Asymmetric dimethylarginine (ADMA), symmetric dimethylarginine (SDMA) and L-arginine in patients with arteriogenic and non-arteriogenic erectile dysfunction.

The plasma concentration of asymmetrical dimethylarginine (ADMA), an inhibitor of nitric oxide synthase, has been linked to endothelial dysfunction. We investigated the relation between ADMA, symmetric dimethylarginine (SDMA) and L-arginine concentrations and erectile dysfunction. We compared plasma levels of ADMA, SDMA and L-arginine in 61 men in good health with erectile dysfunction of arteriogenic and non-arteriogenic origin. Diagnosis of erectile dysfunction was based on the International Index of Erectile Function Score and its aetiology was classified with penile echo-colour-Doppler in basal condition and after intracavernous injection of prostaglandin E1. The ADMA and SDMA concentrations were significantly higher in men with arteriogenic erectile dysfunction compared with those with erectile dysfunction of non-arteriogenic origin (p < 0.05) and the concentrations in both subgroups were significantly higher than in controls (p < 0.001). There was a negative correlation between ADMA and International Index of Erectile Function Score only in arteriogenic erectile dysfunction subgroup. L-arginine did not differ significantly neither between the two erectile dysfunction subgroups (p > 0.05) nor between each of the two erectile dysfunction subgroups and controls (p > 0.05). The L-arginine/ADMA and the L-arginine/SDMA ratios in arteriogenic erectile dysfunction subgroups were significantly lower than both in controls (p < 0.05) and in non-arteriogenic erectile dysfunction patients (p < 0.05); the two ratios in non-arteriogenic erectile dysfunction patients did not differ from those in the controls (p > 0.05). We conclude that ADMA and SDMA concentrations are significantly higher and L-arginine/ADMA ratio lower in patients who have arteriogenic erectile dysfunction compared with both patients with non-arteriogenic erectile dysfunction and controls. The negative correlation between ADMA and severity of erectile dysfunction is present only in patients with arteriogenic erectile dysfunction. This study supports the importance to always distinguish arteriogenic from non-arteriogenic erectile dysfunction patients to study the complicate erectogenic mechanisms that lead to erectile dysfunction and also to provide potential therapeutic agents for patients with arteriogenic erectile dysfunction.

Intra-operative testing for parathyroid hormone: the Central Laboratory option.

BACKGROUND: Intra-operative PTH testing in the operating theatre has proven to be an accurate way to verify the removal of all pathological parathyroid tissue in primary hyperparathyroidism. Its limitation is the high cost. An alternative, more cost-effective procedure is proposed: intra-operative PTH dosage at the Central Laboratory. PATIENTS AND METHODS: Fifty-four patients underwent parathyroidectomy with intraoperative dosage of PTH at the Central Laboratory. Three blood samples were taken from each patient: just after the induction of anesthesia, 5 and 10 min after parathyroidectomy. The samples were sent to the Central Laboratory and analysed simultaneously. The results were phoned back to the theatre. The procedure was considered effective when PTH drop was >/=50% from basal value, 10 min after parathyroidectomy. RESULTS: 98.1% of patients proved recovered (average follow- up 31.1 months). The procedure had 3 false negatives, 1 false positive, with sensitivity, specificity, accuracy, positive predictive value and negative predictive value of 94.0%, 75.0%, 92.6%, 97.9%, and 50.0%, respectively. DISCUSSION AND CONCLUSION: The main disadvantage of the presented procedure is the long waiting time. Nevertheless this time is the same as that required for results from intra-operative histological examination, the only alternative to determine surgery effectiveness in centres where portable instrumentation for intra-operative PTH dosage in the operating theatre is not available. The advantage of intra-operative PTH at the Central Laboratory is the very low cost. If results in terms of sensitivity, specificity, accuracy, and cost are taken into consideration, intra-operative dosage of PTH at the Central Laboratory may be deemed a viable alternative to the operating theatre.

Levels of l-arginine and l-citrulline in patients with erectile dysfunction of different etiology.

