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After a steady increase between the 1950s and the 1970s, laryngeal cancer mortality has been levelling off since the early 1980s in men from most western and southern European countries and since the early 1990s in central and eastern Europe. To update trends in laryngeal cancer mortality, we analyzed data provided by the World Health Organization over the last two decades for 34 European countries and the European Union (EU) as a whole. For major European countries, we also identified significant changes in trends between 1980 and 2012 using joinpoint regression analysis. Male mortality in the EU was approximately constant between 1980 and 1991 (annual percent change, APC=-0.5%) and declined by 3.3% per year in 1991-2012. EU age-standardized (world population) rates were 4.7/100,000 in 1990-91 and 2.5/100,000 in 2010-2011. Rates declined in most European countries, particularly over the last two decades. In 2010-11, the highest male rates were in Hungary, the Republic of Moldova, and Romania (over 6/100,000), and the lowest ones in Finland, Norway, Sweden, and Switzerland (below 1/100,000). In EU women, mortality was stable around 0.29/100,000 between 1980 and 1994 and slightly decreased thereafter (APC=-1.3%; 0.23/100,000 in 2000-01). We also considered male incidence trends for nine European countries or cancer registration areas. In most of them, declines were observed over recent decades. Laryngeal cancer mortality thus showed favourable trends over the last few decades in most Europe, following favourable changes in tobacco and, mostly for Mediterranean countries, alcohol consumption. What's new? The study quantifies the favourable trends of laryngeal cancer mortality in most European countries over the last few decades, following reduction in tobacco and, for Mediterranean countries, alcohol consumption.

Assessment of donor-specific alloreactive memory/effector T cell responses using an IFN-γ Elispot assay has been suggested to be a novel immune-monitoring tool for evaluating the cellular immune risk in renal transplantation. Here, we report the cross-validation data of the IFN-γ Elispot assay performed within different European laboratories taking part of the EU RISET consortium. For this purpose, development of a standard operating procedure (SOP), comparisons of lectures of IFN-γ plates assessing intra- and interlaboratory assay variability of allogeneic or peptide stimuli in both healthy and kidney transplant individuals have been the main objectives. We show that the use of a same SOP and count-settings of the Elispot bioreader allow low coefficient variation between laboratories. Frozen and shipped samples display slightly lower detectable IFN-γ frequencies than fresh samples. Importantly, a close correlation between different laboratories is obtained when measuring high frequencies of antigen-specific primed/memory T cell alloresponses. Interestingly, significant high donor-specific alloreactive T cell responses can be similarly detected among different laboratories in kidney transplant patients displaying histological patterns of acute T cell mediated rejection. In conclusion, assessment of circulating alloreactive memory/effector T cells using an INF-γ Elispot assay can be accurately achieved using the same SOP, Elispot bioreader and experienced technicians in kidney transplantation.

Summary Insulin-dependent (type 1) diabetes is a prototypic organ-specific autoimmune disease resulting from the selective destruction of insulin-secreting β cells within pancreatic islets of Langerhans by an immune-mediated inflammation involving autoreactive CD4+ and CD8+ T lymphocytes which infiltrate pancreatic islets. Current treatment is substitutive, i.e. chronic use of exogenous insulin which, in spite of significant advances, is still associated with major constraints (multiple daily injections, risks of hypoglycaemia) and lack of effectiveness over the long term in preventing severe degenerative complications. Finding a cure for autoimmune diabetes by establishing effective immune-based therapies is a real medical health challenge, as the disease incidence increases steadily in industrialized countries. As the disease affects mainly children and young adults, any candidate immune therapy must therefore be safe and avoid a sustained depression of immune responses with the attendant problems of recurrent infection and drug toxicity. Thus, inducing or restoring immune tolerance to target autoantigens, controlling the pathogenic response while preserving the host reactivity to exogenous/unrelated antigens, appears to be the ideal approach. Our objective is to review the major progress accomplished over the last 20 years towards that aim. In addition, we would like to present another interesting possibility to access new preventive strategies based on the ‘hygiene hypothesis’, which proposes a causal link between the increasing incidence of autoimmune diseases, including diabetes, and the decrease of the infectious burden. The underlying rationale is to identify microbial-derived compounds mediating the protective activity of infections which could be developed therapeutically.

