg begun 1 hour after balloon denudation (n = 7). The degree of neointimal thickening 28 days after balloon denudation was significantly less in group 2 than in group 1 (neointimal area: group 1, 0.59 +/- 0.11 mm2; group 2, 0.22 +/- 0.05 mm2; P < .05. Neointima/media: group 1, 0.85 +/- 0.17; group 2, 0.23 +/- 0.05; P < .05). When angiopeptin was started 1 hour after denudation (group 3), however, the neointimal area (0.52 +/- 0.09 mm2) and the neointima/media ratio (0.76 +/- 0.10) were not statistically different from the control group. For analysis of protooncogene induction, rabbits received twice-daily subcutaneous injections of saline (n = 7), angiopeptin 10 micrograms/kg (n = 8), or angiopeptin 100 micrograms/kg (n = 4) begun 24 hours before balloon denudation. The animals were killed 30 minutes after balloon denudation, and total aortic RNA was hybridized with fos and jun probes. Expression of c-fos and c-jun was detected 30 minutes after injury; angiopeptin pretreatment at 20 micrograms.kg-1.d-1 induced a 41% reduction in c-fos expression and a 42% reduction in c-jun expression compared with control animals. The inhibitory effect at the higher dose of angiopeptin was similar. Our results show that the inhibitory effect of angiopeptin on neointimal thickening is related to events that occur very early after injury and suggest that the inhibition of smooth muscle cell activation may be responsible, at least in part, for this effect.

Neuropeptide messenger RNA expression in the 6-hydroxydopamine-lesioned rat striatum reinnervated by fetal dopaminergic transplants: Differential effects of the grafts on preproenkephalin, preprotachykinin and prodynorphin messenger RNA levels

In situ hybridization histochemistry was used to analyse the expression of the messenger RNAs encoding for enkephalin, substance P and dynorphin in the striatum of normal rats, rats subjected to a unilateral 6-hydroxydopamine lesion of the mesostriatal dopamine pathway and lesioned rats bearing intrastriatal transplants of fetal nigral neurons. About half of the rats in each group received twice-daily subcutaneous injections of 5 mg/kg apomorphine and the other half received control injections of saline, for nine days. Three hours after the last injection, the rats were killed by decapitation. Cryostat sections through the striatum were incubated with, 35 S-labeled oligodeoxyribonucleotide probes hybridizing with preproenkephalin, preprotachykinin or prodynorphin messenger RNA. One additional series of sections was incubated with [ 3H]GBR 12935 in order to label dopamine uptake sites. Quantitative evaluation of the hybridization signal was performed both at the macroscopic level (autoradiographic film analysis) and at the cellular level (optical density of silver grains over identified cells). The grafted nigral neurons reversed the lesion-induced up-regulation of preproenkephalin messenger RNA in the whole striatal complex. By contrast, the graft-induced effect on the lesion-induced down-regulation of preprotachykinin messenger RNA was restricted to the region of the host striatum where the graft-derived dopamine fibers exhibited their densest distribution (up to 0.5 mm from the border of the grafts). However, following chronic treatment with apomorphine, preprotachykinin messenger RNA expression approached control levels in a wider portion of the grafted striata (up to 1 mm from the border of the grafts). Basal prodynorphin messenger RNA expression, which was also down-regulated in the lesioned striata, was only partially restored by the transplants. Repeated injections of apomorphine enhanced prodynorphin messenger RNA in the lesioned striata to levels several fold higher than normal. This massive increase in prodynorphin messenger RNA expression was completely prevented by the transplants over a large volume of the host striatum (> 1 mm from the graft-host border), but a trend towards an abnormally high prodynorphin messenger RNA expression was still present in peripheral striatal areas that were not reached by graft-derived dopamine fibers. The present results indicate that fetal nigral neurons transplanted to the 6-hydroxydopamine-lesioned striatum have differential effects on the activity of enkephalin-containing (i.e. mainly striatopallidal) and substance P- or dynorphin-containing (i.e. mainly striatonigral) neurons. An inhibitory control over the activity of striatopallidal neurons is completely restored by the grafts, even in non-reinnervated striatal regions, suggesting that neurohumoral mechanisms underlie this effect. A facilitatory control over the activity of striatonigral neurons is restored by the grafts only in the densely reinnervated portion of the host striatum, which indicates that synaptic transmission is likely to mediate this effect. The ability of the grafts to prevent an abnormal up-regulation of prodynorphin messenger RNA expression by apomorphine indicates, however, that graft-derived dopamine release is also sufficient to induce a complete and widespread normalization of dopamine receptor sensitivity on striatonigral neurons.

The Use of an Indwelling Teflon Catheter for Subcutaneous Heparin Administration During Pregnancy

The use of subcutaneous heparin, the therapy of choice for women requiring anticoagulant prophylaxis during pregnancy, is problematic because of the discomfort produced by repeated injections. An indwelling subcutaneous Teflon catheter that can be left in place for 1 week recently became available for use as an entry port for parenteral therapy. Since the use of this catheter has the potential to overcome some of the problems of long-term heparin therapy, we decided to compare this Teflon catheter with twice-daily subcutaneous injections in women requiring heparin during pregnancy. In a randomized, multiple-crossover study, patients alternated every 2 weeks between having heparin administered through the indwelling Teflon catheter and receiving heparin via subcutaneous injections. After each 4-week cycle, patients completed a questionnaire designed to determine their preferred method of heparin administration. The side effects, doses, and anticoagulant activity of heparin with the two delivery systems were also compared. Twelve patients completed one to five 4-week cycles of heparin therapy. Ten of the patients selected the Teflon catheter as the preferred route of heparin administration (P = .04) and 11 patients reported that the catheter caused less pain and bruising than twice-daily subcutaneous injections (P < .01). Five patients developed urticarial reactions at the sites of heparin injections. These reactions tended to be more severe when the Teflon catheter was used, and two women discontinued using the catheter after the first cycle because of this complication. There were no differences in heparin dose requirements or achieved activated partial thromboplastin times between the two routes of heparin administration. Most pregnant women in our study preferred to have subcutaneous heparin administered through an indwelling Teflon catheter rather than by twice-daily injections. Heparin given through the Teflon catheter was bioavailable and caused less local bruising than twice-daily injections. Urticarial reactions to heparin tended to be more severe with the use of the Teflon catheter and resulted in the discontinuation of the device's use in two of 12 patients.