A surgical model for isolating the pig liver in vivo for gene therapy.

Several studies report results that suggest the need of vascularization blocking for efficient gene transfer to the liver, especially in nonviral gene therapy. In this study, we describe a surgical strategy for in vivo isolation of the pig liver, resulting in a vascular watertight organ that allows the evaluation of several gene injection conditions. The hepatic artery and portal, suprahepatic and infrahepatic cava veins were dissected. Then, liver vascularization was excluded for 5-7 min. In that time, we first injected 200 ml saline solution containing the p3c-eGFP plasmid (20 µg/ml) simultaneously through two different catheters placed in the portal and cava veins, respectively. Vital constants were monitored during the surgery to assess the safety of the procedure. Basal systolic/diastolic blood pressures were 92.8/63.2 mm Hg and dropped to 40.7/31.3 mm Hg at the end of vascular exclusion; the mean basal heart rate was 58 bpm, reaching 95 bpm when the blood pressure was low. Oxygen saturation was maintained above 98% during the intervention, and no relevant changes were observed in the ECG tracing. Peak plasma AST (aspartate aminotransferase) and ALT (alanine aminotransferase) levels were observed after 24 h (151 and 57 IU, respectively). These values were higher, but not relevant, in 60 ml/s injection than in 20 ml/s injection. Efficiency of gene transfer was studied with simultaneous (cava and portal veins) injection of eGFP gene at flow rates of 20 and 60 ml/s. Liver tissue samples were collected 24 h after injection and qPCR was carried out on each lobe sample. The results confirmed the efficiency of the procedure. Gene delivery differed between 20 ml/s (9.9-31.0 eGFP DNA copies/100 pg of total DNA) and 60 ml/s injections (0.6-1.1 eGFP DNA copies/100 pg of total DNA). Gene transcription showed no significant differences between 20 ml/s (15,701.8-21,475.8 eGFP RNA copies/100 ng of total RNA) and 60 ml/s (12,014-36,371 eGFP RNA copies/100 ng of total RNA). The procedure is not harmful for animals and it offers a wide range of gene delivery options because it allows different perfusion ways (anterograde and retrograde) and different flow rates to determine the optimal conditions of gene transfer. This strategy permits the use of cell therapy and viral or non-viral liver gene therapy, especially appropriated to a wide variety of inherited or acquired diseases because of the liver's ability to produce and deliver proteins to the bloodstream.

Near-real time flash drought monitoring system and dataset for Spain.

Flash droughts are characterized by rapid development and intensification, which makes early warning and monitoring difficult. Flash drought monitor (FDM) is a near-real time monitoring system for Spain (https://flash-drought.csic.es) based on the Standardized Precipitation Evapotranspiration Index (SPEI). Flash drought identification was based on rapid and anomalous declines in SPEI at a short time scale (1-month). Thus, FDM enables operational tracking of flash drought conditions in Spain at high spatial resolution (1.1 × 1.1 km) and high temporal frequency (weekly). Likewise, to put flash drought monitoring into a temporal context, the FDM also provides weekly flash drought conditions recorded in Spain from 1961 to the present. The FDM is a useful tool for preparedness and mitigation of flash droughts in Spain. Furthermore, the data provided by the FDM could be useful to develop future studies in relation to the flash drought in Spain.

Digital image analysis workflows for evaluation of cell behavior and tumor microenvironment to aid therapeutic assessment in high-risk neuroblastoma.

Digital pathology and artificial intelligence are promising emerging tools in precision oncology as they provide more robust and reproducible analysis of histologic, morphologic and topologic characteristics of tumor cells and the surrounding microenvironment. This study aims to develop digital image analysis workflows for therapeutic assessment in preclinical in vivo models. For this purpose, we generated pipelines that enable automatic detection and quantification of vitronectin and αvß3 in heterotopic high-risk neuroblastoma xenografts, demonstrating that digital analysis workflows can be used to provide robust detection of vitronectin secretion and αvß3 expression by malignant neuroblasts and to evaluate the possibility of combining traditional chemotherapy (etoposide) with extracellular matrix-targeted therapies (cilengitide). Digital image analysis added evidence for the relevance of territorial vitronectin as a therapeutic target in neuroblastoma, since its expression is modified after treatment, with a mean percentage of 60.44% in combined therapy tumors vs 45.08% in control ones. In addition, the present study revealed the efficacy of cilengitide for reducing αvß3 expression, with a mean αvß3 positivity of 34.17% in cilengitide treated material vs 66.14% in control and with less tumor growth when combined with etoposide, with a final mean volume of 0.04 cm3 in combined therapy vs 1.45 cm3 in control. The results of this work highlight the importance of extracellular matrix-focused therapies in preclinical studies to improve therapeutic assessment for high-risk neuroblastoma patients.

