Daily peer review of abnormal cervical smears in the assessment of individual practice as an additional method of internal quality control.

OBJECTIVE: An important internal quality control system used in the Cancer Prevention and Research Institute cytopathology laboratory in Florence is the peer review procedure, based on the review of all abnormal cytological smears which routinely emerge. Peer review is an important training opportunity for all cytologists, especially for those with less experience. This article shows the results of the peer review procedure. METHODS: Of the 63 754 Papanicolaou (Pap) smears screened in 2011, 1086 were considered to be abnormal [at least atypical squamous cells of undetermined significance (ASC-US+)] on primary screening (selected by a single cytologist) and were subjected to the peer review procedure. The overall performance of the laboratory's cytologists was evaluated using a multiple rater analysis and the comparison of each cytologist with the final diagnosis. Further, the agreement was assessed by means of Cohen's kappa and weighted kappa statistics. RESULTS: In general, a moderate/substantial level of agreement between the ten cytologists and the final diagnoses was evident. Kappa values for each reader compared with the final diagnosis ranged from 0.54 to 0.69. The overall kappa value was 0.62 [95% confidence interval (CI), 0.58-0.66] and overall weighted kappa value was 0.76 (95% CI, 0.74-0.79). The category-specific agreement showed the lowest values for atypical squamous cells, cannot exclude high-grade squamous intraepithelial lesion (ASC-H). CONCLUSION: In summary, peer review represents an important internal quality control in the evaluation and improvement of inter-observer agreement and of the functioning of the laboratory as a whole. Multi-head microscope sessions may improve particularly the reproducibility of borderline diagnoses and, above all, can be an important training contribution for cytologists.

Therapeutic challenges for systemic sclerosis: facts and future targets.

Pulmonary arterial hypertension (PAH) is an important cause of death in systemic sclerosis (SSc), despite the improvement of therapies. An early diagnosis and the use of drugs interfering with the main pathogenic pathways of PAH is pivotal for the improvement of prognosis in primary PAH and PAH secondary to autoimmune rheumatic diseases, mainly SSc. Lately, new specific therapies have been developed targeting prostacyclin, endothelin, and nitric oxide pathways, the major pathogenic pathways leading to endothelial dysfunction in PAH. Epoprostenol improved life expectancy of patients with primary and secondary PAH, but its continuous intravenous administration requires experienced centers. More stable analogues of prostacyclin, administrated by intravenous (iloprost, treprostinil), subcutaneous, inhalatory (treprostinil, iloprost), and oral route (Beraprost) have shown efficacy in PAH. Bosentan, the first oral endothelin receptor antagonist (with affinity for endothelin A and B receptors) improves exercise function and survival in PAH, both primary and secondary to autoimmune rheumatic diseases. This is confirmed also for Sitaxsentan and Ambrisentan, selective A receptor antagonists. Because of its short half-life and systemic side effects, short-term NO inhalation is used only in short-term management of PAH in critically ill adults. Inhibitors of NO degradation, such as sildenafil, a phosphodiesterase (PDE) type 5 inhibitor, improved functional and hemodynamic parameters without significant side effects. Vardenafil and taladafil, longer-acting PDE inhibitors, also have vascular pulmonary selectivity. All these drugs may be used in combination, to maximize their clinical benefit not only in patients unresponsive to single drugs, but also potentially as initial therapy of PAH.

Clinical impact (cost-effectiveness) of qualifying atypical squamous cells of undeterminate significance (ASCUS) in cases favoring a reactive or dysplastic process.

The cost-effectiveness of qualifying ASCUS cases into two different subcategories, favoring a reactive (ASCUS-R) or dysplastic process (ASCUS-S), was evaluated at the Centro per lo Studio e la Prevenzione Oncologica of Florence in a prospective study. The study determined the positive predictive value (PPV) for histologically confirmed cervical intraepithelial neoplasia grade 2 (CIN2) or more (CIN>) severe lesion of the two ASCUS subgroups. ASCUS-S had a PPV (10.78%) comparable to low-grade squamous intraepithelial lesions (LSIL) (11.40%). For ASCUS-R cases, the recommendation of 6-mo repeat cytology prompting colposcopy in cases of persistent ASCUS or more severe cytology was also effective, as it selected a subgroup with a relatively high PPV (10.34%). The cost-effectiveness of a protocol based on ASCUS qualification was compared with two other possible options for nonqualified ASCUS cases: immediate colposcopy and colposcopy in persistent ASCUS at 6-mo repeat cytology. The detection rate of CIN2> was substantially higher using ASCUS qualification (35.9 vs 14.8 or 17.1). The cost per ASCUS subject was euro 24.99, 27.11, or 25.14 and that per CIN2> detected was euro 697, 1,831 or 1,470 for the three options, respectively. The evidence that ASCUS detection option implies a higher detection rate of CIN2> and subsequently a lower cost per CIN2> detection must be considered with caution and deserves confirmation by other comparative studies.

