Allelic polymorphism of human FcγRIIA-H/R131 receptor in American tegumentary leishmaniasis.

FcγRIIA binding to IgG subclasses with different levels of affinity is influenced by the polymorphism in the gene that encodes this receptor. The substitution of arginine (R) for histidine (H) in the 131 position defines three allelic patterns, H/H, R/R, and H/R, resulting in FcγRIIA-H/H131 affinity for IgG2 and higher affinity for IgG3 subclasses. Studies have shown the importance of genetic host factors in leishmaniasis and participation of FcγRs on the macrophage infection by amastigote forms and in the immune response to Leishmania sp. We analysed the influence of allelic diversity patterns of the receptor FcγRIIA on American tegumentary leishmaniasis (ATL). FcγRIIA-H/R131 polymorphism was determined by PCR followed by an allele-specific enzymatic digestion in 88 individuals with ATL and 98 healthy volunteer blood donors (control group). The genotypic and allelic distributions of FcγRIIA-H/R131 were similar among the studied groups as well in mild and severe clinical forms of ATL. Our results suggest no association between this allelic polymorphism and susceptibility or resistance to ATL, neither influencing the development of different clinical forms of this illness.

Whole blood glutathione peroxidase and erythrocyte superoxide dismutase activities, serum trace elements (Se, Cu, Zn) and cardiovascular risk factors in subjects from the city of Ponta Delgada, Island of San Miguel, The Azores Archipelago, Portugal.

Activities of whole blood glutathione peroxidase (GSH-Px) and erythrocyte superoxide dismutase (SOD) and serum levels of selenium (Se), copper (Cu) and zinc (Zn) were measured in 118 apparently healthy subjects aged 20-60 years from the city of Ponta Delgada, Island of San Miguel, The Azores Archipelago, Portugal. Data were analysed by age/gender, lipid profile and blood pressure as cardiovascular risk factors searching for their relevance when assessing reference values for antioxidant biomarkers. GSH-Px was in the same range, but SOD was significantly lower than in other Portuguese populations. Neither activity differed with gender. GSH-Px activity increased with age, namely in normolipidemic men versus the hyperlipidemic group in which a decrease was observed. This suggests a progressive impairment of GSH-Px with age caused by an enhanced production of oxidant species in hyperlipidemia. GSH-Px was 30% lower in male hypertensives versus normotensives. SOD activity did not relate to age or blood pressure but was 17% higher in the hyperlipidemic men versus the normolipidemic group, suggesting a better antioxidant protection by SOD than by GSH-Px in hyperlipidemia and hypertension. Se was higher in men versus women, particularly in the older subjects, and partly related to hyperlipidemia. Zn levels showed a similar dependency on gender, not related to age or lipid profile. Cu levels were much higher in women than in men in all age or lipid profile classes and decreased in hyperlipidemia. They were lowered with age in both genders, particularly in normolipidemic women. The present research therefore suggests that hyperlipidemia and hypertension do affect antioxidant status and should be considered when assessing antioxidant biomarkers in blood.

Diagnóstico por imagen en la enfermedad inflamatoria intestinal / Diagnostic imaging of inflammatory bowel disease

En el presente artículo se consideran las posibilidades de las distintas técnicas de formación de imagen en el estudio de la enfermedad inflamatoria intestinal, con especial referencia al papel desempeñado por la tomodensitometría y la resonancia magnética. Aunque éstas no superan al tránsito baritado o a la enteroclisis convencional en la demostración del detalle mucoso, se muestran muy útiles en la evaluación de las características de la pared de las asas y en la detección de las posibles complicaciones extraluminales: abscesos, adenopatías, afectación mesentérica, etc. La resonancia magnética se presenta con un gran potencial en el estudio del intestino delgado, probablemente en un futuro no lejano desempeñará un importante papel en la evaluación de la enfermedad inflamatoria intestinal, entidad de carácter recurrente, que acaece en pacientes jóvenes, con amplia esperanza de vida, en los que hay que minimizar la exposición a las radiaciones ionizantes. Se presenta un abordaje fisiopatológico de la secuencia de acontecimientos que justifican los hallazgos radiológicos (úlceras aftoides, ulceraciones fisurantes y penetrantes, ulteriores trayectos fistulosos, abscesos, afectación mesentérica, etc.) y se considera una clasificación basada en los hallazgos radiológicos, objetiva, que resulta útil para la toma de decisiones desde el punto de vista terapéutico, justificando así el importante papel que cumple el radiólogo no sólo en el diagnóstico, sino también en el control evolutivo de esta entidad