[Monitoring medication errors in personalised dispensing using the Sentinel Surveillance System method]. / Monitorización de errores de medicación en dispensación individualizada mediante el método del carro centinela.

OBJECTIVE: To assess the efficacy of a new quality control strategy based on daily randomised sampling and monitoring a Sentinel Surveillance System (SSS) medication cart, in order to identify medication errors and their origin at different levels of the process. METHOD: Prospective quality control study with one year follow-up. A SSS medication cart was randomly selected once a week and double-checked before dispensing medication. Medication errors were recorded before it was taken to the relevant hospital ward. Information concerning complaints after receiving medication and 24-hour monitoring were also noted. Type and origin error data were assessed by a Unit Dose Quality Control Group, which proposed relevant improvement measures. RESULTS: Thirty-four SSS carts were assessed, including 5130 medication lines and 9952 dispensed doses, corresponding to 753 patients. Ninety erroneous lines (1.8%) and 142 mistaken doses (1.4%) were identified at the Pharmacy Department. The most frequent error was dose duplication (38%) and its main cause inappropriate management and forgetfulness (69%). Fifty medication complaints (6.6% of patients) were mainly due to new treatment at admission (52%), and 41 (0.8% of all medication lines), did not completely match the prescription (0.6% lines) as recorded by the Pharmacy Department. Thirty-seven (4.9% of patients) medication complaints due to changes at admission and 32 matching errors (0.6% medication lines) were recorded. The main cause also was inappropriate management and forgetfulness (24%). The simultaneous recording of incidences due to complaints and new medication coincided in 33.3%. In addition, 433 (4.3%) of dispensed doses were returned to the Pharmacy Department. After the Unit Dose Quality Control Group conducted their feedback analysis, 64 improvement measures for Pharmacy Department nurses, 37 for pharmacists, and 24 for the hospital ward were introduced. CONCLUSIONS: The SSS programme has proven to be useful as a quality control strategy to identify Unit Dose Distribution System errors at initial, intermediate and final stages of the process, improving the involvement of the Pharmacy Department and ward nurses.

Assessment of the effectiveness and safety of natalizumab for treating relapsing-remitting multiple sclerosis.

OBJECTIVE: Assessing the effectiveness and safety of natalizumab for treating relapsing-remitting multiple sclerosis in a tertiary hospital. METHOD: Observational, prospective study of adult patients treated with natalizumab from May 2007 until February 2009. TREATMENT: 300 mg natalizumab every four weeks. Response criteria: assessment of disease progression, appearance of flare-ups and assessment of magnetic resonance images. Adverse reactions during treatment with natalizumab were recorded. RESULTS: Thirty patients (73% female); average age 34 ± 8.4 years; mean baseline EDSS 3.4 ± 1.3; number of flare-ups in the past year 2.1 ± 1.2. TREATMENT was discontinued in five patients, due to refusal in one case, ineffectiveness in two cases and anaphylaxis in the other two cases. Fourteen patients completed one year of treatment with satisfactory results. A lower EDSS score by 36%, 47%, 31%, 54% and 28% was obtained at 3, 6, 9, 12 and 15 months of treatment respectively. The prevalence of relapse-free patients was 94%, 76% and 54% at 3, 6 and 12 months. MRI imaging studies in 11 patients one year after they began treatment showed no new lesions. Two patients suffered severe anaphylactic shock and another one had an outbreak of urticaria. The presence of neutralising antibodies was the reason for suspending treatment in 6.6% of the patients. CONCLUSIONS: The treatment's effectiveness and safety in our patient group suggest that natalizumab is a treatment for refractory patients or those with aggressive types of multiple sclerosis, although we do not yet know about its long-term effects and the evolution of the appearance of neutralising antibodies.

Timing of spermatogonial stem cell transplantation affects the spermatogenic recovery outcome in mice.