Nitric oxide is a physiologic signal essential to penile erection. l-citrulline (l-Cit) is converted into l-arginine (l-Arg), the precursor from which nitric oxide is generated. The level of l-Arg and l-Cit in the field of male sexual function remains relatively underexplored. The aim of the study was to evaluate the level of serum l-Arg and of l-Cit in a group of patients with erectile dysfunction. Diagnosis and severity of erectile dysfunction was based on the IIEF-5 and its etiology was classified as arteriogenic (A-ED), borderline (BL-ED), and non-arteriogenic (NA-ED) with penile echo-color-Doppler in basal condition and after intracaversous injection of prostaglandin E1. Serum l-Arg and l-Cit concentrations were measured by a cation-exchange chromatography system. l-Arg and l-Cit levels of men with A-ED were compared with those of male with BL-ED and NA-ED. Median level of l-Arg and l-Cit in 122 erectile dysfunction patients (41 A-ED, 23 ED-BL, 58 NA-ED) was 82.7 and 35.4 µmol/L, respectively. l-Arg and l-Cit levels in control patients were not significantly different (p = 0.233 and p = 0.561, respectively) than in total erectile dysfunction patients. l-Arg and l-Cit levels in control patients were significantly higher (p < 0.001 and p < 0.018, respectively) than in A-ED patients, but no difference (p > 0.50) was observed in controls and in both BL-ED and NA-ED patients. Patients with severe/complete-erectile dysfunction (IIEF-5 < 10) had l-Arg or l-Cit level significantly lower (-17%, p < 0.03; -13%, p < 0.04) and were more frequent (p < 0.01 and p < 0.04) under the respective median level (82.7 and 35.4 µmol/L) than those with mild-erectile dysfunction (IIEF-5 = 16-20). l-Arg and l-Cit levels in A-ED were significantly lower (p < 0.007 and p < 0.001, respectively) than in NA-ED patients. Penile echo-color-Doppler revealed that A-ED (peak systolic velocity ≤ 25 cm/sec) was more frequent in men with l-Arg under 82.7 µmol/L or l-Cit under 35.4 µmol/L and in the same population, the median peak systolic velocity values were lower in l-Arg deficient (29 vs. 35; p < 0.04) and also in l-Cit deficient (31 vs. 33, p > 0.3) but without reaching the statistical significance. Our study shows that a significant proportion of erectile dysfunction patients have low l-Arg or l-Cit level and that this condition is more frequent in patients with arteriogenic etiology. Low levels of these nitric oxide synthase substrates might increase the erectile dysfunction risk by reducing the concentration of nitric oxide.

Mechanistic aspects of the relationship between low-level chemiluminescence and lipid peroxides in oxidation of low-density lipoprotein.

In this study oxidation of low-density lipoprotein (LDL) induced by different Cu2+ concentrations was investigated. Lipid peroxidation was assessed by monitoring low-level chemiluminescence (LL-CL), conjugated diene hydroperoxide (CD) and alpha-tocopherol (TocOH), the major lipophilic antioxidant in LDL. At high Cu2+ concentration, LDL oxidation was characterised by CD formation, LL-CL emission and TocOH consumption. At low Cu2+ concentration, CD formation was independent of LL-CL and occurred in the presence of TocOH. Thus, two different mechanisms lead to lipid peroxide formation in LDL. The combination of CD assay and LL-CL monitoring makes it possible to distinguish the autocatalytic mechanism of CD formation and that associated with TocOH, found at a high and a low rate of initiation, respectively.

Clinical significance of serum pancreatic enzymes in the quiescent phase of chronic pancreatitis.

The most commonly used serum enzymes in pancreatic diseases are total amylase, pancreatic isoamylase, lipase and trypsin. To determine which of these enzymes is the most useful in the diagnosis of clinically quiescent chronic pancreatitis and which enzyme best reflects exocrine functional reserve, we studied 22 healthy control subjects, 44 patients with gastrointestinal, liver and biliary tract diseases, and 25 patients with chronic pancreatitis. On the basis of duodenal intubation, the latter were divided into two subgroups: one group of 13 patients with slight to moderate secretion deficiency and another of 12 patients with severe exocrine insufficiency. Of the enzymes studied, lipase, trypsin and pancreatic isoamylase are equally suitable for the evaluation of function in severe chronic pancreatitis, but not for the early diagnosis of the disease. Results for total amylase are not reliable so that its use in the study of chronic pancreatitis is not advisable.

