Plasma L-arginine and metabolites of nitric oxide synthase in patients with left-to-right shunt after intracardiac repair.

STUDY OBJECTIVE: Human plasma L-arginine serves as a substrate pool for endothelial-derived nitric oxide (NO) synthase. In this pilot study, we tested the hypothesis that plasma L-arginine and other metabolites of the L-arginine NO pathway could correlate with postoperative pulmonary hypertension after cardiopulmonary bypass (CPB). DESIGN: Forty-two patients (median age, 0.5 years; range, 0.1 to 28 years) with atrial septal defect (n = 15), ventricular septal defect (n = 18), atrioventricular canal (n = 8), and aortopulmonary window (n = 1) were enrolled. The influence of patient age, preoperative pulmonary hypertension, duration of CPB, plasma L-arginine, guanosine 3', 5'-cyclic monophosphate (cGMP), and nitrate on postoperative pulmonary hypertension during the first 24 h after CPB was studied by logistic regression. RESULTS: Nineteen of 42 patients were found to have preoperative pulmonary hypertension. Thirteen of 42 patients showed persistent pulmonary hypertension after intracardiac repair with a mean pulmonary artery pressure (PAP) of 38 mm Hg (range, 23 to 55 mm Hg) at 24 h after CPB. L-arginine concentrations in plasma were significantly lower 24 h after CPB than before: 52 mumol/L (range, 18 to 95 mumol/L) vs 79 mumol/L (range, 31 to 157 mumol/L). Plasma cGMP levels were higher and plasma nitrate levels were lower immediately after weaning from CPB (p < 0.0033). On logistic regression analysis, only patient age (p = 0.02) and preoperative PAP (p = 0.01) were related to postoperative pulmonary hypertension. CONCLUSION: Low plasma L-arginine does not relate to persistent pulmonary hypertension in patients with left-to-right shunt after CPB and intracardiac repair.

Serum hyaluronic acid levels are elevated in arthritis patients, but normal and not associated with clinical data in patients with fibromyalgia syndrome.

BACKGROUND: Fibromyalgia syndrome (FM) is a disease with widespread chronic pain and many nonspecific symptoms. Hyaluronic acid (HA) is a disputed marker for the diagnosis of FM. The aim of the study is to clarify the discrepant results reported so far. METHODS: Serum concentrations of HA were measured with a radiometric assay (Pharmacia & Upjohn, Sweden) in 41 patients with FM (38 females), 48 with arthritis (35 females) and 31 control subjects (28 females). Correlations of HA levels with clinical parameters (duration of disease, age, gender, medication) and scores of disease severity (e.g. depression and pain) were calculated. If appropriate, partial correlations and analysis of covariance adjusted for confounding variables (e.g. age) were used. RESULTS: HA levels were confirmed to be age-related in the whole study group (r(s) = 0.54; P < 0.001) and each subgroup. Association between HA levels and gender, drug therapy, clinical or psychometric data could not be demonstrated in patients suffering from FM. Analyzing all study participants, HA levels were correlated with the pain disability index (PDI) (r(tau) = 0.27; P < 0.02) and, in arthritis patients only, with duration of disease (r(tau) = 0.82; P < 0.001). Moreover, analysis of covariance revealed that patients with FM had normal HA values as compared with control subjects and only patients with arthritis had significantly higher levels than both other groups. CONCLUSIONS: The present study with a quite large cohort including patients with arthritis and FM demonstrates that serum levels of HA in FM are neither elevated nor associated with any relevant clinical data of this disease and, therefore, have no diagnostic or prognostic value in Germans.

Activity and concentration of lipoprotein lipase in post-heparin plasma and the extent of coronary artery disease.

Numerous studies have found polymorphisms in the lipoprotein lipase (LPL) gene to be associated with the risk of coronary artery disease (CAD), implicating LPL in the development of atherothrombotic disease. It remains controversial, however, whether LPL acts in a pro- or anti-atherogenic fashion. We quantitated activity and concentration of LPL in post-heparin plasma from 194 male patients undergoing coronary angiography. HDL cholesterol was significantly associated with LPL activity quartiles (1.09+/-0.26 the highest vs. 0.96+/-0.25 mmol/l the lowest quartile, P<0.01). There was also a trend towards higher total (5.61+/-1.33 vs. 5.16+/-1.44 mmol/l, P=0.059) and LDL cholesterol (3.92+/-1.39 vs. 3.46+/-1.06 mmol/l, P=0.09) with higher LPL activity. In contrast, measures of CAD extent showed no differences between LPL quartiles (P>0.30 for prior myocardial infarction, number of diseased vessels, Gensini and extent scores). Additionally, there was no difference in LPL activity (CAD: n=158, 168+/-70 nmol/ml/min, no CAD: n=36, 180+/-89 nmol/ml/min, P=0.47) or concentration (280+/-121 ng/ml and 288+/-111 ng/ml, P=0.72) between patients with and without CAD. Our data show that, in spite of an association with lipoprotein parameters, LPL in post-heparin plasma is unrelated to the presence or the extent of CAD. Therefore, lipoprotein lipase determination in plasma does not appear to be a useful marker in the assessment of CAD risk.