Although therapeutic advancements have made Hodgkin's lymphoma (HL) a largely curable disease, trends in HL mortality have been variable across countries. To provide updated information on HL mortality in the Americas, overall and 20-44 years age-standardized (world population) mortality rates from HL were derived for the 12 Latin American countries providing valid data to the World Health Organization database and with more than two million of inhabitants. For comparative purpose, data for the United States and Canada were also presented. Trends in mortality over the 1997 to 2008 period are based on joinpoint regression analysis. Declines in HL mortality were registered in all Latin American countries except in Venezuela. In most recent years, HL mortality had fallen to about 0.3/100,000 men and 0.2/100,000 women in Argentina, Brazil, Chile, Colombia, Ecuador and Guatemala, that is, to values similar to North America. Despite some declines, rates remained high in Cuba (1/100,000 men and 0.7/100,000 women), Costa Rica and Mexico as well as in Venezuela (between 0.5 and 0.6/100,000 men and between 0.3 and 0.5/100,000 women). In young adults, trends were more favorable in all Latin American countries except Cuba, whose rates remained exceedingly high (0.8/100,000 men and 0.6/100,000 women). Thus, appreciable declines in HL mortality were observed in most Latin America over the last decade, and several major countries reached values comparable to North America. Substantial excess mortality was still observed in Cuba, Costa Rica, Mexico and Venezuela, calling for urgent interventions to improve HL management in these countries. What's new? Mortality from Hodgkin's lymphoma differs between developed and less-developed regions of the world, with mortality rates having declined markedly in the former but less so in the latter. In this study of Hodgkin's lymphoma mortality in 12 Latin American countries, selected based on population size and completeness of mortality data, appreciable declines were registered for the period 1997-2008 in most of the countries assessed, with rates reaching those observed in North America. However, substantial excesses remained in Cuba, Mexico, and Venezuela, calling for urgent intervention in these middle-income countries. Copyright

Cytokines play a role in alopecia areata. We used immunohistochemical and in situ hybridization studies to demonstrate the persistence of pro-inflammatory as well as apoptotic mechanisms in skin biopsies from patients with chronic alopecia areata. In situ hybridization allows the visualization of the distribution of immunocompetent cells in vivo. We studied skin biopsies from 11 untreated alopecia areata patients and two normal controls. In situ hybridization was performed on frozen sections using 35S-radio-labeled riboprobes, specific for IL-1beta, IL-2, IL-6, INFgamma, and granzyme B mRNA. Immunohistochemistry was carried out using an anti-IL-1beta monoclonal antibody, and a monoclonal antibody directed against the human Fas protein. We demonstrated the presence of cells labeled with IL-1beta, IL-6, INFgamma, and granzyme B antisense probes. Similarly, cells labeled with anti-IL-1beta were found in 10 of 11 cases. The labeled cells were located in the mononuclear peri- and intrafollicular infiltrate. Cells expressing granzyme B were found in close contact with the follicle. Fas positivity was demonstrated in four of four cases at the level of the cytoplasmic membrane of the hair follicle keratinocytes. These results, based on visualizing the labeled cells, demonstrate that pro-inflammatory cytokines are produced by the mononuclear cell infiltrate in close contact with follicles in alopecia areata. Furthermore, they demonstrate for the first time that apoptotic mechanisms involving granzyme B and Fas-Fas ligand pathways may play a major role in the persistence of chronic alopecia areata.

CXCL10, a gamma-interferon-induced chemokine seems to play a relevant role in lung involvement that occurs in systemic sclerosis (SSc). The objective of this study was to assess the serum level of CXCL10 in interstitial lung disease (ILD) associated with SSc.Serum level of CXCL10 was assayed in 23 healthy volunteers (60.0 years; 58.0-67.3) and 29 SSc patients (63.1 years; 60.1-69.4) by ELISA method. Pulmonary function tests (PFTs), lung CT-scan and echocardiogram were also performed in the patients. Serum levels from patients and healthy controls were compared and a comparison among SSc patients between those with and without ILD, as documented by lung CT-scan, was also performed.Median CXCL10 level from patients with SSc was significantly higher than that from healthy volunteers (110.0 pg/ml; 60.8-223.8 versus 52.0; 41.3-65.8; p0.001). Fifteen out of the 29 patients had ILD on lung CT-scan; the median CXCL10 level from SSc patients with ILD was significantly higher than that from SSc patients without ILD (210.0 pg/ml; 115.0-307.5 versus 76.0; 55.0-110.0; p=0.02).Our findings suggest that CXCL10 is specifically increased in the lung involvement of SSc and plays a role in scleroderma lung disease.

CD3-specific monoclonal antibodies were the first rodent monoclonals introduced in clinical practice in the mid 1980s as approved immunosuppressants to prevent and treat organ allograft rejection. Since then compelling evidence has been accumulated to suggest that in addition to their immunosuppressive properties, CD3-specific antibodies can also afford inducing immune tolerance especially in the context of ongoing immune responses. Thus, they are highly effective at restoring self-tolerance in overt autoimmunity, a capacity first demonstrated in the experimental setting, which was recently transferred to the clinic with success.