Clinical pharmacokinetics of factor VIII in patients with classic haemophilia.

Studies of Factor VIII pharmacokinetics in haemophiliacs can be classified into 2 groups depending on whether single-dose or multiple-dose Factor VIII curves are used. This review analyses information published so far in both these areas, with particular emphasis on the choice of appropriate models for pharmacokinetic analysis. Single-dose studies of Factor VIII kinetics have previously used a wide variety of methods for pharmacokinetic analysis (empirical methods of Factor VIII level prediction, graphical techniques for semilog analysis, 1-compartment and 2-compartment models). However, Factor VIII poses unique problems to the pharmacokineticist because decay curves can be either monophasic (monoexponential) or biphasic (biexponential) for unknown reasons, and because Factor VIII concentrations are generally subject to significant assay error. Problems of compartmental analysis that occurred in previous studies are highlighted, and a model-independent non-compartmental approach for analysing Factor VIII curves is proposed. To date, fewer data have been published on multiple-dose kinetics of Factor VIII. From a clinical point of view, repeated-dose regimens are most commonly required in patients undergoing surgery and in patients with severe bleeding. A fairly well defined 'therapeutic window' of optimal Factor VIII plasma concentrations has been identified, particularly in surgical patients. This fact has spurred research aimed at applying to haemophilia patients the pharmacokinetic dosing methods commonly used for therapeutic monitoring of drugs (e.g. Bayesian method for dosage individualization). A few papers have already been published in this field, and this review summarises problems encountered by previous investigators, and evaluates comparatively the pharmacokinetic methods used.

Pharmacological activity of some pyrrolo and pyrazolo pyrimidine derivatives.

4,7-Dihydro-4-ethyl-2-phenylpyrazolo-[1,5-a]pyrimidin-7-one (FPP028) is the prototype of a class of pyrazolo[1,5-a]pirimidine derivatives that has been shown to possess marked antiinflammatory and analgesic properties; 1,4-dihydro-1-ethyl-7-phenylpyrrolo[1,2-a]pyrimidin-4-one (FPP129) is a new compound belonging to a series of pyrrolo[1,2-a]pirimidine derivatives that has been synthetized in an attempt to reduce certain toxic effects observed with FPP028. To evaluate these two compounds more fully, in the present study we determined the LD50 of FPP129 in mice and assessed its activity in different experimental inflammation models. Ulcerogenic properties of FPP129 were evaluated by the test of stress-induced ulcer in rats. We also studied the in vitro and ex vivo effects of both FPP028 and FPP129 in a series of platelet-aggregation experiments using either arachidonate or collagen as aggregating agents. In the carraggeenan-induced paw edema, FPP129 exerted a marked anti-inflammatory activity with an ED50 of 22.2 mg/kg. FPP129 inhibited also the edema induced by concanavalin-A or Paf-acether. The in vitro anti-aggregatory activity of FPP028 was shown to be much less than that of indomethacin, thus confirming the hypothesis that the compound is a very weak inhibitor of cyclooxygenase. Ex vivo, both FPP028 and FPP129 were shown to exert no antiaggregatory effect in rabbits after administration of doses equal to the ED50. No ulcerogenic activity was found with FPP129, this result being consistent with previous observations on FPP028. Our data show tha FPP028 and FPP129 do not share a typical mechanism of action (cyclooxygenase inhibition) found with the majority of nonsteroidal antiinflammatory drugs. Nonetheless, the antiinflammatory properties of these compounds, together with their lack of ulcerogenic activity, indicate that both FPP028 and FPP129 are potentially interesting for future therapeutic use.

Effect of oral treatment with pantethine on platelet and plasma phospholipids in IIa hyperlipoproteinemia.

In a single-blind, crossover, completely randomized study, the effects of oral treatment with pantethine or placebo on fatty acid composition of plasma and platelet phospholipids were investigated in 10 IIa hyperlipoproteinemic patients. A significant decrease of total cholesterol and total phospholipids was observed both in plasma and in platelets after a twenty-eight-day treatment. In plasma, pantethine induced a decrease of the ratio sphingomyelin/phosphatidylcholine. Moreover, a relative increase of n3-polyunsaturated fatty acids both in plasma and in platelet phospholipids and a decrease of arachidonic acid in plasma phospholipids were observed. These results indicate that pantethine can affect plasma and platelet lipid composition with possibly favorable influences on the determinants of cell membrane fluidity.