This article considers the possibilities of different image acquisition techniques in the study of inflammatory bowel disease, with special emphasis on the roles of tomodensitometry and magnetic resonance imaging. Although these techniques are not superior to barium follow through studies or conventional enteroclysis at depicting the mucosa in detail, they are very useful in the evaluation of the characteristics of the walls of the bowel loops and in the detection of possible extraluminal complications, such as abscesses, adenopathies, and mesenteric involvement. Magnetic resonance shows great potential for the study of the small bowel and is likely to play an important role in the study of inflammatory bowel disease in the near future. This recurring disease affects young patients with long life expectancies in whom exposure to ionizing radiation needs to be minimized. We explain the radiologic findings through a physiopathologic approach to the sequence of events (aphthoid ulcers, fissuring and penetrating ulcerations, formation of fistulous tracts, abscesses, mesenteric involvement) that give rise to them. We propose an objective classification based on the imaging findings that is useful for therapeutic decision making, and we describe the important role of the radiologist in the follow-up as well as in the diagnosis of this entity

Valoración clínico-radiológica de las vertebroplastias / Clinical and radiologic assessment of vertebroplasty

Introducción. La vertebroplastia percutánea consiste en la inyección de cemento acrílico en los cuerpos vertebrales debilitados, con el fin de obtener un efecto analgésico y la estabilización mecánica de la columna vertebral. Objetivo. Valorar las características y efectividad de las vertebroplastias realizadas en el Hospital Universitario Miguel Servet de Zaragoza. Pacientes y métodos. Se han estudiado, de forma retrospectiva, 147 vertebroplastias efectuadas entre 1994 y 2003, a 95 pacientes (60 mujeres y 35 hombres), con un rango de edad entre 19 y 84 años. La vía de abordaje utilizada actualmente es la transpedicular oblicua, que con una sola punción consigue una buena expansión del cemento dentro del cuerpo vertebral. Resultados. La lesión ósea más frecuentemente tratada con vertebroplastia es la fractura secundaria a osteoporosis, con un 65% de los casos, le sigue con un 23% los hemangiomas y, en menor proporción, las metástasis osteolíticas, fracturas traumáticas, linfomas y mielomas. Para valorar la reducción del dolor empleamos la escala analógica visual (VAS), pasando de un VAS previo a la vertebroplastia de 8,88 a un VAS de 2,78 después del tratamiento. Solamente el 7,4% de los pacientes tuvieron complicaciones sintomáticas. Conclusión. La vertebroplastia es un procedimiento seguro y eficaz, de elección en el tratamiento del dolor vertebral refractario a la medicación, que permite una rápida incorporación del paciente a su vida habitual y una reducción de los costes hospitalarios

Introduction. Percutaneous vertebroplasty consists of the injection of acrylic cement into weakened vertebral bodies to achieve pain relief and mechanical stability of the spine. Objective. To evaluate the characteristics and effectiveness of the vertebroplasties performed at the Hospital Universitario Miguel Servet in Zaragoza. Patients and methods. This is a retrospective study of 147 vertebroplasties performed in 95 patients (60 women and 35 men; age range: 19 to 84 years). The oblique transpedicular approach, which achieves adequate cement injection with a single puncture, is currently used. A visual analogue scale (VAS) was used to evaluate pain before and after the procedure. Results. The osseous lesion most often treated by vertebroplasty is fracture secondary to osteoporosis, accounting for 65% of the cases in this series, followed by hemangiomas (23%), and osteolytic metastases, traumatic fractures, lymphomas, and myelomas. Prior to vertebroplasty, the mean VAS score was 8.88 versus 2.78 after the treatment. Only 7.3% of the patients had symptomatic complications. Conclusion. Vertebroplasty is safe and efficacious; it is the treatment of choice for vertebral pain refractory to medication. It enables patients to return to their habitual lifestyle quickly and thus helps reduce hospital stays and costs

Expectativas hacia la jubilación del personal de la Universidad de Santiago de Compostela / Retirement expectations of the staff of the University of Santiago de Compostela

