Development of Three Molecular Diagnostic Tools for the Identification of the False Codling Moth (Lepidoptera: Tortricidae).

Three molecular protocols using qPCR TaqMan probe, SYBR Green, and loop-mediated isothermal amplification (LAMP) methods were set up for the identification of larvae and adults of an African invasive moth, Thaumatotibia leucotreta (Meyrick, 1913) (Lepidoptera: Tortricidae). The DNA extracts from larval and adult samples of T. leucotreta were perfectly amplified with an average Ct value of 19.47 ± 2.63. All assays were demonstrated to be inclusive for T. leucotreta and exclusive for the nontarget species tested; the absence of false positives for nontarget species showed a 100% of diagnostic specificity and diagnostic sensitivity for all assays. With the SYBR Green protocol, the Cq values were only considered for values less than 22 (cutoff value) to prevent false-positive results caused by the late amplification of nonspecific amplicons. The limit of detection (LoD) for the qPCR probe protocol was equal to 0.02 pg/µl while a value equal to 0.128 pg/µl for the qPCR SYBR Green assay and LAMP method were established, respectively. The intrarun variabilities of reproducibility and repeatability in all the assays evaluated as CV%, ranged between 0.21 and 6.14, and between 0.33 and 9.52, respectively; the LAMP values were slightly higher than other assays, indicating a very low interrun variability. In order for an operator to choose the most desirable method, several parameters were considered and discussed. For future development of these assays, it is possible to hypothesize the setup of a diagnostic kit including all the three methods combined, to empower the test reliability and robustness.

Prostate cancer: from the pathophysiologic implications of some genetic risk factors to translation in personalized cancer treatments.

Several pathologies affect human prostate, such as prostate cancer (PC), which is the most common non-skin malignant cancer in Western male populations. A complex interaction between genetic and environmental factors (i.e. infectious agents, dietary carcinogens) and hormonal imbalances has been reported to have a fundamental role in PC pathophysiology by evoking chronic inflammation. Thus, chronic inflammation drives prostate carcinogenesis and neoplastic progression. No adequate biomarkers exist until now to guide PC prognosis and treatment. Accordingly, the research has particularly focused its attention on genetic variants in genes, codifying molecules of signaling innate immune/inflammatory and steroid metabolism pathways, which are able to modify the PC genetic susceptibility and clinical disease outcome. Single-nucleotide polymorphisms (SNPs) may operate in combination to create a 'risk profile'. Combinations of several inflammatory and sex steroid hormone pathway SNPs are found in PC patients. Thus, their combinations might be used as promising biomarkers in a pre- and post-treatment clinical PC setting. Indeed, their identification may hold promise for the realization of a personalized PC medicine. Many of these aspects are summarized in this report through the elucidation of literature data and the results of our studies.

Pathological implications of Th1/Th2 cytokine genetic variants in Behçet's disease: Data from a pilot study in a Sicilian population.

Cytokines act as pleiotropic polypeptides able to regulate inflammatory/immune responses and to provide important signals in physiological and pathological processes. Several cytokines (Th1, Th2, and Th17) seem to be involved in the pathophysiology of Behçet's disease, a chronic immune-mediated disease characterized by oral and genital lesions and ocular inflammation. Its individual susceptibility seems to be modulated by genetic variants in genes codifying these cytokines. Th1 and Th17 seem to be involved in the disease's active phases, and Th2 seems to affect the development or severity of the disease; however, contrasting data are reported. In this study, some genetic variants of the Th1/Th2 cytokine genes were investigated in Sicilian patients and age- and gender-matched controls. Three very significant associations with Behçet's disease were detected, and combined genotypes associated with increased disease risk were identified. Results obtained point to the key role of Th1/Th2 cytokine genetic variants in disease susceptibility.

Analysis of IL-6, IL-10 and IL-17 genetic polymorphisms as risk factors for sepsis development in burned patients.