This study aimed at evaluating the screening of thyroïditis and coeliac disease, in a population of children and adolescents with type 1 diabetes, and at comparing the appearance of antibodies specific for these 2 diseases as a function of age.The study included 370 children and adolescents, 179 girls and 191 boys, aged 13.8 +/- 4.4 yr and with diabetes for 7.1 +/- 3.8 yr. Auto-immune thyroïditis was screened using antimicrosomal and antithyroglobulin auto-antibodies, at a mean rhythm of 3 tests per patient (1 every 2 yr), associated with dosages of TSH and FT4. Coeliac disease was screened using antigliadin (+/- antiendomysium) auto-antibodies, at a mean rhythm of 2 tests per patient, and was confirmed by duodenojejunal biopsy. Antithyroïd auto-antibodies were correlated with age following the "censured data analysis" type approach.Antithyroïd autoantibodies were found in 42 patients (11.4%), of whom 9 were treated for hypothyroïdism and 1 for Basedow disease, and coeliac disease autoantibodies were found in 9 patients (3.2% of tested patients). The cumulated frequency of antithyroïd auto-antibodies increased regularly with age and was significantly higher in girls, reaching 28% in girls and 12% in boys around 18 yr of age. As a consequence of this evolution, antithyroïd auto-antibodies were frequently found at the time of diagnosis of diabetes when it declared after 10 yr of age, while they often became positive secondarily when diabetes occurred before 10 yr of age. Coeliac disease specific auto-antibodies appeared much earlier and were found at the time of diagnosis of diabetes or at the first screening test.Antithyroïd autoantibodies are increasingly frequent with age in children with type 1 diabetes, and become very elevated in girls. The rhythm for screening should be adapted to this evolution of autoantibodies with age, which is very different between thyroïditis and coeliac disease.

Regulation of the immune response to self-antigens is a complex process that depends on maintaining self-tolerance while retaining the capacity to mount a robust immune response to foreign antigens. Autoreactive T cells specific for these autoantigens are present in most normal individuals but are kept under control by multiple diverse peripheral tolerance mechanisms. In the last few years, there has been a re-emergence of suppressor cells as among the most central of these regulatory mechanisms. These cells, which express CD4, CD25, and CD62L, develop in the thymus and survive in a CD28-dependent manner in the periphery to maintain the homeostatic equilibrium of immunity and tolerance. In this review, we will summarize studies of these regulatory cells as they relate to autoimmune diseases and more specifically to type 1 diabetes and attempt to address some of the many outstanding questions. Finally, evidence is provided to support the ability of anti-CD3 mAbs to stimulate the regulatory T cells and reset the rheostat of immune tolerance in an animal model of autoimmune diabetes, the NOD mouse.

Asthma is a major public health problem, with variable trends in several countries. We analysed mortality trends from asthma in Italy and Spain between 1980 and 1996. Overall asthma-related mortality at all ages increased between 1980 and 1987 in both sexes in Italy, from 16.6 in 1980-1981 to 29.0 in 1986-1987 per million males, and from 8.0 in 1980-1981 to 13.8 in 1986-1987 per million females, but decreased thereafter to reach 14.6 per million in males and 8.7 in females in 1996. The downward trends after 1987 were consistent in middle age and elderly population, but asthma mortality tended to rise in children and young adults over the last few years. In Spain, overall age-standardized mortality rates from asthma declined in men from 37.8 in 1980-1981 to 10.1 in 1996, and from 19.5 in 1980-1981 to 13.2 per million females in 1996. In women, the fall in mortality rates was smaller, and overall mortality was higher than in males since early 1990s. Trends of asthma mortality in Italy and Spain were favourable over the last decade.

Several studies reported data on the increased risk of erectile dysfunction (ED) in populations of diabetic men, but few presented data separately for Type 1 and Type 2 subjects. No comparison data for these diabetic subgroups are available with regard to risk factors for ED.Eligible for the study were men aged 20-69 years with a diagnosis of insulin-dependent (Type 1) or non-insulin-dependent (Type 2) diabetes who were observed on randomly selected days in 178 diabetes centres in Italy. Erectile dysfunction was defined as a failure to achieve and maintain an erection sufficient for satisfactory sexual performance.The study population consisted of 1383 Type 1 and 8373 Type 2 men. The prevalence of ED increased with age for both groups. After taking into account the effect of age Type 2 men (37/100 men) tend to report ED less frequently than Type 1 men (51/100 men). A significant positive relationship was reported between ED and poor metabolic control and smoking for both Type 1 and Type 2 men, whereas high body mass index (BMI) increased only the risk of ED in Type 1 cases.The study offers a quantitative estimate of the prevalence of ED and its main risk factors in Type 1 and Type 2 diabetic subgroups.

Objective: To investigate the relation between wine drinking and intake of selected indicator foods, which may vary in various populations. Design: Cross-sectional analysis of the comparison group of a case- control study. Setting: A network of teaching and general hospitals from six Italian areas. Subjects: 5642 control subjects (3261 females and 2381 males) aged 20‐74 y (median age 58 y), admitted for acute, non-neoplastic conditions unrelated to alcohol consumption. Participation rate was over 95%. Intervention: Trained interviewers collected information using a structured and validated questionnaire. The average intakes of selected food items were computed, together with the multivariate odds ratios (OR) of eating above the median of each food. Results: No appreciable difference in either sex for any food indicator considered (fruit, raw vegetables, cooked vegetables, salad and fish) was observed between abstainers, wine, and other alcoholic beverage drinkers. If anything, female wine drinkers reported less frequently high consumption of salad (ORa 0.8) and raw vegetables (ORa 0.8), both estimates being of borderline significance. Conclusions: In no instance did wine drinkers or mixed drinkers (who include a large proportion of wine drinkers, too) show an association with indicators of healthy diet. Sponsorship: Italian Association for Cancer Research, Milan, Italy. Descriptors: drinking patterns; wine intake; dietary habits; alcoholic beverages; alcohol intake; cross-sectional study; diet European Journal of Clinical Nutrition (2000) 54, 177‐179