Age related changes in platelet lipid composition.

Platelet lipid composition was investigated in 52 healthy subjects aged 20 to 68 years with similar dietary habits and living in a narrow geographic area in order to search possible changes referrable to aging. No significant variations were observed when platelet cholesterol, total phospholipids and different phospholipid fractions were considered, whereas cholesterol/phospholipid (C/PL) molar ratio significantly increased with aging (p less than 0.01). Moreover, a significant increase in 16:0 + 16:1 fatty acids was found in phosphatidylcholine (PC) and in sphingomyelin (SP) (r = 0.62, p less than 0.001 and r = 0.30, p less than 0.05 respectively) and a decrease in 18:2 n6 in the phospholipid fractions considered (at least p less than 0.05). These results indicate that modifications in platelet lipid composition occur with aging and that they could affect platelet functions so playing a role in the onset of atherosclerosis and in thrombotic phenomena occurring with increasing frequency in the elderly.

Platelet function and coagulation studies in patients with mitral valve prolapse.

Twenty-eight consecutive patients with mitral valve prolapse (MVP), seven of whom had previous cerebrovascular disorders (CVD), were studied for platelet function and coagulation tests. While platelet function tests were found to be normal with the exception of platelet aggregation rate (PAR), there was a significant rise of factors VIII vWF:Ag (Von Willebrand) and (FPA) fibrinopeptide A. Six cases had high levels of both these factors, suggesting the existence of a particular subset of patients with MVP, with a higher risk of thromboembolic episodes, although only three out of seven patients with previous CVD had either FPA or VIII vWF:Ag levels. The broad spectrum of subjects with MVP probably explains the different results obtained when studying platelet function and coagulation factors. Therefore, larger population studies and prolonged follow-up of cases with either coagulation abnormalities similar to the ones found in the present report and/or altered platelet function tests are suggested to discover if it is possible to detect patients with a potential for thromboembolism.

Increased thromboxane A2 generation and altered membrane fatty acid composition in platelets from patients with active angina pectoris.

Lipid composition of platelet membranes and thromboxane A2 (TxA2) generation by platelets were investigated in eighty-seven anginal patients (forty-two with resting angina in active phase and forty-five with effort stable angina or rest angina in inactive phase) and in forty-five clinically healthy subjects of similar age. All subjects were on the same dietary regimen and the adherence to diet was checked by analysis of red blood cell lipids. Platelets from active angina patients produced more TxA2 than platelets from both inactive patients and controls (p less than 0.001). Moreover patients with active angina had higher arachidonic acid (AA, p less than 0.001) and lower eicosapentaenoic (EPA) and docosahexaenoic acid (DHA) levels in phosphatidylcholine (PC, p less than 0.001), than inactive patients and controls. AA and EPA changes in membrane PC significantly correlated with TxA2 production (p less than 0.001) but not with coronary pathoanatomy. Plasma lipids, content of cholesterol, total phospholipids (and their saturated and unsaturated fatty acids) and the different phospholipid fractions in platelet membrane were not different in the three groups. Present results indicate that in platelets from anginal patients phospholipid fatty acid composition is at least in part independent of plasma composition and that in active angina there are modifications leading to increased TxA2 formation and possibly contributing to the occurrence of ischemic attacks.

A microcomputer program for individualizing factor VIII dosage in hemophilia patients undergoing major surgery.

A pharmacokinetic program that allows individualization of Factor VIII dosage regiments in hemophilia patients undergoing major surgery is described. The program, which is designed for the IBM PC microcomputer and compatible machines, is based upon the one-compartment open model with instantaneous input. In the framework of such a pharmacokinetic model, it is assumed that the elimination of Factor VIII is faster during the early post-operative period and that it decreases progressively over the following days. Since Factor VIII half-life is dependent on the time elapsed since the operation (short half-life values during the early post-operative period, longer half-life values thereafter), the pharmacokinetic model is a nonlinear one. A first-order 'variation' rate constant is used to describe the prolongation of Factor VIII half-life from the initial value immediately after surgery to the final value achieved several days later. Individualized estimation of the patient's kinetic parameters (initial half-life, 'variation' rate constant and volume of distribution) is performed through the Bayesian method. Therefore, for such estimation the program exploits the Factor VIII plasma levels measured in the individual patient as well as the population pharmacokinetic data of Factor VIII. After estimating the individual's Bayesian parameters, the program predicts the dosage regimen that will elicit the desired time-course of Factor VIII plasma levels. If requested, the program is able to calculate the least-squares estimates for the parameters of the pharmacokinetic model and dosage prediction can also be made on the basis of such estimates. The least-squares estimates are useful for calculating population pharmacokinetic parameters according to the Standard Two-Stage method. Some examples of clinical use of the program are presented.