Involucrados en la importancia que el fenómeno jubilación tiene en la actualidad, hemos llevado a cabo un estudio sobre las expectativas hacia la misma de un grupo de trabajadores de la Universidad de Santiago de Compostela (USC), cuestionando parámetros como edad de jubilación, sostenibilidad del Sistema de Pensiones, y motivados especialmente en la repercusión subjetiva sobre los individuos, a través de la administración de un cuestionario que mide actividad, vinculación social, expectativas hacia la jubilación y satisfacción laboral y vital, junto con variables de identificación personal (edad, sexo, estado civil, etc). A través del programa estadístico SPSS realizamos un análisis descriptivo entre las variables de estudio, lo que nos ha llevado a concluir que las actitudes hacia la jubilación son de esperanza e ilusión, con un interés manifiesto por seguir en la vida activa, donde el mayor temor suscitado es ver limitado el poder adquisitivo

Since we are involved in the importance of the retirement nowadays, questioning parameters as age of retirement, sustainability of pension systems, and we pay special attention on the subjective impact on individuals, we have carried out an study about the expectations on retirement, among a group of workers in the University of Santiago de Compostela (USC). In order to do so, we have made a survey, measuring activity, social link, expectations on retirement and labour and vital satisfaction, as well as personal identification variables (age, sex, marital status, etc). By using the SPSS software, we made a descritpive analysis among the variables of study. This lead us to conclude that the attitude about retirement is hope. We also found a stated interest to continue the active life. The biggest fear is to see a restriction of the purchasing power

An assessment of time-dependent effects of lead exposure in algerian mice (Mus spretus) using different methodological approaches.

Time-dependent effects of lead (Pb) toxicity were studied in Algerian mice (Mus spretus) treated with Pb acetate via drinking water (1 g Pb acetate/L) for different periods of exposure (15, 45, and 90 d). End points included the determination of hepatic Pb concentration and the assessment of some morphophysiological, biochemical and cytogenetical parameters. A control group receiving distilled water was also monitored for comparative purposes. Hepatic Pb accumulation increased with the time of exposure and was significantly higher in treated mice when compared to controls. In association with significant body mass loss in Pb-exposed mice, for 15 and 45 d, a significant increase in the relative spleen mass was observed after 45 d of intoxication. Pb-exposed mice also showed significant decreases in red blood cells, hematocrit, and mean corpuscular hemoglobin. On the contrary, changes in plasma transferases (aspartate aminotransferase and alanine aminotransferase) and hepatic superoxide dismutase activities did not reach statistical significance. A significant increase in the frequency of micronucleated polychromatic bone marrow erythrocytes was also found in the 90-d-exposed mice, compared to nontreated mice and the other exposed groups. Exposure to Pb acetate resulted also in a slight time-dependent decrease of the polychromatic-normochromatic ratio. These results support the concept that a long-term chronic exposure to Pb induced alterations upon some morphophysiological and genetic parameters in Algerian mice.

Trace element status (Se, Cu, Zn) in healthy Portuguese subjects of Lisbon population: a reference study.

Serum levels of selenium, copper, and zinc were systematically determined in healthy subjects of the Lisbon population. The sample consisted of 183 blood donors of both genders who were divided into two age groups: 20-44 and 45-70 yr of age; relationships with gender, age, the lipid profile, and tobacco consumption were investigated. In the female group, the intake of oral contraceptives and pregnancy were considered for the youngest women, and hormonal replacement therapy (HRT) was taken into account for the oldest ones. Serum concentrations of these elements were in the same range as those found for populations of other European countries. Differences between genders were observed for the three elements studied, with serum selenium and zinc concentrations higher in men and copper levels higher in women. Age-dependent differences were found for selenium: The oldest subjects (regardless gender) presented the highest concentrations of selenium in serum as further demonstrated by the positive correlation with age. In both pregnant and contraceptive-using women, copper was greatly increased, confirming the influence of estrogen status and/or oral contraceptive intake on increased serum copper levels. However, in postmenopausal women, HRT did not significantly affect serum copper levels. Selenium, copper, and zinc status were not different between normolipidemic and hyperlipidemic subjects for the same gender and age range, but selenium levels tended to increase with hyperlipidemia when considering the whole group of subjects. With respect to the lifestyle, higher serum zinc levels were found in tobacco-consuming men. Albumin serum levels were similar for all considered subgroups, except for the pregnant women, for whom a decrease in this parameter was observed. The present study allowed one to obtain reference values for this healthy group of population, which will serve for a comparative study with groups having pathological conditions, such as cardiovascular disease.