The potential therapeutic benefits of CD3 monoclonal antibodies, such as OKT3, have been limited by their immunogenicity and their propensity to activate a severe cytokine release syndrome. This has constrained the clinical use of OKT3 to the treatment of acute rejection episodes of organ allografts.We have humanized a rat CD3 antibody and created a single amino acid substitution in position 297 of the IgG1 heavy chain to prevent glycosylation and, consequently, binding of the therapeutic antibody to Fc receptors and to complement. This antibody has been given as first line antirejection therapy in nine kidney transplant recipients with biopsy-proven acute rejection episodes.None of the patients demonstrated any antiglobulin response nor any significant cytokine release syndrome. Seven of the nine showed evidence of resolution of their rejection, although some patients experienced re-rejection.These findings suggest that CD3 antibodies can be engineered to lose their toxicity while retaining their potency as immunosuppressants. Nonactivating humanized CD3 monoclonal antibodies now merit further investigation in the management of transplant patients and in therapy of autoimmune diseases.

The first administration of CD3 monoclonal antibodies, such as anti-human CD3 (OKT3), induces a massive release of several cytokines, including tumor necrosis factor alpha (TNF-alpha), interferon (IFN)-gamma, interleukin (IL)-2, IL-3, IL-6, and granulocyte-macrophage colony-stimulating factor.Cytokine levels in patient's sera were measured by specific ELISA. In vitro cultures were performed using OKT3-stimulated peripheral blood mononuclear cells and/or whole blood from patients and normal controls.Here we describe that OKT3 administration to human renal allograft recipients also leads to a significant release of IL-10. Contrasting with most OKT3-induced cytokines, such as TNF-alpha whose release is transient, IL-10 levels show a more progressive increase, they peak only by 4-8 hr after the first OKT3 injection and persist longer. Thus, significant IL-10 levels are still detectable at the time of the second and the third OKT3 injection. Administration of corticosteroids, 1 hr before the first OKT3 injection, significantly reduced both TNF-alpha and IL-10 release. Experiments were performed to evaluate the source(s) of IL-10 and its (their) influence on the initial T-cell activation. When stimulated in culture with soluble OKT3, the production of IL-10 was dependent on the cooperation between T lymphocytes and monocytes. It is important that, as assessed through the use of a specific neutralizing antibody, the endogenous IL-10 produced in the co-culture system exerted a negative feed-back on the release of the other pro-inflammatory CD3-induced cytokines, which was reproducible.These results are supportive of a major role of IL-10 in the down-modulation of the OKT3-triggered T-cell activation cascade.

A hospital-based case-control study was conducted in the Heilongjiang Province of northeast China between May 1993 and May 1995, A total of 129 histologically confirmed brain cancer cases (73 gliomas and 56 meningiomas) and 258 matched controls were interviewed in 6 major hospitals to examine the influence of dietary factors in developing brain cancer, Information was obtained about frequency of consumption of 57 food items. Odds ratios (ORs) were obtained from conditional logistic regression, including allowance for socio-demographic factors, alcohol, tobacco and total energy intake. Consumption of fresh vegetables (OR = 0.29 for the highest quartile compared with the lowest one), and specifically of Chinese cabbage and onion, fruit (OR = 0.15), fresh fish (OR = 0.38) and poultry (OR = 0.16) was inversely related to the risk of developing brain cancer. A protective effect was also seen for vitamin E intake, calcium and, although non-significantly, beta-carotene and vitamin C. Risk of brain cancer increased with consumption of salted vegetables (OR = 2.54) and salted fish. (C) 1999 Wiley-Liss, Inc.

Considerable progress has been made in the knowledge of the epidemiology of gestational trophoblastic disease (GTD) in the last few years. There are two main and widely known points related to this disease: its geographical distribution and the different frequency in the various classes of age. GTD is more frequent in South-East Asia, India and Africa, and is rare in European and North American populations. For example, in the United States, the frequency of GTD was 108 per 100,000 pregnancies in the 1970's. In Europe, particularly in Italy, frequencies are lower. In northern Italy, the frequency of hydatidiform mole, in the period 1979-1982, was equal to 62 per 100,000 pregnancies, but in Indonesia and in China, the reported rates were 993 and 667 per 100,000 pregnancies respectively. GTD disease is more frequent in the extreme classes of age (under 20 and over 40 years) and the risk may be more than 100 times greater over 50 years. Besides these risk factors, the possible role of both genetic (familiarity, blood groups) and environmental factors (diet, cigarette smoking, etc.) has been investigated on the onset of GTD. This paper reviews the epidemiologic knowledge on GTD.