Pharmacokinetic profile of a locally administered doxycycline gel in crevicular fluid, blood, and saliva.

BACKGROUND: With the help of so-called controlled release delivery systems, the half-life period of locally administered antibiotics in gingival crevicular fluid (GCF) can be extended significantly. The aim of this study was to characterize the delivery profile of a new one-component 14% doxycycline free amine gel for local application. Pharmacokinetics of doxycycline (DOXY) were analyzed in GCF, saliva, and serum. METHODS: Twenty patients with persisting or recurring pockets (probing depths > or = 5 mm and bleeding on probing) after mechanical treatment (surgical or non-surgical) took part in the study. In each patient 1 periodontal defect was treated with DOXY gel. Samples of GCF, saliva, and serum were obtained before application of DOXY gel; 15 minutes after application; at 2 and 5 hours; and at 1, 2, 3, 4, 7, 9, and 11 days after application. Separation and quantitative measurement of DOXY was performed with high performance liquid chromatography and UV detection at lambda = 260 nm. RESULTS: Coefficients of variation were lower than 2% (intraassay) and 4% (interassay), respectively. For concentrations between 50 to 1000 microg/ml, we found a linear relationship between expected and measured DOXY values (linear coefficient of correlation: r = 0.998). Within the first 5 hours after application, concentration of DOXY in GCF (maximum after 15 minutes 19.97 +/- 5.85 mg/ml) and saliva (maximum after 15 minutes 17.83 +/- 2.84 mg/ml) was similar. Then concentration fell to a lower level (28.90 +/- 19.44 microg/ml) compared to GCF (577.41 +/- 127.34 microg/ml) after 3 days. Up to 10 days after application, the concentration of DOXY in GCF was 34.24 microg/ml. With the exception of 1 patient, all serum samples were DOXY-negative. CONCLUSIONS: 1) After subgingival application of biodegradable 14% doxycycline gel, mean doxycycline levels in GCF that exceeded 16 microg/ml could be maintained for at least 12 days. Thus, the antimicrobial agent may be classified as a controlled release device. 2) The antibiotic effect was limited mainly to the subgingival sites of application of the doxycycline gel. 3) The doxycycline gel possesses the pharmacokinetic and clinical properties to deliver efficacious levels of antibiotics to the periodontal pocket and to maintain these levels for at least 1 week without the need of further drug retention by a periodontal dressing.

The CFTR associated protein CAP70 interacts with the apical Cl-/HCO3- exchanger DRA in rabbit small intestinal mucosa.

DRA (down regulated in adenoma) is an intestinal anion exchanger, acting in parallel with NHE3 to facilitate ileal and colonic NaCl absorption. Furthermore it is involved in small intestinal bicarbonate secretion. Because DRA has a PDZ interaction motif, which may influence its properties, we searched for DRA-interacting PDZ adapter proteins in the small intestine. Using an overlay assay with the recombinant DRA C-terminus as a ligand, a 70 kDa protein was labeled, which was restricted to the brush border membrane in rabbit duodenal and ileal mucosa and was not detected in the colon. Destruction of the C-terminal PDZ interaction motif abolished this band, suggesting a specific protein-protein interaction. The 70 kDa protein was identified as CAP70 (CFTR associated protein of 70 kDa) by an anti-CAP70 antibody and by two in vitro binding assays after cloning CAP70 from rabbit duodenum and ileum. The interaction was recapitulated in HEK cells transfected with DRA and PDZK1, the human orthologue of CAP70. Corresponding to the overlay assay, no CAP70 mRNA or protein was detected in the colon. In vitro protein-protein interaction studies revealed specific binding of DRA to the 2nd and 3rd PDZ domain, while CFTR is known to interact with PDZ1, PDZ3, and PDZ4. The composition of macromolecular complexes assembled by CAP70 in the distal small bowel is unknown. Its restricted expression shows that it cannot be involved in NaCl absorption in the proximal colon. We suggest that CAP70 mediates regulatory functions specific to the small intestine.

Comparison of the pharmacokinetic profiles of two locally administered doxycycline gels in crevicular fluid and saliva.