Spermatogonial stem cell transplantation (SSCT) is a strategy that has demonstrated to be feasible to restore spermatogenesis in animal models when it is performed shortly after the gonadotoxic onset to destroy their endogenous germ cells. However, in the case of boys subjected to fertility preservation, future transplantations will be performed with a delay of many years. In order to study how timing of SSCT affects donor-derived spermatogenic recovery in mice, we compared the percentage of spermatogenic tubule cross-sections within testes of 59 C57BL/6NCrl mice distributed in 6 groups: group 1, untreated mice controls (n = 9); group 2, mice that received a single dose of busulfan 40 mg/kg (n = 10); group 3, mice that received two additional doses of busulfan 10 mg/kg every 5 weeks (n = 10); group 4 (SSCT-A), mice subjected to a standard SSCT performed 5 weeks after a single injection of busulfan 40 mg/kg (n = 10); group 5 (SSCT-B), mice subjected to a delayed SSCT performed 15 weeks after a single injection of busulfan 40 mg/kg (n = 10); and group 6 (SSCT-C), mice subjected to a delayed SSCT with two additional doses of busulfan 10 mg/kg every 5 weeks (n = 10). Spermatogenic recovery in standard SSCT-A and SSCT-C groups ranged between 22.29 and 22.65%, compared with a lower recovery rate of 11.54% showed in the SSCT-B group. However, donor contribution resulted higher in standard SSCT-A, representing a 69.71% of cross-sections, compared with the rest of conditions ranging from 34.69 to 35.42%. Overall, we concluded that a delay in the SSCT from the gonadotoxic onset decreases the efficiency of donor-derived spermatogenic recovery in mice.

The complex multi-sectoral impacts of drought: Evidence from a mountainous basin in the Central Spanish Pyrenees.

We analyzed the impacts of drought severity on a variety of sectors in a topographically complex basin (the upper Aragón basin 2181 km2) in the Central Spanish Pyrenees. Using diverse data sources including meteorological and hydrological observations, remote sensing and tree rings, we analyze the possible hydrological implications of drought occurrence and severity on water availability in various sectors, including downstream impacts on irrigation water supply for crop production. Results suggest varying responses in forest activity, secondary growth, plant phenology, and crop yield to drought impacts. Specifically, meteorological droughts have distinct impacts downstream, mainly due to water partitioning between streamflow and irrigation channels that transport water to crop producing areas. This implies that drought severity can extend beyond the physical boundaries of the basin, with impacts on crop productivity. This complex response to drought impacts makes it difficult to develop objective basin-scale operational definitions for monitoring drought severity. Moreover, given the high spatial variability in responses to drought across sectors, it is difficult to establish reliable drought thresholds from indices that are relevant across all socio-economic sectors. The anthropogenic impacts (e.g. water regulation projects, ecosystem services, land cover and land use changes) pose further challenges to assessing the response of different systems to drought severity. This study stresses the need to consider the seasonality of drought impacts and appropriate drought time scales to adequately assess and understand their complexity.

[Assessment of a reconciliation and information programme for heart transplant patients]. / Evaluación de un programa de conciliación e información al paciente trasplantado cardíaco.

INTRODUCTION: The objective is to assess a pharmaceutical care programme for heart transplant patients upon patient admission and discharge. MATERIAL AND METHODS: Observational study of heart transplant patients, performed during the first quarter of 2007. Upon admission, the patient was interviewed regarding home treatments, adherence, allergies and adverse effects, and his/her prescriptions were compared with the last discharge report (drug reconciliation). At time of discharge, treatment was checked against the last hospital prescription (reconciliation) and an informative report was drawn up and personally delivered to the patient. Subsequently, a satisfaction questionnaire was carried out by telephone. Drug-related problems were recorded using Atefarm software. RESULTS: The programme was applied to 24 patients upon admission and 23 upon discharge. No drug interactions were detected. Treatment adherence was higher than 90%. 37.5% of patients informed of an adverse reaction. Medication-related problems were identified in 16 patients (45.7%) for 6.6% of medications, most of which (38%) were for infection prophylaxis; medication omission was the most frequently-detected error. Positive evaluation of the information that was received was higher than 90%. CONCLUSIONS: Pharmacotherapeutic follow-up upon admission and discharge resolves and prevents problems while improving patient information and satisfaction. Limitations on personnel prevent the population's requests from being met.

[Quality of drug treatment process through medication errors in a tertiary hospital]. / Calidad del proceso farmacoterapéutico a través de errores de medicación en un hospital terciario.