Infection risk, sepsis and mortality after severe burn are primarily determined by patient age, burn size, and depth. Whether genetic differences contribute to otherwise unexpected variability in outcomes is unknown. We sought to determine whether there was an association between IL-6, IL-10 and IL-17 polymorphisms with cytokine production and development of sepsis. We evaluated 71 patients with burns ≥15% TBSA and 109 healthy subjects. The genotypes of IL-6 (-174C/G), IL-10 (-819C/T and -1082A/G) and IL-17 (7488T/C) polymorphisms were identified applying polymerase chain reaction protocols. The cytokine levels in serum were determined with enzyme-linked immunoabsorbent assays. Our results demonstrated no significant differences in the genotype frequencies studied between burn patients and healthy subjects. No significant associations were found among IL-6 and IL-17F genotypes and the related cytokine serum levels. Only IL-10 promoter -1082GG genotype was related to an increased IL-10 production in burned patients. In addition, septic subjects bearing -1082G/G genotype have shown the highest and non-septic bearing -1082A/* genotypes the lowest IL-10 serum levels. All together these data seem to indicate that genetically determined individual difference in IL-10 production might influence the susceptibility to septic complications in burned patients and suggest that these markers might be useful in burned patient management.

B cells compartment in centenarian offspring and old people.

Immunosenescence is considered a major contributory factor to the increased frequency of morbidity and mortality among elderly. On the other hand centenarians are considered the best example of successful ageing. To gain insight into mechanisms of immunosenescence and its clinical relevance, a possible model is represented by centenarians and/or their offspring. Nowadays centenarians are not more a curiosity, but in Europe are 1/8000 inhabitants and it has been demonstrated that the centenarian offspring, who are typically in their 70s and 80s, have a survival advantage when compared with age-matched controls whose parents died at an average life expectancy. Then again, studies on immunosenescence focus mainly on T cell impairment, although B cells are also affected. So, in the present preliminary report, we have studied B cell compartment in two classes of individuals, old people and centenarian offspring. B cell compartment was analysed using IgD and CD27 antibodies which characterize naïve B cells (IgD(+) CD27(-)), memory unswitched B cells (IgD(+)CD27(+)), memory switched B cells(IgD(-)CD27(+)) and double negative B cells (DN) (IgD(-)CD27(-)), i.e. exhausted memory cells. As expected, in both cohorts we observed a decreased B cell count. However, in centenarian offspring, naïve B cells are more abundant whereas exhausted memory cells (DN B cells, IgD(-)CD27(-)) do not show the increase that we have previously demonstrated in healthy elderly donors. These data are similar to that found in previously experiments on young subjects. So, our preliminary results show that centenarian offspring do not have the typical trend of memory/naive B cell subsets observed in elderly people and this is in agreement with the higher levels of IgM in the serum of centenarian offspring in comparison with data obtained in age-matched controls. This reservoir of naive B cell might be one of the causes that make centenarian offspring able to keep fighting off new infections, hence prolonging their life. So, B cell subset changes could represent a hallmark of successful or unsuccessful ageing and could be used as a biomarker of human life span, potentially useful for the evaluation of anti-ageing treatment.

Age-related inflammation: the contribution of different organs, tissues and systems. How to face it for therapeutic approaches.

A typical feature of ageing is a chronic, low-grade inflammation characterized by a general increase in the production of pro-inflammatory cytokines and inflammatory markers ("inflamm-ageing"). This status may slowly damage one or several organs, especially when unfavorable genetic polymorphisms and epigenetic alterations are concomitant, leading to an increased risk of frailty together with the onset of age-related chronic diseases. The contribution of different tissues (adipose tissue, muscle), organs (brain, liver), immune system and ecosystems (gut microbiota) to age-related inflammation ("inflamm-ageing") will be discussed in this review in the context of its onset/progression leading to site-restricted and systemic effects. Moreover, some of the possible strategies and therapies to counteract the different sources of molecular mediators which lead to the age-related inflammatory phenotype will be presented.

Immune-inflammatory responses and oxidative stress in Alzheimer's disease: therapeutic implications.