BACKGROUND: Controlled-release delivery systems enable the clinician to extend the half-life period of locally administered antibiotics in gingival crevicular fluid (GCF) significantly. The aim of this split-mouth study was to compare the pharmacokinetic profile of two different doxycycline gels (DOXY and ATRI) for topical subgingival application. Pharmacokinetics of both doxycycline gels were analyzed in GCF and saliva. METHODS: In 10 patients suffering from severe periodontitis, 10 pairs of contralateral defects (pocket depth > or =5 mm/bleeding on probing or > or =6 mm) were randomly assigned either to the first application of DOXY or ATRI. Fourteen days after the topical application of the first antibiotic gel, the application of the second gel in the contralateral defect took place. Clinical examinations at baseline showed no significant differences between sites treated with DOXY and ATRI. Samples of GCF and saliva were drawn baseline, 2, 5 and 24 h after application, 2, 3, 4, 7, 9 and 11 days after application. Separation and quantitative measurement of both doxycycline-gels was performed with HPLC- and UV-detection at lambda=260 nm. RESULTS: In saliva specimens, time-dependent changes of mean doxycycline concentration were almost identical for both doxycycline-gels and declined from a maximum 2 h after application (ATRI: 6653.90+/-3096.14 microg/ml; DOXY: 5386.60+/-1542.02 microg/ml [arithmetic mean+/-SEM]) to zero values 9 days after application. In crevicular fluid specimens, sites treated with ATRI exhibited a faster decrease of mean doxycycline concentration (1085.30, 264.00, 273.94, and 258.00 microg/ml measured 2, 5, 24, and 48 h after application) than sites treated with DOXY (1388.38, 1300.40, 803.73, and 235.10 microg/ml). The faster decrease of ATRI compared with DOXY could not be proved to be statistically significantly different. CONCLUSIONS: Both doxycycline gels showed pharmacokinetics of controlled-release delivery systems.

Anti-p53 in breast cancer: concordance of different assay procedures and association with p53 antigen expression.

OBJECTIVE: Anti-p53 levels detected by different methods were compared in a predefined group of patients with breast cancer and correlated with p53 antigen expression in the corresponding tumors. METHODS: P53 autoantibodies were investigated in 165 patients with primary breast cancer using ELISAs with recombinant or native p53. Immunoblot and indirect immunofluorescence (Huh7) were used for confirmation, p53 antigen expression in the tumor was determined immunohistochemically. RESULTS: Using ELISA, overall 18/165 positives (11%) were detected, with only partly concordant results between the assays. Five positive sera were confirmed by immunoblot, and three also by indirect immunofluorescence. Anti-p53-positive patients detected by more than two assays showed accumulated p53 in the tumor (6/6) and mostly suffered from recurrent tumors (4/6; p = 0.02). In these cases, a trend towards a shortened disease-free interval was found (26 vs. 49 months; n.s.). In patients with a positive or borderline result in only one of the serological methods, there was no increased rate of p53 accumulation compared to anti-p53-negative patients (4/19 versus 35/126). CONCLUSIONS: Lack of assay standardization may partly explain the divergence in reports on anti-p53 and its clinicopathological associations. We speculate that, in different groups of patients, anti-p53 might be induced by different mechanisms.

Evaluation of Cobas Integra 800 under simulated routine conditions in six laboratories.

The new selective access analyser Cobas Integra 800 from Roche Diagnostics was evaluated in an international multicentre study at six sites. Routine simulation experiments showed good performance and full functionality of the instrument and provocation of anomalous situations generated no problems. The new features on Cobas Integra 800, namely clot detection and dispensing control, worked according to specifications. The imprecision of Cobas Integra 800 fulfilled the proposed quality specifications regarding imprecision of analytical systems for clinical chemistry with few exceptions. Claims for linearity, drift, and carry-over were all within the defined specifications, except urea linearity. Interference exists in some cases, as could be expected due to the chemistries applied. Accuracy met the proposed quality specifications, except in some special cases. Method comparisons with Cobas Integra 700 showed good agreement; comparisons with other analysis systems yielded in several cases explicable deviations. Practicability of Cobas Integra 800 met or exceeded the requirements for more than 95% of all attributes rated. The strong points of the new analysis system were reagent handling, long stability of calibration curves, high number of tests on board, compatibility of the sample carrier to other Roche systems, and the sample integrity check for more reliable analytical results. The improvement of the workflow offered by the 5-position rack and STAT handling like on Cobas Integra 800 makes the instrument attractive for further consolidation in the medium-sized laboratory, for dedicated use of special analytes, and/or as back-up in the large routine laboratory.