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Reports the conduction of biochemical tests for diagnosis of pheochromocytoma. Evaluation of urine and plasma; Reliability of metanephrine in the confirmation of the disease; Applicability of the test in patients with intermittent hypertension.

Shiga toxin producing Escherichia coli (STEC) are noninvasive enteric pathogens that may cause hemorrhagic colitis (HC) and diarrhea-associated hemolytic uremic syndrome (D+ HUS). We hypothesized that development of D+ HUS is associated with increased serum procalcitonin (PCT) levels. PCT was measured by an immunoluminometric assay in 113 patients. Concentrations of PCT were different in normal controls, disease control groups (rotavirus enteritis, HC due to non-STEC pathogens, chronic renal failure), and children with uncomplicated O157:H7 HC or D+ HUS. Children with D+ HUS showed higher PCT levels than those with uncomplicated O157:H7 HC, and increased concentrations were noted in cases requiring peritoneal dialysis. Severely increased concentrations were observed in children with D+ HUS on d 5 or 6 after the onset of enteritis, whereas serial measurements in those with uncomplicated O157:H7 HC remained within the normal range throughout the first week of illness. PCT was correlated with serum concentrations of lipopolysaccharide (LPS)-binding protein and serum levels of alanine aminotransferase. Using two separate sets of real-time PCR primers, we were unable to detect elevated PCT mRNA transcripts in nonadherent undifferentiated (monocytic) or differentiated (macrophage-like) THP-1 cells stimulated with purified Shiga toxin-1 and/or LPS. Our data show that serum levels of PCT are associated with the severity of illness in children with D+ HUS. Further studies are needed to determine the role of PCT in the pathogenesis of thrombotic microangiopathy associated to childhood D+ HUS.

A significant decrease of DPP IV activity has been found in patients with severe sepsis in relationship to the increase of procalcitonin. These findings might be explained by the high concentration of other substrates for DPP IV present in these patients. It can be hypothesized that this enzymatic decrease is bound to some changes in immunomodulation. Further studies will be necessary to elucidate the clinical importance of these findings.

Procalcitonin (PCT) has been described as an early and discriminating marker of bacteria-associated sepsis in patients. However, little is known of its source and actions, especially with regard to its relation to tumor necrosis factor (TNF). TNF is responsible for the release of several other mediators of sepsis e.g., chemokines. We tested the hypothesis that plasma PCT levels during sepsis differ with regard to the degree of TNF availability. Severe hyperdynamic sepsis was induced in baboons (n = 14) by i.v. infusion of live E. coli (approximately 2 x 10(9) colony-forming units/kg) over 2 h. Animals were pretreated 2 h before E. coli either with 1 mg/kg humanized anti-TNF antibody (CDP571) or placebo (Ringer solution). Plasma PCT levels at baseline was barely detectable, but increased to about 4000 pg/mL at 4 h after E. coli infusion. Levels were maximal between 8 and 24 h and had returned nearly to baseline at 72 h. Although no TNF could be measured in the treated group, PCT levels were not different between the placebo and the TNF antibody treatment group. We conclude that PCT levels are not dependent on the systemic presence of TNF in an E. coli sepsis model in baboons. Such sepsis induced PCT release is clearly different from the previously reported PCT release during infusion of rhTNF in volunteers or chimpanzees.

Release of toxins in the organism trigger a cascade of biological and chemical events. The process involves a large number of molecules produced under genetic control in a perfectly regulated chronological order. Certain molecules (TNFx, IL-1, Il-2.) have inflammatory proprieties, generally producing systemic Inflammatory Response Syndrome (SIRS). Other less numerous molecules (IL-4, IL-10) have antiinflammatory actions. Finally, soluble receptors and these cytokines contribute to the decrease in the quantity of active cytokines at the receptor level. Dozens of other molecules, many of which remain to be fully understood, are also found in the blood stream. They can, for example, facilitate white cell adhesion (ICAM, VCAM, selectins.). Some of them (G-CSF. CM-CSF. IL-5) stimulate production of granulocytes, monocytes, eosinophils. More recently, certain peptides, like macrophage inhibiting factor (MIF) and procalcitonin (PCT) have been added to the list of molecules involved in bacterial infections. Coagulation factors are also very rapidly released in response to toxinic aggression triggering disseminated intravascular coagulation. Later, acute-hase inflammation proteins, such as C-reactive protein (CRP), haptoglobin, serum amyloid A, etc. are found in largely increased quantities. All these molecules are potential markers of inflammation. Only a few have however been retained for routine assay due to their fragility and their short-half life or due to analytical difficulties. CRP and PCT can however be used to differentiate viral infections from bacterial infections and are thus routinely used in clinical applications. In the second part of this review, we briefly discuss therapeutic perspectives for severe infections and septic shock. There have been many attempts to neutralize different cytokines but results have been disappointing to date. There are however many possibilities currently under study, particularly neutralization of endotoxins, immunomodulation, use of recombinant C and S proteins, etc.