Alzheimer's disease (AD) is a heterogeneous and progressive neurodegenerative disease which in Western society mainly accounts for clinical dementia. AD has been linked to inflammation and oxidative stress. Neuro-pathological hallmarks are senile plaques, resulting from the accumulation of several proteins and an inflammatory reaction around deposits of amyloid, a fibrillar protein, Abeta, product of cleavage of a much larger protein, the beta-amyloid precursor protein (APP) and neurofibrillary tangles. Inflammation clearly occurs in pathologically vulnerable regions of AD and several inflammatory factors influencing AD development, i.e. environmental factors (pro-inflammatory phenotype) and/or genetic factors (pro-inflammatory genotype) have been described. Irrespective of the source and mechanisms that lead to the generation of reactive oxygen species, mammalian cells have developed highly regulated inducible defence systems, whose cytoprotective functions are essential in terms of cell survival. When appropriately activated, each one of these systems has the possibility to restore cellular homeostasis and rebalance redox equilibrium. Increasing evidence, support the notion that reduction of cellular expression and activity of antioxidant proteins and consequent augment of oxidative stress are fundamental causes for ageing processes and neurodegenerative diseases., including AD. The better understanding of different molecular and cellular inflammatory mechanisms is crucial for complete knowledge of AD pathophysiology, hence for its prevention and drug therapy. Accordingly, two lines of preventive therapeutics can be outlined, the first based on anti-inflammatory drugs, the second one on anti-oxidative properties.

A pilot study on prostate cancer risk and pro-inflammatory genotypes: pathophysiology and therapeutic implications.

Host genetic factors are crucial risk determinants for many human cancers. In this framework, an interesting model is represented by prostate cancer (PC), which is featured by a complex pathophysiology with a strong genetic component. Multiple genes seem to influence PC risk and several single nucleotide polymorphisms (SNPs) of candidate genes modifying PC susceptibility have been identified. It is noteworthy the potential association of common SNPs in pro-inflammatory genes with PC risk, since chronic inflammation is assumed to play a key role in prostate carcinogenesis. With the aim to identify candidate genes as an experimental basis to develop new strategies for both prevention and treatment of PC, we have investigated the potential role of common SNPs of a gene cluster (TLR4, TLR2, PTGS2 and 5-Lo), involved in innate and inflammatory response, in PC cases, age-matched controls and centenarians from Sicily. Six SNPs were genotyped and their association with PC risk determined. Statistical analysis evidenced a significant association of some pro-inflammatory gene SNPs with an increased risk of PC. Furthermore, significant differences were observed comparing the three groups in the combined presence of a "high responder" pro-inflammatory profile. Overall, the present results suggest the likely association of these SNPs and PC risk, clearly motivating the need of larger studies to confirm the role of these genes in PC development and/or progression.

Risk profiles in type 2 diabetes (metabolic syndrome): integration of IL-10 polymorphisms and laboratory parameters to identify vascular damages related complications.

Recently it has been reported that low serum IL-10 levels are associated with an increased susceptibility for metabolic syndrome and type 2 diabetes mellitus (T2DM). We investigated whether the -1087G/A (rs1800896), -824C/T (rs1800871), -597C/A (rs1800872) IL-10 polymorphisms were associated with type 2 diabetes in a study on a cohort of Italian Caucasians comprising 490 type 2 diabetic and 349 control subjects. Stratifying the data according to IL-10 genotypes, trends for the progressive increase of glucose and neutrophil levels were observed in -1087GG vs. -1087GA vs. -1087AA positive diabetic patients (-1087GG<-1087GA<-1087AA). In addition, evaluating the laboratory parameters according to the -597/-824/-1087 derived haplotypes a significant increase of neutrophils was found in diabetic vs. non-diabetic -597A/ -824T/-1087A positive subjects (Student t test = 3.707, p<0.01). In an attempt to integrate clinical laboratory and immunogenetic data to determine whether these factors taken together define sufficient risk sets for type 2 diabetes we performed the grade-of-membership analysis (GoM). GoM allowed to identify a population of subjects negative for IL-10 -824T allele, 74.4% of which were diabetic patients characterised by vascular damages (Chronic kidney failure and/or Myocardial Infarction), reduction of haematocrit, increase of blood urea nitrogen, creatinin and monocyte levels. These data seem to suggest that -597A/-824T/-1087A negative subjects are more prone to the major type 2 diabetic vascular damages and allow to hypothesise that the contemporary evaluation of some simple hematochemical parameters and IL-10 SNPs may allow identifying diabetic patients with the worse prognostic profile, needing both better complication prevention planning and therapeutic strategies.

Association between the polymorphisms of TLR4 and CD14 genes and Alzheimer's disease.