Fecal chymotrypsin: a new diagnostic test for exocrine pancreatic insufficiency in children with cystic fibrosis.

The purpose of this report is to evaluate whether a new, simple, non-invasive method for chymotrypsin measurement in stools is useful for the diagnosis of exocrine pancreatic insufficiency in cystic fibrosis (CF). A hundred children aged from 2 months to 12 years were tested: 50 children had been admitted for chronic diarrhoea, 15 for cystic fibrosis and 40 acted as controls. Chymotrypsin in stools was assayed using a kinetic measurement with Succ-Ala-Ala-Pro-Phe-pNa as substrate in a simple photometric assay. In 13 of 15 children with cystic fibrosis, stool enzyme levels were always remarkably low, while all control subjects and all children not presenting cystic fibrosis had normal stool levels of chymotrypsin. Our data suggest that stool chymotrypsin measurement is a simple and reliable "tubeless" test for the evaluation of exocrine pancreatic insufficiency in children with cystic fibrosis.

Increased urinary N-acetyl-beta-glucosaminidase (NAG) excretion in young insulin-dependent diabetic patients.

We evaluated urinary N-acetyl-beta-glucosaminidase (NAG) excretion in overnight and in second morning urine in 50 young diabetic patients, aged 7.4-25 years with a disease duration from 2-19.6 years. In all patients we evaluated urinary NAG and creatinine excretion, in both overnight and second morning urine, glycosuria, fasting blood glucose and HbA1c levels, insulin requirement, blood pressure, and the presence of microangiopathic complications. Urinary NAG excretion was also evaluated in 69 age- and sex-matched controls. NAG was determined using 3-cresolsulfonphtaleinyl-beta-N-acetylglucosaminide as substrate (Boehringer Mannheim, Germany). In the diabetic patients NAG/Cre ratios were significantly higher than in controls both in overnight and second morning urine (P < 0.0005, respectively). We observed significantly higher NAG/Cre ratio levels in the second morning than in overnight urine, both in controls and in diabetics (P < 0.0005, respectively). Elevated (above 2 S.D. of the mean) NAG/Cre ratios were found in 17/50 patients (34%) in overnight urine and in 29/50 (58%) in second morning urine. No correlation was observed between NAG/Cre ratio levels and age, duration of disease, pubertal stage, body mass index, fasting blood glucose, glycosuria, insulin requirement and blood pressure. The patients with one or more complications did show NAG/Cre ratio levels significantly higher than those without complications (P < 0.005) in second morning urine, but not in overnight urine. Our study has demonstrated an increased rate of urinary NAG excretion in young IDDM patients, in particular in those with microangiopathic complications.

Exocrine pancreatic function in children and adolescents with insulin-dependent diabetes mellitus.

Exocrine pancreatic function was evaluated in 21 diabetic children on the basis of a p-aminobenzoic acid (PABA) test and a determination of fasting serum amylase, pancreatic isoamylase, lipase, trypsin and elastase levels. Fecal chymotrypsin was also measured. Compared to the controls, the diabetic children had significantly lower levels of trypsin (P less than 0.001) and elastase (P less than 0.02). Fecal chymotrypsin appeared to be significantly lower (P less than 0.01) in diabetic children than in controls but in all patients fecal chymotrypsin values registered above the limit considered to be normal. No significant correlation was observed between pancreatic enzyme concentrations, serum and urinary PABA values, and chronologic age, HbA1 and insulin requirement. Only for serum PABA a significant negative correlation with duration of disease (P less than 0.01) has been observed. These data show that exocrine pancreatic function may be abnormal in children with IDDM.

Effects of malnutrition on the acute phase response of plasma proteins in patients undergoing total gastrectomy.