Aims: To assess the sensitivity, specificity, and predictive value of procalcitonin (PCT) in differentiating bacterial and viral causes of pneumonia., Methods: A total of 72 children with community acquired pneumonia were studied. Ten had positive blood culture for Streptococcus pneumoniae and 15 had bacterial pneumonia according to sputum analysis (S pneumoniae in 15, Haemophilus influenzae b in one). Ten patients had Mycoplasma pneumoniae infection and 37 were infected with viruses, eight of whom had viral infection plus bacterial coinfection. PCT concentration was compared to C reactive protein (CRP) concentration and leucocyte count, and, if samples were available, interleukin 6 (IL-6) concentration., Results: PCT concentration was greater than 2 microg/l in all 10 patients with blood culture positive for S pneumoniae; in eight of these, CRP concentration was above 60 mg/l. PCT concentration was greater than 1 microg/l in 86% of patients with bacterial infection (including Mycoplasma and bacterial superinfection of viral pneumonia). A CRP concentration of 20 mg/l had a similar sensitivity but a much lower specificity than PCT (40% v 86%) for discriminating between bacterial and viral causes of pneumonia. PCT concentration was significantly higher in cases of bacterial pneumonia with positive blood culture whereas CRP concentration was not. Specificity and sensitivity were lower for leucocyte count and IL-6 concentration., Conclusions: PCT concentration, with a threshold of 1 microg/l is more sensitive and specific and has greater positive and negative predictive values than CRP, IL-6, or white blood cell count for differentiating bacterial and viral causes of community pneumonia in untreated children admitted to hospital as emergency cases.

Objective: To evaluate the accuracy of procalcitonin (PCT) in predicting bacterial infection in ICU medical and surgical patients., Setting: A 10-bed medical surgical unit., Design: PCT, C-reactive protein (CRP), interleukin 6 (IL-6) dosages were sampled in four groups of patients: septic shock patients (SS group), shock without infection (NSS group), patients with systemic inflammatory response syndrome related to a proven bacterial infection (infect. group) and ICU patients without shock and without bacterial infection (control group)., Results: Sixty patients were studied (SS group:n=16, NSS group,n=18, infect. group,n=16, control group,n=10). The PCT level was higher in patients with proven bacterial infection (72+/-153 ng/ml vs 2.9+/-10 ng/ml,p=0.0003). In patients with shock, PCT was higher when bacterial infection was diagnosed (89 ng/ml+/-154 vs 4.6 ng/ml+/-12,p=0.0004). Moreover, PCT was correlated with severity (SAPS:p=0.00005, appearance of shock:p=0.0006) and outcome (dead: 71.3 g/ml, alive: 24.0 g/ml,p=0.006). CRP was correlated with bacterial infection (p<10(-5)) but neither with SAPS nor with day 28 mortality. IL-6 was correlated with neither infection nor day 28 mortality but was correlated with SAPS. Temperature and white blood cell count were unable to distinguish shocked patients with or without infection. Finally, when CRP and PCT levels were introduced simultaneously in a stepwise logistic regression model, PCT remained the unique marker of infection in patients with shock (PCT> or =5 ng/ml, OR: 6.2, 95% CI: 1.1-37,p=0.04)., Conclusion: The increase of PCT is related to the appearance and severity of bacterial infection in ICU patients. Thus, PCT might be an interesting parameter for the diagnosis of bacterial infections in ICU patients.

Objective: Procalcitonin concentration increases in bacterial infections but remains low in viral infections and inflammatory diseases. The change is rapid and the molecule is stable making it a potentially useful marker for distinguishing between bacterial and viral infections., Patients and Methods: Procalcitonin (PCT) was determined with an immunoluminometric assay on plasma collected at admission in 436 infants and children hospitalized for bacterial or viral infection. It was compared with C reactive protein, interleukin-6 and interferon-alpha measured on the same sample., Results: PCT was 41.3 +/- 77.4 micrograms/l in children with septicemia or bacterial meningitis (n = 53), 0.39 +/- 0.57 microgram/l in children with viral infection (n = 274) and 3.9 +/- 5.9 micrograms/l in children with a localized bacterial infection who had a negative blood culture (n = 109). PCT was > 1 microgram/l in 126 children with a localized or systemic bacterial infection (sensitivity 78%). PCT was < 1 microgram/l in 258 children with a viral infection (specificity 94%). For differenciation between viral and bacterial infections, CRP value > or = 20 mg/l, IL-6 > 100 pg/ml and interferon-alpha > 0 Ul/ml have 85, 48 and 76% sensitivity and 73, 85 and 92% specificity respectively., Conclusions: In this study, a PCT value of 1 microgram/l or greater had better specificity, sensitivity and predictive value than CRP, IL-6 and interferon-alpha in children for distinguishing between viral and bacterial infections. PCT may be useful in pediatric emergency room for making decision about antibiotic treatments.

Increased concentrations of procalcitonin (PCT) are found in the plasma of patients with thermal injury and in patients with sepsis and severe infection, making this molecule important as a diagnostic and prognostic marker in these diseases. Interestingly, only the truncated form of PCT, PCT(3-116), is present in the plasma of these patients. The enzyme responsible for this truncation is unknown as yet. Here, using capillary zone electrophoresis, mass spectrometry and Edman sequence analysis, we demonstrate that dipeptidyl peptidase IV (DP IV, EC 3.4.14.5) is capable of catalyzing the hydrolysis of PCT(1-116), releasing the N-terminal dipeptide Ala-Pro. We hypothesize that PCT(3-116) is the result of the hydrolysis of PCT(1-116) by soluble DP IV of the blood plasma or by DP IV expressed on the surface of cells.