Systemic markers of the redox balance in chronic obstructive pulmonary disease.

Chronic obstructive pulmonary disease (COPD) is highly prevalent and its pathogenesis is still not completely clarified. Clinically stable patients (n=21) and healthy subjects (n=24) were studied for blood markers of oxidative injury and antioxidant status. The plasma concentration of protein carbonyls was significantly increased in COPD patients, both ex-smokers (0.76 +/- 0.28 nmol mg(-1)) and smokers (0.99 +/- 020 nmol mg(-1)) versus controls (0.49 +/- 0.14 nmol mg(-1)) . The concentration of total thiols was slightly enhanced in plasma of the COPD patients (ex-smokers 492 +/- 23 micromol 1(-1) and smokers 505 +/- 36 micromol 1(-1) versus controls 450 +/- 67 micromol 1(-1); p < 0.05). The activity of the antioxidant enzyme superoxide dismutase was increased in erythrocytes (activity in U g(-1) haemoglobin; ex-smokers 4460 +/- 763 and smokers 4114+/- 1060 versus 3015 +/- 851 in controls; p > 0.01), while glutathione peroxidase activity was decreased in total blood (activity in U g(-1) haemoglobin: ex-smokers 27 +/- 9 and smokers 23 +/- 9 versus 47 +/- 25; p < 0.01). Lower levels of selenium in plasma were also found for COPD patients (concentration in mg 1(-1): ex-smokers 0.030 +/- 0.019 and smokers 0.032 +/- 0.024 versus 0.058 +/- 0.023 in controls; p < 0.01), being more evident in those with very low levels of arterial oxygen pressure. In addition, the levels of potassium and rubidium were increased in blood cells of the patient group. All these changes might reflect oxidant damage and an altered electrolytic homeostasis, and can be interpreted as markers of COPD rather than as indicators of smoking habits.

Hepatic elemental contents and antioxidant enzyme activities in Algerian mice (Mus spretus) inhabiting a mine area in central Portugal.

In this study the effects of heavy metals (manganese, iron, copper, zinc) and selenium exposure on the hepatic activity of antioxidant enzymes, superoxide dismutase (SOD) and glutathione S-transferases (GST), were appraised on a seasonal basis in Algerian mice (Mus spretus) inhabiting an active copper mine area. A reference population of the same species was considered for comparative purposes. Different patterns of seasonal variation were found in both populations for the manganese, iron and selenium hepatic concentrations and SOD activity. When the two populations were compared, iron and selenium concentrations were enhanced in mice from the polluted area. In addition, SOD activity was significantly decreased in summer in exposed mice, but no other significant changes in SOD and GST activities between sites throughout the year were recorded. However, when seasonal data within each group of mice were pooled, significant differences were found between sites for the average concentrations of manganese, iron and selenium, which are higher in the polluted site. In addition, significant differences were obtained for the average values of SOD and of GST activities, due to simultaneously higher GST values and slightly lower SOD values in the polluted site. The population from the reference site was more homogeneous for all parameters measured than the population from the polluted area. These results, in particular the higher variability in data collected from mice exposed to heavy metals and selenium, combined with the negative associations between biochemical markers and heavy metals, may suggest, despite the good adaptability of the mice to their habitat, biochemical stress due to diminished environmental quality.

[Detection of phospholipidolytic Candida albicans isolated from saliva of children with Down's syndrome]. / Detecção de Candida albicans fosfolipidolíticas isoladas da saliva de crianças com síndrome de Down.

The childhood is one of the most propitious period of the life to the occurrence of infection by yeasts of the genus Candida. In children with Down's syndrome, besides the predispose factors to bucal candidiasis; macroglossia, bucal muscular incompetence, frequent respiratory diseases, motor difficulty and immunologic deficit are mentioned as additional elements for this fungus disease. It was verified that the children attacked by this syndrome have much more strains of Candida than other children. The aim of this study was to detect the prevalence of phospholipase producer, Candida on the saliva of children with Down's syndrome. Candida albicans was the only identified specie of Candida. The phospholipase production was found in isolated strains from both of study and control. However, the isolated strains of the group of children with Down's syndrome have strongly present phospholipidolitic.