The serum level of six acute phase proteins (APP) has been evaluated preoperatively and for a few days in the postoperative period. Thirty patients undergoing total gastrectomy for gastric cancer have been studied in two subgroups according to their nutritional status. Those with gastric cancer had significantly higher baseline serum levels of alpha 1 acid glycoprotein (alpha1AGP) and C-reactive protein (CRP) than a control group. Malnourished patients also had reduced acute phase response for alpha1AGP and alpha 1 antitrypsin (alpha1AT) a higher increase of CRP and fibrinogen, and a lower decrease for transferrin and retinol binding protein (RBP).

The influence of gender in the evaluation of platelet and plasma catecholamines.

The platelet and plasma levels of catecholamines (CA) were simultaneously studied in a group of normal subjects in order to find possible sex-related changes in the distribution of CA in these two compartments. No significant differences between males and females were observed, but a marked platelet noradrenaline increase was found in the luteal phase as compared to the follicular phase. Furthermore, the platelet and plasma CA levels were strongly correlated in the male group but not in the female group. These results, while confirming the existence of a menstrual-related variability in noradrenergic activity, suggest a sex-related difference in the dynamic balance between platelet and plasma CA levels. The simultaneous assay of platelet and plasma CA enabled this phenomenon to be revealed, whereas the separate evaluation of platelet or plasma CA levels would not have done so.

Reference values of neuroactive amino acids in the cerebrospinal fluid by high-performance liquid chromatography with electrochemical and fluorescence detection.

Sampling and HPLC analysis procedures for CSF amino acid determinations were evaluated. In order to increase sensitivity, a precolumn derivatization of amino acids by o-phthalaldehyde-mercaptoethanol reagent was used. By using fluorimetric and electrochemical detection in series, positive peak identification can be obtained in a single chromatographic run. It is recommended to analyze freshly collected CSF. Amino acids are stable for short periods over a wide range of temperature, but storage at -80 degrees C is recommended. The CSF samples for the calculation of the reference values were taken from 40 healthy subjects, hospitalized for lumbar disk herniation, placed on the same diet and kept drug-free for at least 1 week. The mean values (mumol/l) and the ranges (in parentheses) were: 0.27 (0.09-0.63), 0.62 (0.18-1.15), 5.32 (3.05-11.50), 6.16 (2.90-13.30), 0.16 (0.03-0.22) for aspartic acid, glutamic acid, glycine, taurine and gamma-aminobutyric acid respectively.

Determination of illicit drugs and related substances by high-performance liquid chromatography with an electrochemical coulometric-array detector.

A general method for the simultaneous determination of fifteen common drugs (6-acetylmorphine, 3,4-methylenedioxymetamphetamine, buprenorphin, cocaine, codeine, dihydrocodeine, ethylmorphine, heroin, hydrocodone, lidocaine, methadone, morphine, naloxone, procaine and thebaine) was developed using reversed-phase HPLC and electrochemical detection. The separation of the drugs was achieved by using as the mobile phase 20 mM monobasic sodium phosphate-acetonitrile (90:10) with a gradient to 50% of the organic modifier, on a silica based C18 column (150 x 4.6 mm I.D.) of 3 microns particle size and by the selectivity supplied by an array of eight coulometric electrodes at increasing potential. It was possible to identify and to determine fifteen different drugs in the same chromatographic run in 50 min. The method was tentatively applied to the determination of drugs in extracts of human hair.

PHA-stimulated cellular immune function and T-lymphocyte subsets in major depressive disorders.

We measured some immunological parameters in 20 hospitalized patients with major depression and 20 age- and sex-matched healthy controls. Both enumeration of immune cells, including T-lymphocyte subpopulations, and assay of T-cell function were studied. White blood cells were evaluated with an automated cell counter, T-cell subsets with an immunobead technique, and T-cell function with a phytohemagglutinin-induced proliferation in vitro assay. We found that T-lymphocyte responses to the mitogen were significantly lower in depressed patients than in controls. All the other parameters were normal. These findings suggest that functional but not numerical changes in T-lymphocytes characterize major depressive disorders.

Performance and clinical significance of the new fructosamine assay in diabetic patients.