Background: Procalcitonin (PCT) concentration increases in bacterial infections but remains low in viral infections and inflammatory diseases. The change is rapid and the molecule is stable, making it a potentially useful marker for distinguishing between bacterial and viral infections., Methods: PCT concentration was determined with an immunoluminometric assay on plasma collected at admission in 360 infants and children hospitalized for bacterial or viral infection. It was compared with C-reactive protein (CRP), interleukin 6 and interferon-alpha measured on the same sample., Results: The mean PCT concentration was 46 microg/l (median, 17.8) in 46 children with septicemia or bacterial meningitis. PCT concentration was > 1 microg/l in 44 of 46 in this group and in 59 of 78 children with a localized bacterial infection who had a negative blood culture (sensitivity, 83%). PCT concentration was > 1 microg/l in 16 of 236 children with a viral infection (specificity, 93%). PCT concentration was low in 9 of 10 patients with inflammatory disease and fever. A CRP value > or =20 mg/l was observed in 61 of 236 patients (26%) with viral infection and in 105 of 124 patients (86%) with bacterial infection. IL-6 was > 100 pg/ml in 14% of patients infected with virus and in 53% with bacteria. A secretion of interferon-alpha was found in serum in 77% of viral infected patients and in 8.6% of bacterial infected patients., Conclusions: In this study a PCT value of 1 microg/l or greater had better specificity, sensitivity and predictive value than CRP, interleukin 6 and interferon-alpha in children for distinguishing between viral and bacterial infections. PCT values are higher in invasive bacterial infections, but the cutoff value of 1 microg/l indicates the severity of the disease in localized bacterial infection and helps to decide antibiotic treatment in emergency room. PCT may be useful in an emergency room for differentiation of bacterial vs. viral infections in children and for making decisions about antibiotic treatments.

Objective: In an attempt to differentiate acute pyelonephritis from lower urinary tract infection (UTI), we measured serum procalcitonin levels, a recently described marker of infection. We compared it with other commonly used inflammatory markers and evaluated its ability to predict renal involvement as assessed by dimercaptosuccinic acid (DMSA) scintigraphy., Methods: Serum C-reactive protein, leukocyte counts, and procalcitonin levels were measured in 80 children, 1 month to 16 years of age, admitted for suspected pyelonephritis. Renal involvement was assessed by 99mTe-DMSA scintigraphy in the first 5 days after admission. The examination was repeated at least 3 months later if the first result was abnormal., Results: In lower UTI, the mean procalcitonin (PCT) was 0.38 micrograms/L +/- 0.19 compared with 5.37 micrograms/L +/- 1.9 in pyelonephritis. In these two groups, respectively, leukocyte counts were 10939/mm3 +/- 834 and 17429/mm3 +/- 994, and C-reactive protein (CRP) levels were 30.3 mg/L +/- 7.6 and 120.8 mg/L +/- 8.9. When inflammatory markers were correlated to the severity of the renal lesion as ranked by DMSA scintigraphy, we found a highly significant correlation with plasma levels of PCT, but borderline significance with CRP and none with leukocyte counts. Patients without vesicoureteral reflux had a mean PCT of 5.16 micrograms/L +/- 2.33, which was not significantly different from that in patients with reflux who had a mean PCT of 5.76 micrograms/L +/- 3.49. For the prediction of renal lesions at admission, CRP had a sensitivity of 100% and a specificity of 26.1%. The sensitivity and specificity of PCT were 70.3% and 82.6%, respectively., Conclusion: We conclude that serum PCT levels were increased significantly in children with febrile UTI when renal parenchymal involvement (assessed by DMSA scintigraphy) was present and allowed for prediction of patients at risk of severe renal lesions.

Objectives: In young children with meningitis, blood or cerebrospinal fluid (CSF) analysis cannot differentiate all cases of viral meningitis (VM) from bacterial meningitis (BM). Empirical antibiotic therapy is often given. As new markers are needed, we compared serum proCalcitonin (PCT) with CSF analysis for C-reactive protein (CRP) and interleukin-6 (IL6)., Patients and Methods: PCT was measured with a chemoluminescent assay in the sera of 23 children (aged 3 months to 14 years) hospitalized for BM and in 51 patients with VM., Results: Initial CRP (mean 143.3 mg/l, range 28-351 and mean 13.9, range 1-48), CSF proteins (mean 2.2, range 0.4-4.74 and mean 0.57, range 0.12-2.72) and white blood cell count in CSF (range 240-17500 and 20-3200) in BM and VM respectively, were not sufficiently discriminative to distinguish between BM and VM. Twenty-four of the 51 patients with VM were given antibiotics. IL6 values at admission showed an overlap zone (> 100 pg/ml in 7/19 patients with VM and < 100 pg/ml in 1/8 patients with BM. PCT was discriminative in all cases: mean PCT in BM was 61 micrograms/l (range 4.8-335) and 0.33 in VM (range 0-1.7; p < 0.001). No production of PCT was detected in CSF. After antibiotic therapy, PCT decreased and reached undetectable levels after recovery., Conclusion: PCT is a sensitive and specific marker for early diagnosis of viral meningitis versus bacterial meningitis in children.