Morphological and haematological parameters in the Algerian mouse (Mus spretus) inhabiting an area contaminated with heavy metals.

In the present study morphological (ratio of body weight to body length, weight of internal organs) and haematological parameters (cells count, haemoglobin concentration, haematocrit, mean corpuscular volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration) were used for monitoring potential effects of metal contaminants on the Algerian mouse (Mus spretus). The study also provided the opportunity to evaluate the use of morphological and haematological characteristics as indicators of a metal stress. Data were taken from a riverside population inhabiting a heavy metal-contaminated site and compared with those collected from a reference population of the same species. With the exception of non-reproductive males, all mice weighed less in the metal-polluted site as compared to the reference site. Spleen and kidney masses were also less in contaminated mice but mean haemoglobin concentration was greater in the polluted group versus controls. Responses of mice from the polluted area may indicate physiological stress due to diminished environmental quality. Results also suggest that M. spretus may be used as an indicator in pollution monitoring.

Response of antioxidant enzymes in freshwater fish populations (Leuciscus alburnoides complex) to inorganic pollutants exposure.

Evidence is accumulating indicating the importance of antioxidant enzyme activity measurements in eco-toxicological studies, as they may constitute markers for exposure to a large variety of pollutants. Variation of antioxidant enzymes, superoxide dismutase (SOD) and glutathione S-transferases (GST) and the effect of heavy metals and selenium exposure on these enzymes were investigated in the livers of Iberian endemic minnows (Leuciscus alburnoides complex) captured in a copper (Cu) mining area. Higher hepatic levels of copper and selenium were always observed in fish captured at the polluted site relative to the reference area population, reflecting the environmental monitoring results. A seasonal fluctuation in zinc and selenium levels for both populations was also observed which could be related to gonad maturation. The activity of SOD did not show significant regional alterations, but a seasonal variation occurred presumably associated with the Leuciscus life cycle. The GST activity was higher in the fish population from the polluted area (except in spring) and GST variability was associated with selenium and copper levels when both regions were compared. The increased GST activity was probably a metabolic adaptation to the continuous exposure to higher levels of those elements.

Trace element status (Se, Cu, Zn) and serum lipid profile in Portuguese subjects of San Miguel Island from Azores'archipelago.

Data on selenium, copper and zinc in serum of Portuguese inhabitants of the city of Ponta Delgada-Azores Archipelago are reported for the first time. The subjects are of both sexes, aged 20 to 60 years, and non-abusers of alcohol, tobacco or drugs. Serum concentrations of these elements are in the same range than those found for populations of Lisbon and of some other European countries. Differences between sexes are observed, with serum selenium and zinc levels being higher in males than in females, and the copper levels being higher in females as compared to males. These results can be explained by the hormonal status and/or oral contraceptive steroids intake, particularly for copper. Concerning lipid profile, the majority of individuals have serum lipid parameters within the normal range. In addition, no difference in trace element levels between normo and hyperlipidemic individuals is observed and no conclusive results about the relationship of all evaluated parameters to alcohol, tobacco and drug consumption are observed, in agreement with data obtained in the population of Lisbon.

The effect of age and selenium on some biochemical parameters in rat liver.

Two age groups, 3 and 15 mo, were used to investigate whether age-associated changes in some parameters related to lipid peroxidation occur in the liver of male Wistar rats and to observe possible effects of dietary selenium supplementation (0.25 and 0.50 ppm) for 12 mo on the same parameters. At these experimental conditions, the most important observation was that peroxidation did not change by aging, at least until 15 mo of age. In addition, the activity of Se-dependent glutathione peroxidase (GSH-Px, EC 1.11.1.9) was higher in the liver of the older animals. It is suggested that the enzyme could have a role in the unchanged hepatic peroxidation observed in aged male rats. On the other hand, an effect of dietary selenium supplementation on those parameters was not observed, probably because the selenium levels were still at an adequate plateau.

Effect of selenium supplementation on polyunsaturated fatty acids in rats.