This study examined the association between maternal smoking before and during the first trimester of pregnancy and spontaneous abortion.We have been conducting a hospital-based case-control study on risk factors for spontaneous abortion in the greater Milan area. We collected information from 782 cases of spontaneous abortions and 1543 controls (women who delivered at term healthy infants).With respect to never smokers, the odds ratio (OR) were 0.7 (95%, confidence interval (CI), 0.5-1.0) for women who quit smoking and 1.3 (95% CI, 1.0-1.6) for those who continued during pregnancy. Women who smoked more than 10 cigarettes/day in the first trimester were at increased risk of miscarriage, with an OR of 1.4 (95% CI, 1.0-2.1). No relationship was evident between the number of cigarettes smoked before conception and the risk of abortion. Likewise, no association emerged between paternal smoking and miscarriage. Moreover, no significant interaction or modification effect was obtained when strata of age and other major characteristics were investigated.The risk of abortion associated with cigarette smoking during the first trimester of pregnancy was measurable and noticeable in this population, and accounted for 9% (95% CI, 6-13%) of all cases. The increased risk of spontaneous abortion in women smoking during pregnancy is a further reason to encourage pregnant women to quit.

The relationship between frequency of consumption of whole grain food and risk of selected neoplasms has been analysed using data from an integrated series of case-control studies conducted in northern Italy between 1983 and 1996. The overall dataset included the following incident, histologically confirmed neoplasms: oral cavity and pharynx 181, oesophagus 316, stomach 745, colon 828, rectum 498, liver 428, gallbladder 60, pancreas 362, larynx 242, breast 3,412, endometrium 750, ovary 971, prostate 127, bladder 431, kidney 190, thyroid 208, Hodgkin's disease 80, non-Hodgkin's lymphomas 200, multiple myelomas 120. Controls were 7,990 patients admitted to hospital for acute, non-neoplastic conditions, unrelated to long-term modifications in diet and neat likely to have been caused by tobacco or alcohol use. Odds ratios (OR) for subsequent scores (never/occasional/ frequent) of whole grain food consumption were derived after allowance for age, sex, education, smoking, alcohol intake and body mass index. High intake of whole grain foods consistently reduced risk of neoplasm at:all sites, except thyroid. The ORs for the highest category of consumption were 0.2-0.3 for upper digestive and respiratory tract neoplasms, 0.5 for stomach, colon and gallbladder, 0.7 for rectum, 0.6 for liver, 0.8 for pancreas and prostate, 0.9 for breast and endometrium, 0.6 for ovary, 0.4 for bladder and kidney, 1.3 for thyroid and around 0.5 for lymphomas and myeloma. The rests for trend in risks were significant for all neoplasms, except pancreas, endometrium, Hodgkin's disease and multiple myeloma. No significant heterogeneity was found across strata of age at diagnosis, sex, education, smoking habit, alcohol intake and body mass index. Thus, even in the absence of a univocal and satisfactory biological interpretation, the consistency of the patterns observed indicate that, in this population, higher frequency of whole grain food intake is an indicator of reduced risk of several neoplasms, (C) 1998 Wiley-Liss, Inc.

We conducted a randomized, double-blind, placebo-controlled, rising single-dose study to investigate the effects of recombinant human (rh) interleukin (IL) 10 in renal transplant patients who received OKT3 as induction therapy.Patients received 0.1 (n=6), 1 (n=6), or 10 microg/kg (n=3) rhIL-10 or placebo (n=6) intravenously 30 min before the first injection of 5 mg of OKT3. We monitored IL-10 serum levels, the effect of rhIL-10 on OKT3-induced cytokine production, clinical toxicity, and the incidence of immunization against OKT3.Serum IL-10 levels in the three experimental groups reached 0.8+/-0.2, 7.9+/-1.3, and 118.6+/-7.3 ng/ml (mean+/-SEM), respectively, 30 min after rhIL-10 injection. Peak plasma levels of tumor necrosis factor-alpha (TNF-alpha) were reduced from 2953+/-1599 pg/ml in patients injected with OKT3 and placebo to 447+/-155, 703+/-246, and 459+/-246 pg/ml in patients injected with 0.1, 1, and 10 microg/kg rhIL-10, respectively. Values for 24-hr TNF-alpha area under the curve decreased from 8988+/-3551 pg x hr/ml in control patients to 2284+/-494, 3950+/-955, and 2420+/-931 pg x hr/ml for the 0.1, 1, and 10 microg/kg rhIL-10 dose groups, respectively (P=0.045). There was also a trend toward reduced plasma levels of IL-2, IL-8, and interferon-gamma in rhIL-10-pretreated patients. Although none of the patients who received placebo or 0.1 or 1 microg/kg rhIL-10 developed an IgM antibody response directed against OKT3 during the first 10 days, this occurred in all three patients who received the highest rhIL-10 dose. In two of these patients, neutralization of OKT3 was associated with a reversible acute rejection episode.Pretreatment with doses of up to 1 microg/kg rhIL-10 is safe and reduces the release of TNF-alpha induced by OKT3. However, higher doses might promote early sensitization to OKT3.