Objectives: To describe the initial evolution of serum procalcitonin (PCT) and C-reactive protein (CRP) in previously healthy adult trauma patients and to compare the relationship of the expression of these two proteins with indicators of trauma severity., Design: Prospective, descriptive, longitudinal study., Setting: Surgical ICU in an university hospital., Patients: Twenty-one patients admitted during the first posttraumatic 3 h exhibiting an Injury Severity Score (ISS) between 16 and 50 were enrolled., Measurements: Blood sampling was performed on admission and on posttraumatic days 0.5, 1, 2 and 3 to assess serum levels of PCT and CRP. Total creatine kinase (CKtot) and lactate dehydrogenase (LDHtot) activities in the serum were used as tissue damage indicators., Results: PCT exhibited an early and transient increase in serum levels similar to a more delayed change of CRP levels. Peak PCT and peak CRP were related to the ISS, the extent of tissue damage and the amount of fluid replacement during the first day. During the first 3 posttraumatic days, 90% of the patients exhibited a generalized inflammatory syndrome without infection., Conclusions: An early and transient release of PCT into the circulation was observed after severe trauma and the amount of circulating PCT seemed proportional to the severity of tissue injury and hypovolemia, yet unrelated to infection. The predictive value of both PCT and CRP for a forthcoming multiple organ failure still remains to be clarified.

Procalcitonin (PCT) is a new marker connected to systemic bacterial infection. Blood values are parallel to the severity of the disease. In the present Knowledge on PCT, the usefulness is focused on acute pediatric pathology, ICU, and the follow up of grafts and surgery. This paper dwells on the interest in the differential diagnosis for meningitis (viral versus bacterial). At the opposite of CRP and IL6, a very clear cut off for all the cases has been found. The cut off in this study is about 2-3 micrograms/l. PCT, at the difference of cytokines is a very stable molecule in the blood sample. Also a very small quantity of serum (or plasma) 20 microliters is sufficient for one assay. In the future, a point of care assay will be available and should be very interesting in the emergency wards (pediatric or adult ICU). The origin of PCT seems to be--but perhaps not exclusively--mononuclear cells. The absence of an animal model (except monkeys) is actually a difficulty to progress.

Procalcitonin (ProCT) is a recently described marker of severe sepsis. It was decided to assess the value of proCT as a marker of secondary infection in patients infected with HIV-1. ProCT plasma levels were measured by immunoluminometric assay in a prospective study in 155 HIV-infected individuals: 102 asymptomatic and 53 with lever or suspected secondary infections. The baseline plasma level of ProCT was low (0.5 ng/ml +/- 0.37), even in the latest stages of the disease, and did not differ from the values of healthy subjects (0.54 ng/ml +/- 0.08). EDTA-treated whole blood was collected from patients before starting specific antimicrobial therapy. No elevation of ProCT level was detected in HIV-infected patients with evolving secondary infections including PCP (n = 4), cerebral toxoplasmosis (n = 4), viral infections (n = 9), mycobacterial infections (n = 5), localized bacterial (n = 12) and fungal infections (n = 4), malignancies (n = 3), and in various associated infectious and non-infectious febrile events (n = 13). All these plasma values were lower than 2.1 ng/ml. In contrast, high ProCT plasma levels were detected in one HIV-infected patient with a septicaemic Haemophilus influenzae infection (16.5 ng/ml) and another one with a septicaemic Pseudomonas aeruginosa infection (44.1 ng/ ml), ProCT values decreased rapidly under appropriate therapy. ProCT seems to be a specific marker of bacterial sepsis in HIV-infected patients, as no increase in other secondary infections could be detected in those patients. A rapid determination of ProCT level could be useful to confirm or refute bacterial sepsis for a better management of febrile HIV-infected patients.

We measured the plasma procalcitonin levels in 59 children who were admitted to the hospital because of bacterial or viral meningitis. Eighteen children with acute bacterial meningitis had elevated procalcitonin levels (mean level, 54.5 micrograms/L; range, 4.8-110 micrograms/L). The procalcitonin levels in 41 children with viral meningitis were low (mean level, 0.32 micrograms/L; range, 0-1.7 micrograms/L; P < .0001). Assay of cerebrospinal fluid (CSF) cells and proteins and serum C-reactive protein showed a zone of overlapping values between the two groups. Procalcitonin was not produced in CSF. Plasma procalcitonin levels decreased rapidly during antibiotic therapy. These data suggest that the measurement of plasma procalcitonin might be of value in the differential diagnosis of meningitis due to either bacteria or viruses.

To determine the evolution and significance of circulating procalcitonin (ProCT), IL-6 TNF alpha and endotoxin levels early after thermal injury, we performed a prospective, single unit, longitudinal study. Forty burn patients with total body surface area (TBSA) > 30 per cent were studied, of whom 33 suffered an inhalation injury. Blood samples were taken on the day of admission, every 4 h during the first day and daily during the first week. All patients had increased ProCT and IL-6 levels without any proven infection. Endotoxin and TNF alpha levels remained very low or undetectable. ProCT and IL-levels correlated well with the severity of skin burn injury (respectively, p < 0.006 and p < 0.028, using the non-parametric Kruskal-Wallis test). ProCT levels are not associated with smoke inhalation. ProCT and IL6 are prognostic factors of mortality at the time of admission but less reliable than the clinical UBS (unit burn standard) score. Endotoxin and TNF alpha were undetectable, suggesting that the problem of the early gut bacterial translocation remains to be proven.