The purpose of this work was to study plasma, adipose tissue, and liver fatty acids percentages of Wistar rats that drank water supplemented with several levels of sodium selenite for 1, 3, and 6 mo. In a general way, percentages of saturated, monounsaturated, and polyunsaturated fatty acids of supplemented groups were not different from those obtained with nontreated animals in the analyzed tissues. However, in rats supplemented with 0.5 ppm Se, mainly in adipose tissue, a polyunsaturated fatty acids increase (p < 0.005) was observed for all times of treatment. This could suggest that 0.5 ppm Se supplement probably exercises a protective role on polyunsaturated fatty acids in that tissue. Supplements of 6.0, 15.0, and 54.0 ppm Se did not change unsaturation levels of fatty acids in the analyzed tissues.

Effect of selenium supplementation on some blood biochemical parameters in male rats.

The long-term effect of selenium supplementation on blood glutathione peroxidase (GSH-Px) activity and plasma TBARS' production (as an index of peroxidation) was evaluated in 15-mo-old male rats fed a diet supplemented with 0.25 or 0.50 ppm selenium, for 12 mo. A group of nonsupplemented age-matched rats was the control. In addition, triglycerides, phospholipids, total and free cholesterol, HDL-cholesterol, and HDL-phospholipids levels were measured in plasma. Plasma testosterone levels were also determined in order to control the aging process in these animals. The GSH-Px activity and the peroxidation level were unchanged in all the groups. However, concerning the lipid parameters, a decrease in triglycerides concentration was observed in both treated groups (p < 0.05). Therefore, in these experimental conditions, despite no observed changes in parameters related to lipid peroxidation, selenium seems to be involved with triglycerides metabolism, eventually improving the triglycerides status of aged animals.

Selenium and lipid parameters in plasma of Portuguese subjects.

To obtain further insight into the role of selenium in the development of atherosclerosis, plasma selenium and lipid parameters were determined in 126 Portuguese subjects living in the region of Lisbon, 60 women and 66 men, 20-60 years old, divided up in to three groups according to their plasma lipid profile: Group I consisted of normolipidemic subjects having plasma triglycerides and total cholesterol < 200mg/dL; Group II was composed of hypercholesteremic subjects with total cholesterol > 200mg/dL and plasma triglycerides < 200mg/dL; Group III was composed of hypercholesteremic and hypertriglyceremic subjects having total cholesterol and plasma triglycerides > 200mg/dL. Respective values for mean plasma selenium (+/- SD) in the groups were: 93 +/- 18 micrograms/L, 90 +/- 17 micrograms/L and 96 +/- 18 micrograms/L. A weak significant difference in plasma selenium between women and men was, however, observed in Group I (87 +/- 20 micrograms/L vs. 100 +/- 11 micrograms/L, p < 0.05). Regression analysis showed no significant relationship between plasma selenium and lipid parameters, except in the most hyperlipidemic women of Group III, where a weak correlation between plasma selenium and the HDL-cholesterol/total cholesterol ratio (r = 0.80, p < 0.05) was observed. The present study suggests that the relationship between selenium and HDL-cholesterol has to be further examined, taking into account nutritional, metabolic and genetic factors.

Plasma and liver selenium levels in the rat during supplementation with 0.5, 2, 6, and 15 ppm selenium in drinking water.

Plasma and liver selenium of Wistar rats were determined after 1, 3, and 6 mo supplementation with 0.5, 2, 6, or 15 ppm selenium as sodium selenite in drinking water. Plasma selenium was not different from control values at additional intake of 0.5 ppm but increased above usual levels at higher intakes. A highly significant correlation was observed between the total quantity of selenium ingested and plasma selenium after 1 mo treatment (r = 0.99, p < 0.01), but was less pronounced after 3 and 6 mo (0.94, p < 0.05, and 0.78, p < 0.05, respectively). The decrease in plasma selenium with time of treatment was more pronounced at higher intakes. There was also a highly significant correlation between total selenium intake and liver selenium concentration (r = 0.99, p < 0.01) after 1 mo of treatment, but this time liver selenium did not change with time, and the correlation remained highly significant throughout the investigation. Liver selenium therefore appears as a more sensitive and more representative measure of selenium intake than plasma selenium. Most supplements did not affect body weight and survival of animals, except when the diet was supplemented with 15 ppm for 6 mo; however, alterations in biochemical parameters concerning lipid status and hepatic function were observed at levels above 2.0 ppm.