OBJECTIVE: To assess the quality of drug treatment process in a unit-dose and assisted electronic prescription system in a tertiary hospital, by looking at medication errors. METHODS: A prospective, observational study into medication errors was carried out on 308 hospitalised patients. This was done by assessing medical prescriptions, pharmaceutical validation, prepared and dispensed medication and by directly observing drug administration. The variable, i.e. the medication error, was analysed in the drug treatment process so as to decipher the type and cause of the error. Quality indicators were defined at each stage (percentage relationship between errors and opportunities for error). RESULTS: Of the 308 patients studied, 107 had at least 1 medication error (34.7%). There were a total of 137 errors: omission of allergy and prescription description (20.4%), prescription/validation (28.5%), dispensing (23.4%) and drug administration (27.7%). The most frequent error was dose omission (19.7%) and choice of pharmaceutical product (16.1%). The most common cause of error was forgetfulness or a lack of attention to detail (53.3%). The quality indicators by stage were: 2.3% for omission of the patient's allergies; 0.9% for prescription; 1.6% for prescription/validation; 8.2% for dispensing, and 2.1% for drug administration. CONCLUSIONS: It is estimated that 35 patients in every 100 experience errors in their drug treatment process. Opportunities for improvement are identified based on standardisation and training for professionals in carrying out technical tasks and using technology.

[Medication errors in a tertiary hospital with three different drug delivery systems]. / Errores de medicación en un hospital terciario con tres sistemas de distribución de medicamentos diferentes.

OBJECTIVE: To estimate the proportion of medication errors in a tertiary hospital, global and for each delivery medication system, to describe the error types and the implied medications, and to analyze the factors associated to the same ones. METHODS: Errors were identified from direct observation of 2,242 opportunities for error (administered doses or prescribed doses not given) by 6 couples of observers. Delivery medication systems were stock in ward, unit dose with electronic prescription and unit dose with computerized transcription. Logistic regression was used to evaluate the association between errors and certain factors. RESULTS: The medication error rate was of 7.2% (CI 95%: 6.1-8.3), and 4.4% (CI 95%: 3.6-5.3) of them reached the patient. For delivery systems, the error rate was of 9.5% (CI 95%: 7.4-11.9) for stock in ward, 7.8% (CI 95%: 5.9-10.0) for electronic prescription and 4.7% (CI 95%: 3.4-6.4) for computerized transcription. The highest error frequency was observed in the administration phase (58.4%) and the omitted dose was the most prevalent error (31.7%). The error rate was associated to the pharmacotherapeutic process, the schedule of administration and the unit of hospitalization. CONCLUSIONS: In one of each 14 opportunities for error a medication error takes place. The different delivery medication systems have different error rates.

The latent transforming growth factor-beta-binding protein-1 promotes in vitro differentiation of embryonic stem cells into endothelium.

The latent transforming growth factor-beta-binding protein-1 (LTBP-1) belongs to a family of extracellular glycoproteins that includes three additional isoforms (LTBP-2, -3, and -4) and the matrix proteins fibrillin-1 and -2. Originally described as a TGF-beta-masking protein, LTBP-1 is involved both in the sequestration of latent TGF-beta in the extracellular matrix and the regulation of its activation in the extracellular environment. Whereas the expression of LTBP-1 has been analyzed in normal and malignant cells and rodent and human tissues, little is known about LTBP-1 in embryonic development. To address this question, we used murine embryonic stem (ES) cells to analyze the appearance and role of LTBP-1 during ES cell differentiation. In vitro, ES cells aggregate to form embryoid bodies (EBs), which differentiate into multiple cell lineages. We analyzed LTBP-1 gene expression and LTBP-1 fiber appearance with respect to the emergence and distribution of cell types in differentiating EBs. LTBP-1 expression increased during the first 12 d in culture, appeared to remain constant between d 12 and 24, and declined thereafter. By immunostaining, fibrillar LTBP-1 was observed in those regions of the culture containing endothelial, smooth muscle, and epithelial cells. We found that inclusion of a polyclonal antibody to LTBP-1 during EB differentiation suppressed the expression of the endothelial specific genes ICAM-2 and von Willebrand factor and delayed the organization of differentiated endothelial cells into cord-like structures within the growing EBs. The same effect was observed when cultures were treated with either antibodies to TGF-beta or the latency associated peptide, which neutralize TGF-beta. Conversely, the organization of endothelial cells was enhanced by incubation with TGF-beta 1. These results suggest that during differentiation of ES cells LTBP-1 facilitates endothelial cell organization via a TGF-beta-dependent mechanism.