Objectives: To determine and compare the respective concentrations of tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, soluble TNF receptors, nitrite/nitrate (NO2-/NO3-), and procalcitonin in the plasma of patients with septic shock, cardiogenic shock, and bacterial pneumonia without shock; and to assess the predictive value of these mediators in defining patients with septic shock., Design: Cohort study, comparing normal volunteers (controls) and patients with septic shock, cardiogenic shock, and bacterial pneumonia., Setting: A collaborative study among an intensive care unit, an emergency room, and three research laboratories., Patients: Mediators were measured at various times in 15 patients with septic shock (during the shock phase and during the recovery phase), in seven patients with cardiogenic shock during the shock phase, and in seven patients with severe bacterial pneumonia on day 1 of admission., Interventions: Blood samples were collected at various times during the course of the disease., Measurements and Main Results: TNF-alpha values were highest in the acute phase of septic shock (53 to 131 pg/mL during septic shock), while patients with bacterial pneumonia had intermediate concentrations (32 pg/mL). TNF-alpha concentrations were normal in patients with cardiogenic shock. IL-6 concentrations were highest in patients with acute septic shock (85 to 385 pg/mL). However, in contrast to TNF-alpha concentrations, IL-6 concentrations were normal in patients with bacterial pneumonia and increased in patients with cardiogenic shock (78 pg/mL). Soluble TNF receptors were increased in all three groups vs. controls, with the highest increase in patients with septic shock. NO2-/NO3- concentrations were highest (72 to 140 mM) in patients with septic shock, and were < 40 mM in the other groups of patients. Procalcitonin concentrations were only markedly increased in patients with septic shock (72 to 135 ng/mL, compared with approximately 1 ng/mL in the three other groups). The best predictive value for septic shock was found to be the measurements of NO2-/NO3- and procalcitonin concentrations., Conclusions: These observations showed that increase of proinflammatory cytokines was a consequence of inflammation, not of shock. In this study comparing various shock and infectious states, measurements of NO2-/NO3- concentration and procalcitonin concentration represented the most suitable tests for defining patients with septic shock.

The mechanisms leading to target cell killing by the human NK-like cell line YT2C2 have been studied. YT2C2 cells express CD28 antigen and kill B7-expressing targets by a CD28-mediated mechanism which is inhibited by anti-CD28 mAb (CD28.2). The lysis of B7-negative targets, which are also killed by YT2C2, is insensitive to CD28.2, but can be inhibited by cyclosporin A (CsA). CsA reduces degranulation in YT2C2 as measured by BLT-esterase release assays. A total suppression of B7-negative cell lysis was observed in the presence of EGTA, which blocks both degranulation and perforin polymerization, confirming that lysis of this type of target depends solely upon granule exocytosis. In contrast, an additional extracellular EGTA-resistant component in B7-positive target killing was evidenced. These results were consistently obtained with a panel of B7-positive and B7-negative targets, including a Jurkat subclone transfected to express B7 and its parental cell line. Ca2+-independent killing was completed during the first hour of the cytotoxicity assay, whereas EGTA-sensitive lysis increased throughout the whole incubation time. These two lytic mechanisms used by YT2C2 were found to induce two different modes of cell death. Extracellular Ca2+-dependent killing caused apoptotic death in both B7-positive and B7-negative targets, whereas the EGTA-resistant cytolytic pathway, observed exclusively with B7-positive targets, led to necrosis. CD28 triggering in YT2C2 induces, therefore, an additional mechanism of cell killing, independent of granule exocytosis, the nature of which remains to be identified.

Glucocorticoid-induced apoptosis in the murine interleukin-2-dependent T-cell line CTLL-2 and in freshly isolated thymocytes was studied. It was demonstrated here that in CTLL-2 cells, dexamethasone (methyl in position 16 alpha) was more efficient in inducing apoptosis than betamethasone (methyl in position 16 beta) or triamcinolone (hydroxyl in position 16). In contrast, no such difference between these three molecules was found in murine thymocytes. In addition, we showed that glucocorticoid-induced apoptosis on the two models was mediated through interaction with the glucocorticoid receptor and did not occur in the presence of inhibitors of transcription, translation or an endonuclease-inhibitor. Furthermore, in CTLL-2 cells, apoptosis took place in the presence of EGTA whereas it was prevented in murine thymocytes, thus indicating that calcium plays a different role in these two models. Finally, higher concentrations of interleukin-2 were needed to protect CTLL-2 cells against dexamethasone-induced apoptosis than that induced by betamethasone or triamcinolone. Thus, structural differences at position 16 of the steroid nucleus correlate with a different apoptosis-inducing activity by glucocorticoids which, however, is only evidenced in the calcium-independent CTLL-2 model.

The prognostic value of serum procalcitonin levels in 43 patients with acute melioidosis, an infection with a wide range of clinical manifestations, was assessed. In eight patients with mild localized infections, the median procalcitonin levels were 0.13 ng/mL (range, 0.02-0.46 ng/mL), which were similar to those in 19 healthy controls (median, 0.07 ng/mL; range, 0.03-0.15 ng/mL). In the patients with severe infections, the initial procalcitonin levels were significantly higher in the patients who died (median, 350 ng/mL; range, 63-3,538 ng/mL) than in the survivors (median, 19 ng/mL; range, 0.55-387 ng/mL) (P < .0001); 16 of 19 patients with procalcitonin levels of > 100 ng/mL died, compared with 2 of 16 patients with levels of < 100 ng/mL (relative risk, 6.7; 95% confidence interval, 1.8-25; P = .0001). In those patients who survived, the subsequent procalcitonin levels reflected closely the clinical course of their infection. The serum concentration of procalcitonin correlates well with the severity of Pseudomonas pseudomallei infection and is comparable with other acute-phase markers. However, this prognostic indicator and marker of continuing disease activity is not specific to melioidosis and could be applied to other severe infections.