Clusters of spontaneous abortion among video display terminal (VDT) users in North America and Canada in the late 1970s aroused suspicion about the potential risk of an association between VDT. exposure and pregnancy outcome. This case-control study considered the association between VDT use and the risk of miscarriage. Cases were 508 women admitted for spontaneous abortion to the Clinica Luigi Mangiagalli and a network of obstetric departments in the Milan area. Controls were 1,148 women who gave birth at term to healthy infants on randomly selected days at the same hospitals where cases were identified. No association emerged between VDT exposure and spontaneous abortion, the estimated odds ratio being 1.0 (95% CI: 0.8-1.2). This evidence agrees with studies conducted in different countries by various authors.

We analysed the relationship between coffee and alcohol intake, smoking and risk of multiple pregnancies using data from a case-control study on risk factors for multiple births conducted in Italy. Cases were 133 women who delivered multiple births not related to treatment for infertility (33 monozygotic and 100 dizygotic twins). Controls were 395 women admitted for normal delivery at the same clinic where cases had been identified. The odds ratios (OR) of multiple pregnancy were 1.5[95% confidence interval (CI) 0.8-2.8] and 2.0 (95% CI 1.0-3.7) for women drinking respectively one to two or three or more cups of coffee per day in comparison with non-coffee drinkers. Considering separately dizygotic and monozygotic pregnancies, the estimated OR were respectively for women drinking three or more cups of coffee, 1.7 and 3.1 for dizygotic and monozygotic pregnancies. The risk of multiple pregnancy tended to be higher in women drinkingor= 15 alcohol drinks per week: in comparison with tea-totallers the estimated OR for drinkor = 15 glasses per week were 2.3 and 2.6 respectively for dizygotic and monozygotic pregnancies. Heavy smokers (or = 10 cigarettes per day) were at increased risk of multiple pregnancy: in comparison with never smokers, the estimated OR for multiple pregnancy was 1.6 (95% CI 0.9-2.7). Considering separately the two groups of multiple pregnancy, the OR of dizygotic and monozygotic pregnancy were 1.4 (95% CI 0.8-2.5) and 2.4 (95% CI 0.9-6.1) for women smokingor = 10 cigarettes/day, but the trend in risk with number of cigarettes smoked per day and duration of the habit was not significant.

Recent knowledge into the pathophysiological mechanisms mediating the immune abnormalities characteristic of end-stage renal disease (ESRD) has focused on the dual activation versus deficiency state of immunocompetent cells. Despite major advances in renal replacement therapy, notably hemodialysis, no significant improvement in the immune status of uremic patients has been achieved. After a brief review of the role of T cells and B cells in the normal immune response, the functional and phenotypic T and B cell abnormalities observed in uremic patients are presented. Special emphasis is placed on our recent findings indicating that these abnormalities are observed at an early stage in the course of chronic renal failure, worsen with the progression of uremia, and are exacerbated by the dialysis procedure. The previous hypotheses that could reconcile the so-called Janus-faced behavior of T cells in uremia are updated in light of the recent findings obtained in the search of therapeutic strategies that could counteract the impaired responsiveness of patients with ESRD to vaccination against hepatitis B virus. Perspectives of research aimed at elucidating the respective role of T helper cell subpopulations (Th1 and Th2) could contribute to understanding of the mechanisms of the multifaceted process of uremia-related immune dysregulation and of the rationale for possible immunointervention strategies.

We recently described a case of anaphylaxis occurring at the time of retreatment with OKT3 of a renal allograft recipient in whom, for the first time, high anti-OKT3 IgE levels were documented. This led us to examine a large series of sera from 181 OKT3-treated patients to better define the frequency of IgE sensitization, its fine specificity (anti-isotypic and/or anti-idiotypic) and its relation to the appearance of IgG anti-OKT3 antibodies (Abs). Six patients out of the 181 assayed have developed anti-OKT3 IgE Abs as detected by ELISA. The earliest time of appearance of IgE anti-OKT3 Abs was 10 days after starting OKT3 (range, 10-25). The IgE response peaked by day 18 (range, 11-35) and had usually disappeared at 3 months after treatment. A more careful dissection of the fine specificity of the IgE response revealed that three of the four patients tested had developed an exclusive anti-idiotypic response. In the last patient, an anti-isotypic component was present since anti-OKT3 IgE Abs also reacted with control IgG2a, IgG2b, and IgG3 monoclonal antibodies. Importantly, anti-OKT3 IgE Abs were only detected in heavily sensitized patients also showing high titers of IgG specific Abs by ELISA (or = 1/1000) as well as "blocking" anti-OKT3 antibodies, as assessed by immunofluorescence. We conclude that (1) exposure to OKT3 may lead to specific IgE sensitization that, however, only appears in about 38% of the patients; (2) IgE Abs mostly appear in patients also showing high levels of conventional IgG anti-OKT3 Abs including the presence of "blocking" anti-idiotypic Abs, and (3) IgE Abs may be directed to both idiotypic and isotypic determinants of the monoclonal antibody.