Pharmacokinetics of a new heat-treated concentrate of factor VIII estimated by model-independent methods.

We evaluated the pharmacokinetic properties of Kryobulin TIM3, a new heat-treated Factor VIII concentrate which has recently become available in Europe. Twelve patients with classic hemophilia were studied. In each patient, Factor VIII was given as a single dose (ranging from 11.6 to 30.3 units/kg) after which eight serial blood samples were drawn to characterize the disappearance of Factor VIII from the plasma. Model-independent (noncompartmental) methods were used for pharmacokinetic analysis. The following pharmacokinetic parameters of Factor VIII (mean +/- SD) were estimated: clearance = 3.83 +/- 0.99 ml/h/kg; mean residence time = 15.9 +/- 4.5 h; volume of distribution at steady state = 55.6 +/- 9.3 ml/kg; in-vivo recovery = 129 +/- 29%. The pharmacokinetic parameters of Kryobulin TIM3 obtained in our study are very similar to those previously reported for the untreated concentrate. Thus, our findings suggest that the dosing guidelines previously available for the untreated concentrate need not be revised when using the treated product.

Spontaneous and cold pressor test-induced prostaglandin biosynthesis by human heart.

To investigate prostaglandin biosynthesis by the heart, 21 patients undergoing cardiac catheterization and coronary angiography for congenital or acquired heart diseases other than coronary artery disease were investigated. Prostacyclin (as 6-keto-PGF1 alpha), PGE2, PGF2 alpha and TxA2 (as TxB2) were measured by specific radioimmunoassay in blood from coronary sinus, aorta, and a peripheral vein under resting conditions and following cold pressor test (CPT). PGF2 alpha was always found undetectable. In resting conditions, no significant differences in plasma 6-keto-PGF1 alpha, PGE2, or TxB2 concentrations were found among coronary sinus, aorta, and peripheral venous blood and no transcardiac gradient existed (mean: +0.4 +/- 1.2 pg/ml for 6-keto-PGF1 alpha, +0.1 +/- 0.6 pg/ml for PGE2, and -0.4 +/- 9.9 pg/ml for TxB2). CPT was able to induce a significant increase in 6-keto-PGF1 alpha and PGE2 concentration in blood from the different sampling sites and a significant transcardiac gradient was found following CPT (+11.6 +/- 7.4 pg/ml for 6-keto-PGF1 alpha (p less than 0.01) and +5.2 +/- 3.6 pg/ml for PGE2 (p less than 0.001). TxB2 levels significantly increased in peripheral venous blood (from 18.3 +/- 6.2 to 29.2 +/- 20.3 pg/ml, p less than 0.05), but they did not increase either in coronary sinus (from 21.9 +/- 9.7 to 22.9 +/- 9.8 pg/ml) or in aorta (from 22.3 +/- 4.7 to 19.1 +/- 6.5 pg/ml). Present results indicate that a cardiocoronary prostacyclin and PGE2 synthesis is inappreciable under resting conditions but it becomes remarkable following sympathetic stimulation. On the contrary, no TxA2 cardiocoronary biosynthesis seems to occur in patients free from coronary artery disease.

Decreased number of PGD2 binding sites on platelets from patients with type IIa hyperlipoproteinemia.

Platelets from patients with familial hypercholesterolemia (type IIa hyperlipoproteinemia), a condition associated with a high prevalence of atherosclerosis and its ischemic complications, are claimed to be hyperresponsive to aggregating stimuli. We investigated the platelet responsiveness to and the binding of PGD2, a potent endogenous inhibitor of platelet aggregation via stimulation of adenylate cyclase, in a group of 7 patients affected by IIa hyperlipoproteinemia (IIa HLP) and in a control group of 10 healthy subjects. Inhibition by PGD2 of ADP-induced platelet aggregation was significantly lower in IIa HLP patients than in controls. The number of binding sites for PGD2 of platelets from IIa HLP patients was significantly reduced in comparison with that from controls (93 +/- 19 and 232 +/- 23 receptors/platelet, respectively), whereas the affinity for PGD2 was comparable to that of controls (Kd = 68.8 +/- 19.8 nM in patients and 66.1 +/- 15.9 nM in controls). The reduced number of platelet PGD2 binding sites in IIa HLP patients may account for the impaired sensitivity to PGD2 shown in vitro by platelets and may contribute to the increased tendency to thrombotic manifestations observed in IIa HLP.