[Alerts system associated with computerized physician order entry: analysis and identification of improvement points]. / Sistema de alertas asociado a prescripción electrónica asistida: análisis e identificación de puntos de mejora.

OBJECTIVE: To make a quantitative analysis of the alerts associated with a computerized physician order entry system and identify opportunities to improve the system. METHOD: A retrospective observational study in a general hospital with 750 beds, 500 of which have a computerized physician order entry system installed. The frequency per type and medication of 525,691 alerts produced for a year in the prescription of drug treatments to 15,466 patients was analysed, entering these on a database. The system includes three categories of alert relating to the drug, the characteristics of the patient and the hospital medicine policy. By means of a failure mode and effects analysis, opportunities for improving the system were identified and corrective measures were suggested. RESULTS: It has been observed that from the total of 1,084 drugs, 20 of them produce 34% of alerts. The ten most frequently active ingredients involved are: potassium chloride, acenocumarol, imipenem, lorazepam, diazepam, mycophenolate, enoxaparin, tacrolimus, calcium carbonate and cyclosporine. The most frequent alerts generated during electronic prescription are associated with duplicated therapy (35.4%), renal failure (27.6%) and risk due to advanced age (17.2%), with these groups accounting for 80.2% of the total. The excess of alerts and information provided by the alerts were identified as priority improvement points. CONCLUSIONS: The system produced excessive alerts which led to the risk of them being ignored and reducing the capacity to prevent adverse drug events. Modifications are required for the design of the alert system, which also needs to be continuously updated.

Estrogen receptor agonists and immune system in ovariectomized mice.

Several data implicate the immune system in bone lost after estrogen deficiency, however, some of the effects on the immune system of estrogen deficiency or of estrogen receptor (ER) modulation are not well established. In this study, the effect of ER agonists on the immune system in ovariectomized mice is analyzed. Mice were ovariectomized and were administered 17beta-estradiol (E2), raloxifene (RAL) or genistein (GEN). The effect of a 4-week treatment on bone turnover and on several parameters that reflect the status of the immune system was studied. Results show that ovariectomy provoked both uterine atrophy and thymic hypertrophy. Although RAL corrected thymic hypertrophy, only E2 corrected both. Ovariectomized mice showed increased levels of serum calcium and cathepsin K gene expression and decreased levels of serum alkaline phosphatase (ALP) activity, which suggests that there is a persistent alteration in bone metabolism. Moreover, ovariectomy increased B-cells and CD25+ cells, and decreased the percentages of T-cells and Cbfa1 gene expression in bone marrow (BM). All ER agonists corrected, although to different degrees, changes induced by the ovariectomy. Furthermore, results showed that it is essential to adjust ER agonist doses to avoid immunosuppression, since all ER agonists decreased BM T-cell levels.

[Pharmaceutical care self-assessment process for hospitalized patients through the Valor program]. / Autoevaluación del proceso de atención farmacéutica en pacientes hospitalizados a través del programa Valor.

OBJECTIVE: To assess the quality of pharmaceutical care for inpatients using qualitative criteria as established in the Valor program. METHOD: Evaluation study through 43 explicit structural, process, and outcome criteria within the Valor program, in which pharmacists in the Unit Dose Functional Unit may assess themselves along a compliance scale from 0 to 100%. This Unit provides daily individualized pharmaceutical care to 550 patients in an adult general and surgery hospital. Mean scores per pharmacist and item are estimated for the 2003-2005 period. RESULTS: Mean compliance assessments for all 14 interannual "structural items" were 53, 57, and 64%; those for all 13 "process items" were 52, 51, and 46%; and those for all 15 "outcome items" were 18, 28, and 26%. A variability of 20% was documented for structure and process evaluations, and of 50% for outcome assessments. CONCLUSIONS: Every autoevaluation raises to the equipment the necessity to establish improvements and to enhance communication, and the application of standardized procedures in the pharmaceutical care process.

Renal and hepatic integrity in long-term sevoflurane sedation using the anesthetic conserving device: a comparison with intravenous propofol sedation in an animal model.