The performance of the assay and the clinical usefulness of a new short-term index (Fructosamine Plus: FP) of metabolic control in diabetes was evaluated. The FP concentration was determined with the Hitachi 704, Cobas Mira, and Cobas Fara analyzers. The "within run" and "between run" CVs were less than 2.67% and less than 4.26%, respectively. The reference interval (determined from data on 125 blood donors) was 191 to 288 mumol/l (mean 240 mumol/l). In poorly controlled diabetic patients FP was significantly higher than in the well controlled ones. Moreover FP was significantly correlated with HbA1c and blood glucose levels. Our preliminary data seem to demonstrate that relating FP to total protein does not improve the clinical value of the new index.

Levels of human erythrocyte membrane-bound and cytosolic glycohydrolases are associated with oxidative stress in erectile dysfunction patients.

Oxidative stress (OS) and production of NO, by endothelium nitric oxide synthetase (eNOS), are involved in the pathophysiology of erectile dysfunction (ED). Moreover, OS induces modifications of the physicochemical properties of erythrocyte (RBC) plasma membranes and of the enzyme content of the same membranes. Due to their role in signalling early membrane alterations in OS-related pathologies, several plasma membrane and cytosolic glycohydrolases of human RBC have been proposed as new markers of cellular OS. In RBC, NOS can be activated and deactivated by phosphorylation/glycosylation. In this regulatory mechanism O-ß-N-AcetylGlucosaminidase is a key enzyme. Cellular levels of O-GlcNAcylated proteins are related to OS; consequently dysfunctional eNOS O-GlcNAcylation seems to have a crucial role in ED. To elucidate the possible association between RBC glycohydrolases and OS, plasma hydroperoxides and antioxidant total defenses (Lag-time), cytosolic O-ß-N-AcetylGlucosaminidase, cytosolic and membrane Hexosaminidase, membrane ß-D-Glucuronidase, and α-D-Glucosidase have been studied in 39 ED patients and 30 controls. In ED subjects hydroperoxides and plasma membrane glycohydrolases activities are significantly increased whereas Lag-time values and cytosolic glycohydrolases activities are significantly decreased. These data confirm the strong OS status in ED patients, the role of the studied glycohydrolases as early OS biomarker and suggest their possible use as specific marker of ED patients, particularly in those undergoing nutritional/pharmacological antioxidant therapy.

Performance assessment of a glucose control protocol in septic patients with an automated intermittent plasma glucose monitoring device.

BACKGROUND & AIMS: The optimal level and modality of glucose control in critically ill patients is still debated. A protocolized approach and the use of nearly-continuous technologies are recommended to manage hyperglycemia, hypoglycemia and glycemic variability. We recently proposed a pato-physiology-based glucose control protocol which takes into account patient glucose/carbohydrate intake and insulin resistance. Aim of the present investigation was to assess the performance of our protocol with an automated intermittent plasma glucose monitoring device (OptiScanner™ 5000). METHODS: OptiScanner™ was used in 6 septic patients, providing glucose measurement every 15' from a side-port of an indwelling central venous catheter. Target level of glucose was 80-150 mg/dL. Insulin infusion and kcal with nutritional support were also recorded. RESULTS: 6 septic patients were studied for 319 h (1277 measurements); 58 [45-65] hours for each patient (measurements/patient: 231 [172-265]). Blood glucose was at target for 93 [90-98]% of study time. Mean plasma glucose was 126 ± 11 mg/dL. Only 3 hypoglycemic episodes (78, 78, 69 mg/dL) were recorded. Glucose variability was limited: plasma glucose coefficient of variation was 11.7 ± 4.0% and plasma glucose standard deviation was 14.3 ± 5.5 mg/dL. CONCLUSIONS: The local glucose control protocol achieved satisfactory glucose control in septic patients along with a high degree of safeness. Automated intermittent plasma glucose monitoring seemed useful to assess the performance of the protocol.

Daily monitoring of biomarkers of sepsis in complicated long-term ICU-patients: can it support treatment decisions?