In order to evaluate the relevance of in vitro methods for immunotoxicity assessment, the effects of pharmaceutical drugs on lymphoproliferative and cytotoxic functions of mouse splenocytes and human peripheral blood mononuclear cells (hPBMC) were studied. A comparison of sensitivity of immune cells from different origins to an in vitro exposure to different xenobiotics was performed using non-immunosuppressive (cimetidine and furosemide) and immunosuppressive (azathioprine (AZA), cyclosporine A (CSA), and dexamethasone (DEX)) drugs. For CSA, sensitivity of both rat and mouse splenocytes following in vitro exposure was compared to the one of hPBMC. Immune function tests included lymphoproliferative response to mitogenic lectins (concanavalin A (Con A) and phytohemagglutinin (PHA-P)) or to allogeneic cells (mixed leukocyte response (MLR)) and cytotoxicity assays (cytotoxic-T lymphocyte (CTL) and natural killer (NK) cell-mediated cytolysis). Additionally, to evaluate how well in vitro assays represent the in vivo situation, a comparison of the effect of cyclosporine A on the same immune function tests following in vivo or in vitro exposure was performed. The data obtained show numerous similarities in the effects observed following in vitro exposure of rodent or human cells to the drugs and a very similar sensitivity of rat and mouse cells to CSA in vitro. Discrepancies between human and rodent cells such as lymphoproliferative response to PHA-P following exposure to DEX or sensitivity of CTL-mediated cytolysis to CSA do exist. In vitro assays were very representative of the in vivo situation, both in the rat and in the mouse, following CSA exposure, except for NK cell activity in the rat. These data show the usefulness of in vitro systems for immunotoxicity assessment. They allow direct comparison of rodent and human systems, and could be representative, for drugs altering specifically the immune system like CSA does, of the in vivo situation.

As procalcitonin concentrations have been shown to be elevated in patients with septicemia and gram-negative infections in particular, we proceeded to investigate the effect of endotoxin, a product of gram-negative bacteria, on procalcitonin concentrations in normal human volunteers. Endotoxin from Escherichia coli 0113:H10:k, was injected i.v. at a dose of 4 mg/kg BW into these healthy volunteers. Blood samples were obtained before and 1, 2, 4, 6, 8, and 24 h after injection of the endotoxin. Each patient's cardiovascular and overall clinical status was monitored over this period. The patients developed chills and rigors, myalgia, and fever between 1-3 h. Tumor necrosis factor-alpha levels increased sharply at 1 h and peaked at 90 min, reaching the baseline concentration thereafter by 6 h. Interleukin-6 levels increased more gradually, peaking at 3 h and reaching the baseline concentration at 8 h. The procalcitonin concentration, which was undetectable (< 10 pg/mL) at 0, 1, and 2 h, was detectable at 4 h and peaked at 6 h, maintaining a plateau through 8 and 24 h (4 ng/mL). There was no elevation of calcitonin concentrations, which remained below 10 pg/mL, the lowest sensitivity of the assay. Procalcitonin was measured by a two-antibody immunoradiometric assay specific for this peptide, with no cross-reactivity with calcitonin, katacalcin, or calcitonin gene-related peptide. We conclude that endotoxin induces the release of procalcitonin systemically, that this increase is not associated with an increase in calcitonin, and that the increase in procalcitonin associated with septicemia in patients may be mediated through the effect of endotoxin described here. Whether procalcitonin participates in the mechanisms underlying inflammation remains to be investigated.

We recently reported the characterization of an antigen designated VRA09, identified by a monoclonal antibody and overexpressed on the surface of vincristine-resistant human ovarian carcinoma cells. In the present study, we analyze the distribution of this antigen in normal and tumor tissues. Its pattern of expression appears to differ from that described for other drug-resistance- and/or tumor-associated antigens. In normal tissues, the antigen has a restricted histological distribution and appears to be localized in mesoderm-derived tissues. In tumor tissues, VRA09 expression was mainly detected in serous ovarian tumors. Indeed, VRA09 is strongly expressed in papillary serous cystadenocarcinomas and their metastases, and more specifically in the basement membranes of serous tumors of borderline malignancy. In contrast, no immunostaining was observed in normal ovarian tissue or benign tumors. The detection of this antigen may help to identify serous ovarian tumors by distinguishing tumors of low malignancy from cystadenocarcinomas.

To characterize the membrane changes associated with cisplatin resistance, we raised monoclonal antibodies (MAbs) against a cisplatin-resistant subline (OV1/DDP) derived from a human ovarian carcinoma cell line (OV1/p). An MAb, designated OCP02, was selected for its particularly high affinity for the resistant cell line. It bound 3.1-fold higher to OV1/DDP cells than to OV1/p cells and recognized an M(r) 45K antigen. This antigen appeared to be present in several normal and tumorous tissues. Its distribution in normal tissues was mainly detected in tissues involved in secretory processes, suggesting that this antigen could be related to a transport mechanism in normal cells as well as in drug-resistant cells.

Traditional methods for toxicological assessment have indicated that the immune system is a frequent target of toxic insult following subchronic or chronic exposure to xenobiotics. However, most of the xenobiotics evaluated in standardized protocols were environmental chemicals and correlation with available clinical data was not possible. The purpose of this work was to evaluate the potential immunosuppressive effects of pharmaceutical drugs using a standardized protocol developed for immunotoxicological assessment. Two groups of pharmaceutical drugs were utilized: (a) drugs without known immunosuppressive effect linked to their utilization in human therapy (cimetidine, furosemide, indomethacin, amoxicillin, and procainamide) and (b) immunosuppressive drugs (azathioprine, cyclosporine A, and dexamethasone). Ex vivo tests using B6C3F1 mice were performed after a 28-day repeat dose regimen and assessed: (a) immunopathology, (b) cell-mediated immunity, (c) humoral immunity, and (d) nonspecific immunity. Host resistance to Listeria monocytogenes was also assessed following exposure to immunosuppressive drugs. The results showed that (a) immunopathology and immune function assays were necessary to detect all immunotoxicants and (b) the effects observed with nonimmunotoxic drugs were sometimes statistically significant but the biological significance of these effects is unlikely.