Concomitant immune deficiency and activation of immuno-competent cells, together with a disequilibrium between inflammation-inducing cytokines and their specific natural inhibitors is the basis of our current understanding of immune system dysregulation in patients with chronic uraemia. These anomalies may even be accentuated by dialysis. Clinically, bacterial infections, viral hepatitis and amyloidosis all play important roles. Humoral factors include abnormal immunoglobulin response to specific antibodies and complement activation. The response of T lymphocytes, long sought as the origin of the immunodeficiency associated with chronic uraemia, is also significantly decreased in these patients. The decreased antibody responses to specific stimuli may be related to B cell dysfunction. Monocyte and polymorphonuclear cell reactions are also perturbed. A deficiency in natural killer cells is observed although the mechanisms involved and the consequences are still debated. The factors determining the anomalies leading to immune system dysregulation in chronic uraemia and dialysis and their relationship with the reduction in active nephron mass as well as their metabolic and/or endocrine consequences remain to be fully described. A better understanding of the mechanisms involved should lead to new strategies for immuno-intervention in patients with chronic renal failure and help in optimizing haemodialysis.

Hepatitis B virus (HBV) DNA sequences have been detected in leucocytes from HBV-infected individuals. The aim of this study was to assess the specificity of HBV for a special leucocyte subset in nine healthy chronic HBV carriers, nine HBs antigen-positive patients with chronic active hepatitis, 16 HBs antigen-negative haemophiliacs with HBc and/or HBs antibodies, and 10 patients with HBV-related systemic necrotizing vasculitis. HBV-DNA sequences were found by Southern blot hybridization in the leucocytes of 15 out of the 44 (34%) patients. The prevalence was not significantly different between the four groups. HBV-DNA was found in the CD4+ cells (9/11) as well as in the CD8+ cells (4/11), B cells (4/12) and monocytes (2/12). In conclusion, leucocytes, and particularly CD4+ lymphocytes, are frequently infected by HBV in patients with HBV serum markers.

The monoclonal antibody OKT3 is a potent immunosuppressant inducing both T-cell depletion and antigenic modulation of the CD3/T-cell receptor complex. CD3 + lymphocytes are cleared from the circulation within 1 hour of intravenous administration of OKT3. Periodic monitoring of remaining or reappearing CD3 + lymphocytes gauges the adequacy of OKT3 dosing and OKT3 consumption by anti-OKT3 antibodies. The maximum acceptable concentration of CD3 + lymphocytes for maintaining the effectiveness of OKT3 therapy remains controversial due to variable sensitivity of the methods used. Intravenously administered OKT3 has a half-life of approximately 18 hours and results in trough serum levels ranging from 500 to 1000 ng/ml in 3 to 4 days. The antibody response to OKT3 is oligoclonal, and both IgM and IgG antibodies to OKT3 may develop, showing restricted specificity; anti-isotypic and/or anti-idiotypic antibodies can be detected. The IgG anti-idiotypic antibodies are the ones that neutralize the therapeutic activity of OKT3, and their specific detection involves the use of an immunofluorescence method that tests their ability to block the binding of OKT3 to normal T cells. The production of OKT3 antibodies has been shown to be affected by concomitantly administered immunosuppressive agents. Studies have demonstrated that OKT3 can be successfully reused in allograft recipients who do not present anti-idiotypic blocking antibodies. The acute clinical syndrome regularly observed upon the first OKT3 injection is related to massive, although transient, release of several cytokines, which may be easily monitored with currently available radioimmunologic or immunoenzymatic tests.

Murine monoclonal antibodies specifically directed at the CD3 molecular complex are widely used in clinical transplantation. They are potent inhibitors of immune function and may be administered in association with conventional immunosuppressive drugs. However, their use has been complicated by the severe clinical syndrome that invariably follows the first injections of the antibody. We have shown that this reaction relates to transient T cell activation, with production of a cohort cytokines. This in vivo activating effect is unique to anti-CD3 MoAbs. Furthermore, this property is highly monocyte-dependent, engaging Fc receptors, and thus varies according to the isotype of the administered MoAb. Another anti-T cell MoAb such as the BMA031 directed towards monomorphic determinants of the T cell receptor (TCR) does not promote such massive in vivo activation.

Phenotypic analysis showed a significant increase of CD3+/CD57+ lymphocyte subpopulation in 88 kidney-transplanted patients when compared with non-transplanted patients. No clear correlation was found between the increase of this cell subpopulation and renal function, immunosuppressive regimen, and allogenic or viral stimuli, when each of these factors was considered. Conversely, there was a significant correlation between the proportion of CD3+/CD4+/CD57+ cells and the post-transplant follow-up period. Finally, functional study of CD3+/CD57+ cells suggested that they have suppressive properties, in vitro.