A calculator program for clinical application of the Bayesian method of predicting plasma drug levels.

A pharmacokinetic program that allows individualization of drug dosage regimens through the Bayesian method is described. The program, which is designed for the Hewlett-Packard HP-41 CV calculator, is based upon the one-compartment open model with either instantaneous or zero-order absorption. Individualized estimation of the patient's kinetic parameters (clearance and volume of distribution) is performed by analyzing the plasma levels measured in the patient as well as considering the population data of the drug. After estimating the individual kinetic parameters by the Bayesian method, the program predicts the dosage regimen that will elicit the desired peak and trough plasma levels at steady state. For comparison purposes, the least-squares estimates for clearance and volume of distribution are calculated, and dosage prediction can also be made on the basis of the least-squares estimates. The least-squares estimates can be used to calculate population pharmacokinetic parameters according to the Standard Two-Stage method. Several examples of clinical use of the program are presented. The examples refer to patients with classic hemophilia who were treated with Factor VIII concentrates. In these patients, the Bayesian kinetic parameters of Factor VIII have been estimated through the calculator program. The Bayesian parameter estimates generated by the HP-41 have been compared with those determined by a Bayesian program (ADVISE) designed for microcomputers.

Kinetic evaluation of four Factor VIII concentrates by model-independent methods.

Single-dose kinetics of 4 Factor VIII concentrates (Kryobulin, Hemofil T, Koate, cryoprecipitate) were studied in 41 patients with haemophilia-A. Model-independent methods were adopted for calculating the kinetic parameters (area under the curve, clearance, area under the moment curve, mean residence time, volume of distribution at steady-state). No substantial difference was observed in the kinetic characteristics of the 4 Factor VIII concentrates. A considerable interindividual variability of the calculated kinetic parameters was demonstrated for all concentrates. Our findings support the need to individualize Factor VIII dosage.

Changes in fatty acid composition of the single platelet phospholipids induced by pantethine treatment.

In a single-blind cross-over study the effect of oral treatment with pantethine on plasma and platelet lipid composition was evaluated in 20 patients with dyslipidaemia (7 IIa, 7 IIb and 6 Iv type). In plasma significant decreases of total cholesterol and triglycerides with increase of high density lipoprotein-cholesterol were observed. In platelets pantethine treatment significantly reduced phospholipid and cholesterol content. In addition gas-chromatographic analysis showed a reduction of saturated and monounsaturated and a relative increase of polyunsaturated fatty acid content of platelet phospholipids. A selective relative increase was observed of some n-3 polyunsaturated fatty acids like eicosapentaenoic and docosahexaenoic acid whereas arachidonic acid decreased. The present study indicates a favourable influence of pantethine not only on plasma but also on platelet lipids which could be of value in delaying the development of atherosclerosis in dyslipidaemic patients.

Cryoprecipitate and Factor VIII commercial concentrates: in vitro characteristics and in vivo compartmental analysis.

In vitro and in vivo characteristics of cryoprecipitates and three commercial factor VIII concentrates (Kryobulin, Hemofil and Koate) were comparatively studied. Factor VIII:C/VIIIR:Ag ratio was very low in all commercial concentrates without differences among them. Conversely, the decrease of factor VIIIR:WFRCof was proportional to the degree of purity. Factor VIII:C/VIIIR:WFRCof ratio was shown to be a reliable index of factor VIII complex denaturation. Crossed electroimmunoassay showed a faster migration of factor VIIIR:Ag only in commercial concentrates. Using a two-compartmental open model that accounts for endogenous synthesis of factor VIII:C, single-dose kinetics of factor VIII:C were studied in 23 patients with classic hemophilia. Good agreement between measured and fitted values of factor VIII:C plasma concentration was observed. beta half-life was shorter in high purity concentrates and longer in intermediate purity concentrates and cryoprecipitates.

A calculator program for individualizing factor VIII dosage.

We developed a calculator program to individualize Factor VIII dosage on the basis of the concentration-time data obtained after a single test-dose. The program is designed for the Hewlett-Packard 41-CV calculator. The calculation procedure is the 1977 version of the Sawchuk and Zaske method. The program performs model parameter estimation through a nonlinear iterative least-squares technique (the modified Gauss-Newton method). We tested our program in three patients with classic hemophilia who required multiple-dose treatment with Factor VIII. In each patient, individual kinetic parameters were estimated from the serial plasma levels measured after the test-dose. The predicted concentration-time curve resulting from all the administered doses was calculated based on the estimated kinetic parameters. Good agreement between predicted and measured levels during the multiple-dose regimen was observed in all patients.