Antigenic challenge is known to influence brain catecholamine turnover, e.g. hypothalamic norepinephrine activity, but little is known about effects on the activity of serotoninergic neurons, i.e. the release of the neurotransmitter at nerve terminals. In the present study, we first investigated the changes of central serotonin (5-HT) metabolism in Fischer 344 male rats at 2, 3, 4 and 5 days following i.v. immunization with sheep red blood cell (SRBC). Major decreases in 5-HT levels were evident in the hypothalamus (Hy) and cortex (Cx) at a time which corresponded to the late phase of the production of specific antibodies to SRBC measured with a plaque-forming cell assay (PFC). A pretreatment with an immunosuppressive drug, cyclosporin A (CsA; 12.5 mg/kg by gavage for 7 days) prevented the decreases in cortical 5-HT levels. Concomitantly, a 2-fold increase in the basal 5-HT release at frontocortical nerve terminals was observed by using in vivo microdialysis in awake rats on Day 3 following SRBC inoculation. This effect was totally suppressed by CsA. Our data suggest that the decrease in brain 5-HT levels that occurs after antigen administration may reflect a specific short-lasting CsA-dependent-release of 5-HT at frontocortical nerve terminals at a time (Day 3 or 4) when the splenic immune response is maximal.

The effect of one intraperitoneal (i.p.) injection of ketanserin (K) (20mg/kg) on the levels of monoamines and neuropeptide Y (NPY) in some central and peripheral tissues was examined in normotensive (WKY) and spontaneously hypertensive (SHR) rats. In WKY rats, K induced a depletion of norepinephrine (NE) in the hypothalamus, the medulla, the atria and the caudal artery together with a decrease in dopamine level (DA) in the hypothalamus and in serotonin level (5-hydroxytryptamine, 5-HT) in the medulla. The reduction reached 25% to 35%. In SHR, NE and DA levels in the hypothalamus, and NPY levels in the caudal artery were lower than in WKY, while NE was higher in the adrenals. After treatment with K, NE, 5-HT, 5-HIAA (5-hydroxyindoleacetic acid) and NPY in the medulla were increased by about 30% to 50% while NE in the caudal artery was reduced by about the same value as in WKY. These results indicate that K-induced release of monoamines is not linked to NPY release. Moreover, monoamine and NPY sensitivity to K differ in SHR and WKY rats.

Human neuropeptide Y (hNPY) is a potent vasoconstrictor peptide of 36 aminoacid residues. We isolated hybridomas secreting four monoclonal antibodies directed against various epitopes of neuropeptide Y and studied their cross-reactivity with peptide YY (PYY) and the pancreatic polypeptide (PP), two peptides sharing sequence homologies with hNPY (respectively 70% and 50%). The antibody NPY02 is an IgG1 with a Ka of 5.5 x 10(10) liters/mol. It binds to the 11-24 region of NPY (IC50 = 2 x 10(-7)M), does not recognize PP but cross-reacts weakly with PYY. Antibodies NPY03 and NPY05 are IgG2 with respective Ka's of 6.7 x 10(9) and 2.5 x 10(10) liters/mol. They interact with a C-terminal epitope on NPY (NPY 27-34, IC50 = 2 x 10(-9) M for NPY03 and NPY 32-36, IC50 = 1 x 10(-9) M for NPY05). These two antibodies cross-react with PYY whereas only NPY05 binds PP. NPY05 is unable to bind the free acid form of neuropeptide Y. The 32-36 COOH free subpeptide is recognized 50,000 less efficiently by NPY05 than its amidated form. Antibody NPY04 is an IgG3 with a Ka of 3.8 x 10(8) liters/mol. It recognizes a N-terminal epitope between aminoacids 1 and 12 (IC50 = 2.5 x 10(-6) M). NPY04 interacts weakly with PYY but not detectably with PP. These results obtained with 4 different monoclonal antibodies demonstrate the presence of at least four epitopes on hNPY, two of them being continuous. These antibodies will be used to study the interaction of NPY with its receptor and to develop sensitive and specific assays for determination of NPY concentrations in biological fluids.

Sampling of blood or tissues in experimental animals need frequently a general anesthesia. Some anesthetic agents are known to modify plasma levels of catecholamines; moreover NPY is co-released with noradrenaline or adrenaline under certain conditions. Thus, we investigated the effect of three general anesthetic agents, chloral, chloralose and pentobarbital (pentobarbitone) on the levels of monoamines and NPY in rat hypothalamus, medulla, atria and adrenals. Monoamines were analysed by high pressure liquid chromatography using an electrochemical detection and NPY was measured by radioimmunoassay (immunoradiometric assay, IRMA). Chloralose or pentobarbitone did not affect monoamine tissue levels. In contrast, chloral increased 5-hydroxyindolacetic acid and to a lesser extent serotonin levels in the hypothalamus. NPY tissue levels did not change whatever the anesthetic used. These results must be taken into account in pharmacological or toxicological studies which need the use of a general anesthesia.