BACKGROUND: Diagnosis/grading of infection and the systemic response to infection may be difficult on admission to the intensive care unit, but it is even more complicated for severely ill patients with long intensive care stays. The ACCP-SCCM criteria are difficult to apply for such patients, and objective, validated biomarkers would be of great use in this setting. METHODS: Long-term (>6 days) critically ill patients in the general ICU of University Hospital were prospectively enrolled in the study. All patients were assessed daily by the attending physician using the ACCP-SCCM classification. C-reactive protein (CRP, mg/dL), procalcitonin (PCT, ng/mL), and interleukin-6 (IL-6, pg/mL) of daily stored sera were measured after each patient's discharge. After discharge, an independent, overall clinical evaluation and an a posteriori ACCP-SCCM classification were chosen as the reference standard for all comparisons. The assessor was aware of the patient's clinical course but was blinded to levels of biomarkers. RESULTS: We studied clinical variables and biomarkers of 26 patients over a total of 592 patient days. The day-by-day ACCP-SCCM classification of the attending physician overestimated the severity of the inflammatory response to infection. The diagnostic discriminative ability of severe-sepsis/septic-shock for PCT was high (ROC area 0.952 [0.931-0.973]) and had a best threshold value of 1.58 (83.7% sensitivity, 94.6 % specificity). IL-6 had better discriminative ability than CRP, but both were worse than PCT. CONCLUSION: PCT > 0.43 ng/mL could add to the clinical propensity for sepsis vs. SIRS not related to infection. Values higher than 1.58 ng/mL may support the bedside clinical diagnosis of severe-sepsis. PCT between 0.5 and 1.0 suggest tight daily monitoring of clinical conditions and re-evaluation of PCT.

Determination of free DOPA and 3-O-methyl-DOPA in human plasma by high-performance liquid chromatography with electrochemical detection.

A procedure was devised for the determination of the unconjugated non-protein-bound fraction of 3,4-dihydroxyphenylalanine (DOPA) and 3-O-methyl-DOPA (3-OMD) in plasma using a reversed-phase liquid chromatographic system coupled with electrochemical detection. Sample preparation involves rapid isolation of the unbound drugs from the drug-protein complex by ultrafiltration through a membrane with a molecular weight cut-off of 10,000 dalton. One chromatographic run requires less than 10 min. The relative standard deviation is less than 3% for the within-assay imprecision and less than 4% for the between-assay imprecision. The detection limits for DOPA and 3-OMD are 0.2 and 1.3 ng/ml, respectively.

Biological variability of myoglobin in healthy elderly and younger subjects.

To study the effect of age on serum myoglobin more clearly, the analytical, intra-individual and inter-individual components of variation were estimated from duplicate analyses of specimens collected from 18 healthy elderly subjects [ages 74-97 years; 9 men (EM)], and 14 healthy younger subjects [ages 25-31 years; 7 men (YM)] over a period of 6 weeks. The mean values (microgram/L) were EM: 53.7; EW: 44.9; YM: 34.2; YW:24.8. Estimated analytical (CVA), intra- (CVI) and inter-individual (CVG) variations as CV% were: CVA: 2.2. CVI: EM: 13; EW: 9.9; YM: 12.4; YW: 9.6. CVG: EM: 37.6; EW: 28; YM: 18.5; YW: 13.4. The data obtained were used to derive the desirable analytical goal for imprecision (i.e., < or = 6.5% in EM; < or = 4.9% in EW and < or = 6.2% in YM; < or = 4.8% in YW); inaccuracy (i.e., < or = 9.9% in EM; < or = 7.7 in EW and < or = 5.5% in YM; < or = 4.12% in YW); the change required for serial results to be significantly different (i.e., 36% in EM; 28% in EW and 34% in YM; 27.2% in YW), the numbers of specimen collections required to produce a more precise estimate of the homeostatic set point of an individual within 5% (i.e., 26 in EM; 16 in EW and 24 in YM; 15 in YW), and the index of individuality (i.e., 0.34 in EM; 0.35 in EW and 0.67 in YM; 0.71 in YW). This study shows that intra-individual biological variation of myoglobin in healthy elderly subjects is not different from that in young subjects. Inter-individual variation, instead, is greatly influenced by differences in age and sex.