INTRODUCTION: Critically ill patients are sedated with intravenous agents because the use of inhaled agents is limited by their potential risk of toxicity. Increasing levels of inorganic fluorides after the metabolism of these agents have been considered potentially nephrotoxic. However, hepatic involvement after prolonged administration of sevoflurane has not yet been studied. The present study evaluated the potential renal and hepatic toxicity caused by prolonged administration (72h) of sevoflurane. METHODS: For this experimental, prospective, randomized, controlled trial, 22 Landrace x Large-White female pigs were randomly assigned to two groups: intravenous propofol (P) or inhaled sevoflurane via the AnaConDa™ device (S, end-tidal 2.5 vol%). The P group remained sedated for 108h with propofol. In the S group, sevoflurane was administered for 72h and then changed to propofol for the remaining 36h in order to observe the kinetics of fluoride after discontinuation of sevoflurane. Serum creatinine was the primary outcome variable, but inorganic fluoride concentrations and other renal, hepatic, and cardiorespiratory variables were also measured. RESULTS: Both groups of animals were comparable at baseline. No differences were found between the two groups for plasma creatinine and urea or creatinine clearance throughout the study. Fluoride levels were significantly higher in the sevoflurane group. No correlation was found between inorganic fluoride and serum creatinine values. No significant differences were observed for hepatic function. Hemodynamic, respiratory, and blood gas variables were comparable between the groups. CONCLUSIONS: Long-term sedation with sevoflurane using AnaConDa™ or propofol does not negatively affect renal or hepatic function.

Aflibercept in exudative age related macular degeneration refractory to ranibizumab.

PURPOSE: The aim of this study is to determine the effectiveness, safety and cost of aflibercept in the treatment of wet age-related macular degeneration (ARMD) refractory to ranibizumab. METHODS: Retrospective observational study was conducted on patients diagnosed with wet ARMD, and previously treated with ranibizumab. Efficacy variables assessed were changes in visual acuity (BCVA) and anatomical improvements in the most affected eye. Factors associated with improvement of BCVA with aflibercept were also studied. Adverse events related to the aflibercept administration were recorded. Cost analysis data were collected from the hospital perspective, and only taking the direct medical costs into account. Cost-effectiveness analysis was calculated using the aflibercept treatment cost, and effectiveness calculated as BCVA gained. RESULTS: A total of 50 eyes corresponding to 46 patients were included. The median follow-up period was 4.6 months (range: 1.0-6.0). Improvement in visual acuity after the first 2 doses and at the end of the follow-up period was observed in 32.0 and 28.0% of treated eyes, respectively. None of the variables studied was associated with an improvement in the BCVA after treatment. No significant differences were found in the average monthly cost between treatments. CONCLUSIONS: Aflibercept is shown to be an effective treatment in a significant number of patients resistant to treatment with ranibizumab, presenting a cost similar to that generated during the final stages of treatment with ranibizumab.

New practical treadmill protocol for clinical use.

A new continuous treadmill protocol (USAFSAM) has been designed using a constant treadmill speed (3.3 miles/hour) and regular equal increments in treadmill grade (5%/3min). The constant treadmill speed requires only initial adaptation in patient stride, reduces technician adjustments and produces less electrocardiographic motion artifact than do protocols using multiple or higher treadmill speeds, or both. The regular equal increments in treadmill grade are easy to implement and provide a larger number of work loads than do protocols that are discontinuous or require larger changes in work load. The USAFSAM protocol was compared with the older Balke-Ware protocol in 26 healthy men (aged 30 to 59 years). Each fasting subject completed two maximal treadmill tests from each protocol. Measurements included minute heart rate from the electrocardiogram, auscultatory blood pressures and oxygen consumption obtained with standard techniques. Similarities in between-protocol measurements for submaximal and maximal treadmill efforts were impressive; differences were small and unimportant. Further, both protocols showed equal reproducibility for the measurements noted. Importantly, time to maximal effort was reduced by 24% with the USAFSAM protocol. The USAFSAM treadmill protocol has since been used in more than 500 clinical and screening examinations, thus confirming its advantages and practicality for routine clinical stress testing. Normal reference values previously established for the Balke-Ware protocol are shown to apply to the new USAFSAM protocol as well.

Potentiation by vasopressin of adrenergic vasoconstriction in the rat isolated mesenteric artery.