Triggering of the T-cell antigen receptor complex and some other surface molecules is coupled to the phosphodiesterase (phospholipase C)-mediated hydrolysis of membrane phosphoinositides, in particular, phosphatidylinositol-4,5-biphosphate (PiP2). PiP2 hydrolysis generates two products, inositol 1,4,5-triphosphate and diacylglycerol, which act in concert as second messengers to increase the free intracellular calcium concentration and activate protein kinase C, respectively, thereby stimulating subsequent events leading to cellular activation and proliferation. Transmembrane signalling in T-lymphocytes represents a potential target for designated drugs as well as immunotoxicants. Immunotoxic effects of polycyclic aromatic hydrocarbons are discussed in the view of interaction with transmembrane signalling in the T-lymphocyte.

The aim of this study was to compare carcinoembryonic antigen (CEA) and CA19-9 for the detection of local recurrences and distant metastases after complete resection of gastric carcinoma. At least one postoperative measurement of CEA and CA19-9 was performed in 54 patients. Among these, 32 had recurrence (59%) with a median follow-up of 618 days. Significantly higher sensitivity was observed for CA19-9 in comparison with CEA (68.8% vs 38.2% respectively), but specificity was slightly lower (81.8% vs 95.6% respectively). Increased CEA plasma level never preceded the diagnosis of recurrence while increasing CA19-9 preceded diagnosis in 13 patients (40.6%) from 1 to 22 months (median = 4.5 months). Increasing the normal range of CA19-9 to 80 UI/mL (2.5 x N) raises the specificity to 100% with acceptable sensitivity (53.1%). This study shows that CA19-9, compare with CEA, allows diagnosis of recurrence more often and earlier in the follow-up of resected gastric cancer.

Two monoclonal antibodies of the IgG2 subclass, designated A 01 and B 11, were prepared against two synthetic peptides corresponding to the COOH-terminal sequence of the human epidermal growth factor receptor (EGF-R), in order to detect EGF-R in a radioimmunometric assay and by immunohistochemistry. Characterization of these Mabs showed that they recognized two different eptiopes on the original peptides with Kd of 1.7 x 10(-8) M and 1.3 x 10(-7) M, respectively, without crossreaction. The A 431 antigen recognized by A 01 and B 11 had an apparent molecular weight of approximately 170,000 and was able to specifically link to EGF. Thus, A 01 and B 11 are directed against an antigenic site on the human EGF-R. With Western blot analysis and immunostaining, A 01 was shown to be EGF-R specific. In addition to the EGF-R, B 11 recognized two unidentified soluble proteins present in the cytoplasm of the SKBR-3 cell line but different from the c-erb B-2 oncoprotein expressed by these cells. Mabs A 01 and B 11 were used in an IRMA for the determination of EGF-R using the A 431 cell line as a source of EGF-R. Mab A 01 was also shown to be a useful tool for immunohistochemical detection of EGF-R.

In order to characterize the membrane changes related to Vinca alkaloid resistance, we raised monoclonal antibodies (mAbs) against a Vincristine resistant subline (OV1/VCR) derived from a human ovarian adenocarcinoma cell line (OV1/p). Among three monoclonal antibodies selected for a higher binding to OV1/VCR than to OV1/p cells, one designated OVR09, recognized a Mr 92,000 protein. This protein appears to be gradually overexpressed along the drug resistance establishment in vitro, and to decrease slowly in absence of drug. Further, mAb OVR09 showed a much higher binding to the vinblastine resistant epidermoid tumor cell line KbV1 than to its parental counterpart. The Mr 92,000 protein was also detected in various tumor cell lines and in an ovarian carcinoma surgical sample.

Due to their extreme specificity, monoclonal antibodies (Mabs) have been very useful for new analytical progresses in the field of tumor markers. For example, it has been possible to analyse only calcitonin, independently of its precursors in the patients with medullary thyroid carcinomas. These precursors have been estimated and found increased in the absence of calcitonin for many cancer patients, specially with small cell lung tumors. The assay of HCG and its subunits with a great specificity and sensitivity is an other interesting example of the analytical power of Mabs. The usefulness of the assay of beta subunit is now clear in the diagnostic and follow up of choriocarcinoma and perhaps in the screening of high risk pregnancies. An other example can be derivated of our works on neuropeptide Y (NPY). The assay of NPY is highly difficult, due to its great similarity with other neuropeptides (PYY and PP). Mabs have been used to solve these analytical problems. The assay of NPY is useful not only for patients with neuroendocrine tumors, but also experimentally in the rat hypothalamus for research in nutrition.

Neuropeptide Y (NPY) and cholecystokinin (CCK) are two peptides involved in opposite ways in the control of food intake. A possible interaction between NPY and CCK has not yet been well defined. Two CCK derivatives with agonistic and antagonistic properties were studied with regard to their effects on brain and plasma NPY levels. The CCK agonist decreased NPY levels in plasma and in the hypothalamus but not in the other brain areas assayed. The CCK antagonist reversed the agonist-induced decrease in both plasma and hypothalamus. These results suggest a negative relation between NPY and CCK peptides, which is not surprising given their opposite role in feeding regulation. The hypothalamus, a preferential site of this regulation, appears to be the brain area most involved in the NPY-CCK interaction. The plasma NPY level variations closely reflect the hypothalamic profile, suggesting a direct release of NPY by a mechanism that remains to be investigated.