Murine monoclonal antibodies specifically directed at the CD3 molecular complex are widely used in clinical practice. They are potent inhibitors of immune function and may be administered in association to conventional immunosuppressants. The main purpose of this review is to summarize our present knowledge on the CD3 molecule, to discuss the available data on the clinical use of anti-CD3 MoAb (therapeutic effectiveness, mode of action and side effects) and the recent information derived from the use of anti-CD3 MoAb in experimental models that are opening new perspectives in the applications of anti-CD3 and will hopefully lead to the extension of its clinical use to settings different from transplantation.

OKT3 is considered to be effective in the prophylaxis and treatment of rejection. However, anti-OKT3 immunization is still a major side-effect. Only the IgG antiidiotypic component of the response is responsible for neutralization of OKT3 therapeutic activity by inhibiting OKT3's binding to T cells (i.e., blocking antibodies). It has recently been reported that successful OKT3 retreatment could be performed in patients showing circulating anti-OKT3 antibodies assessed by ELISA, which does not distinguish between blocking and nonblocking antibodies. The aim of this study was to investigate the role of these two types of anti-OKT3 antibodies for their capacity to interfere with effective OKT3 retreatment. Twelve cadaver renal allograft recipients who received OKT3 inducing therapy, were given a second 10 day-course of OKT3 for treatment of rejection. Circulating CD3+ cells were sequentially monitored. Anti-OKT3 antibodies were detected by using both conventional ELISA and an immunofluorescence inhibition test specifically detecting blocking antibodies. OKT3 and the patient's serum were incubated for 30 min at 4 degrees C, and 2 x 10(5) normal T cells were added (30 min at 4 degrees C). After washing, the cells were incubated with FITC goat antimouse antiserum. Fluorescent cells were counted using a FACS analyzer. In 10 patients, at the end of the 10-day second course, less than 10% circulating CD3+ cells were detected. None of these patients had detectable antibodies in the IF inhibition assay at the beginning of retreatment, irrespective of anti-OKT3 antibody titers detected by ELISA. In contrast, in two patients, OKT3 therapy was ineffective: more than 50% circulating CD3+ cells were detected and OKT3 treatment had to be interrupted soon after it was initiated. In both of them, blocking antibodies were detected by the IF-inhibition assay. These results suggest that specific detection of blocking antiidiotypic antibodies by the IF inhibition assay is a reliable parameter for predicting the feasibility of OKT3 retreatment, avoiding misuse of this expensive therapy.

This study analyzed the capacity of an anti-human tumour necrosis factor (TNF) monoclonal antibody, CB006 (murine IgG1, Celltech), to prevent the OKT3-induced acute clinical syndrome. Fourteen renal allograft recipients undergoing prophylactic OKT3 therapy were included. CB006 was administered as a single i.v. injection, 0.4 mg/kg (Group I, 7 patients) and 2 mg/kg (Group II, 7 patients), one hour prior to the first OKT3 administration. Nineteen consecutive patients that were part of a randomized multicenter trial constituted the historical control group. In all patients CB006 was perfectly well tolerated and significantly decreased the frequency of the common OKT3-associated acute symptoms. None of the CB006 pretreated patients showed severe life threatening symptoms (hypotension, respiratory distress or neurotoxicity), observed in 10 per cent of historical controls. At variance with controls, in CB006 treated patients gastrointestinal symptoms (vomiting, diarrhea) and pyrexia (body temperature greater than or equal to 39 degrees C) appeared in low frequency, were mild and short lasting, never promoting major prostration of the patients due to electrolytes and fluid loss. Importantly, the presence in some patients of these mild symptoms, correlated with detectable bioactive TNF in the circulation thus reflecting incomplete blockade by CB006 of OKT3-induced TNF. CB006 pharmacokinetics data further stressed the need for adequate dosage adaptation. CB006 did not affect the biological or clinical effectiveness of OKT3. None of the patients showed evidence of anti-CB006 xeno-sensitization.

The data presented extend to a larger series of 27 consecutive renal allograft recipients treated prophylactically with OKT3 our previous observation that the acute OKT3-induced clinical syndrome is related to massive release in the circulation of some cytokines, among which are tumor necrosis factor and interferon gamma. In addition, a pilot randomized study was set up including 12 consecutive patients receiving high-dose corticosteroid treatment (0.5 g solumedrol) either before or at the same time as the first OKT3 injection. Results confirm that when corticosteroids are given in sufficient amount and, importantly, 1 hr before the first OKT3 injection, they significantly decrease the release of both tumor necrosis factor and interferon gamma. In addition, the pretreatment with corticosteroids may totally abolish the IL-2 release induced by OKT3. Given the key role the massive although transient cytokine release plays in determining the OKT3-induced acute syndrome, these results provide the biological basis supporting a precise kinetics of administration of high-dose corticosteroids to better decrease the severity of the clinical reaction.