1. The aim of the present study was to investigate in rat mesenteric artery rings whether low concentrations of vasopressin could modify the contractile responses to noradrenaline and electrical stimulation of perivascular nerves. 2. Vasopressin (10[10]-10[-7] M) caused concentration-dependent contractions (pD2 = 8.36+/-0.09). The V1-receptor antagonist d(CH2)5Tyr(Me)AVP (10[-9]-10[-8] M) produced parallel rightward shifts of the control curve for vasopressin. Schild analysis yielded a pA2 value of 9.83 with a slope of 1.10+/-0.14. 3. Vasopressin (3 x 10[-10] and 10[-9] M) caused concentration-dependent potentiation of the contractions elicited by electrical stimulation (2-8 Hz; 0.2 ms duration for 30 s) and produced leftward shifts of the concentration-response curve for noradrenaline. The V1-receptor antagonist induced concentration-dependent inhibitions of potentiation induced by vasopressin. The selective V1-receptor agonist [Phe2, Orn8]-vasotocin (3 x 10[10] and 10[-9] M) induced potentiation of electrical stimulation-evoked responses which was also inhibited in the presence of the V1 antagonist (10[-8] M). In contrast, the V2-receptor agonist deamino-8-D-arginine vasopressin (desmopressin 10[-8]-10[-7] M) did not modify the electrical stimulation-induced responses and the V2-receptor antagonist [d(CH2)5, D-Ile2, Ile4, Arg8]-vasopressin (10[-8]-10[-7] M) did not affect the potentiation evoked by vasopressin. 4. In artery rings contracted by 10(-6) M noradrenaline in the presence of 10(-6) M guanethidine and 10(-6) M atropine, electrical stimulation (2, 4 and 8 Hz) produced frequency-dependent relaxations which were unaffected by 10(-9) M vasopressin but abolished by 10(-6) M tetrodotoxin. 5. Vasopressin also potentiated contractions elicited by KCl and contractions induced by addition of CaCl2 to KCl depolarized vessels. The augmenting effects were inhibited by the V1 antagonist. 6. In the presence of the calcium antagonist nifedipine (10[-6] M), vasopressin failed to enhance the contractile responses to electrical stimulation, noradrenaline and KCl. 7. The results demonstrate that low concentrations of vasopressin strongly potentiate the contractions to adrenergic stimulation and KCl depolarization. This effect appears to be mediated by V1 receptor stimulation which brings about an increase in calcium entry through dihydropyridine-sensitive calcium channels.

Proteolytic control of growth factor availability.

Most growth factors are released from cells in a form that does not permit immediate interaction with their high affinity receptors. An important mechanism for presentation of these released latent growth factors is activation by the plasminogen activator-plasmin system. The involvement of this system in the biology of Transforming Growth Factor-beta (TGF-beta) is reviewed.

Endothelin-1-induced potentiation of adrenergic responses in the rabbit pulmonary artery: role of thromboxane A(2).

To examine whether low concentrations of endothelin-1 potentiate the vasoconstrictor response to adrenergic stimulation, we recorded the isometric response of rings of rabbit pulmonary artery to electrical stimulation and noradrenaline. Endothelin-1 (10(-10) M) potentiated the contractions induced by electrical stimulation and noradrenaline. The endothelin ET(B) receptor antagonist (2,6-dimethylpiperidinecarbonyl-gamma-methyl-Leu-N(in)-[Methoxycarbonyl]-D-Trp-D-Nle) (BQ-788, 10(-6) M), but not the endothelin ET(A) receptor antagonist cyclo(D-Asp-Pro-D-Val-Leu-D-TRP) (BQ-123, 10(-6) M), inhibited the potentiating effects of endothelin-1. Pretreatment with the cyclooxygenase inhibitor indomethacin, the thromboxane synthase inhibitor furegrelate and the thromboxane receptor antagonist [1S-[1alpha,2alpha(Z),3alpha,4alpha]]-7-[3-[[[[(1-oxoheptyl)amino]acetyl]amino] methyl]-7-oxabicyclo-[2.2.1]hept-2-yl]-5-heptenoic acid (SQ-30741) (all at 10(-5) M) prevented the potentiation induced by endothelin-1 on adrenergic stimulation. The Ca(2+) channel antagonist nifedipine (10(-6) M) did not affect the potentiation induced by endothelin-1. The results indicate that endothelin-1 potentiates the responses to electrical stimulation and noradrenaline by activating endothelin ET(B) receptors. This potentiation depends on the production of cyclooxygenase-generated factors, probably thromboxane A(2).