Alzheimer's disease (AD) is a heterogeneous and progressive neurodegenerative disease which in Western society mainly accounts for clinical dementia. Inflammation plays a key role in AD and dissecting the genetics of inflammation may provide an answer to the possible treatment. Hence, the better understanding of different molecular and cellular inflammatory mechanisms is crucial for complete knowledge of AD pathophysiology, and for its prevention and drug therapy. Accordingly, in the present study we evaluated whether the pro-inflammatory polymorphisms of lipopolysaccaride-receptors, +896A/G Toll-Like Receptor (TLR4) and -260C/T CD14, are risk factors for AD. The study included both 626 AD patients (427 women and 199 men; age range: 53-98 years; mean age: 74.88+/-8.44) from Northern Italy and age and gender matched controls. Our results demonstrate that the +896A/G TLR4 single nucleotide polymorphism (SNP) is associated with AD, whereas no association has been observed with -260C/T CD14 SNP. Furthermore, no differences have been observed evaluating the combined presence of +896A+TLR4/-260T+CD14 "high responder"(proinflammatory-profile). However, our results showing the involvement of TLR4 in AD pathophysiology, strengthen the suggestion that systemic inflammation plays a key role in AD. Carriers of high responder SNP, affected by mild cognitive impairment might, be the ideal target for a preventive treatment with biologics as monoclonal antibodies directed against the pro-inflammatory cytokines to decrease the level of systemic inflammation involved in AD pathophysiology.

Laboratory parameters in centenarians of Italian ancestry.

Studies aimed at collecting reference parameters for haematochemical analysis in the elderly are scarce and for the oldest old subjects even more rare. In order to establish the reference values for the most common laboratory text in long living individuals, we measured haematochemical parameters in >100 years old subjects and in aged subjects as control. Six hundred and two centenarians accepted to be enrolled in the study. A case history containing the complete anamnesis, clinical examinations, evaluation of the clinical cognitive and functional tests, was prepared for each centenarian. Blood samples from 120>100 years old subjects free of chronic or acute Illness (i.e. Alzheimer's disease, metabolic diseases, cardiovascular disease, stroke, neoplastic and infectious diseases) were analysed. A population of 381 healthy old subjects (age range 65-85 years old), recruited in the same geographic areas and with the same clinical characteristic of the health centenarians, was utilized as control. Significant differences were observed for blood glucose, ALT, cholesterol and platelet levels, reduced in centenarians respect to the old subjects, whereas blood urea nitrogen levels were found significantly increased in centenarians. In conclusion, reference values of the healthy adults can generally been utilized also for the healthy oldest old group, with the notable exception of the above mentioned laboratory parameters that appear to be modified in long living subjects.

Taking advantage of viral immune evasion: virus-derived proteins represent novel biopharmaceuticals.

In healthy individuals, natural and adaptive immune responses are able to control virus entry into the host. In particular, CD8(+)-mediated cytotoxicity, sustained by the intervention of CD4+ cells, represents the major key event leading to virus eradication. On the other hand, viruses are able to evade from host immune response via several mechanisms, and special emphasis will be placed on hepatitis C virus and chronic Epstein-Barr infections also in view of personal data. Virokines, viroreceptors, and serpins greatly contribute to viral immune escape, and, among virokines, interleukin (IL)-10 has been object of intensive studies. Finally, all these products have been used as biopharmaceuticals, and, for instance, viral IL-10, chemokine-binding proteins, and serpins exhibit in animal models immunosuppressive, anti-inflammatory, and antiatherogenic activities. As far as their use in human trials is concernded, many cautions are required in order to avoid deleterious side effects and, in particular, the purity of the product, its route and frequency of administration, as well as the immune status of the patient should be taken into serious account.

Interleukin-6 gene polymorphism is an age-dependent risk factor for myocardial infarction in men.

Several studies show that inflammatory components may contribute to atherosclerosis and increase the risk for myocardial infarction (MI). Interleukin-6 (IL-6) is a key pro-inflammatory and immune-modulatory cytokine of relevance for cardiovascular diseases. In this case-control study, 200 patients with MI and 257 healthy controls were genotyped for the polymorphism present in -174 promoter region of the IL-6 gene. Plasma concentrations of IL-6 and C-reactive protein (CRP) in a group of patients and controls were measured. The -174 C allele was associated with an increased risk of developing MI (OR = 2.886, c.i. = 1.801-4.624, P = 0.0001) in older patients, while no association was found in younger ones. The IL-6 plasma levels were higher in patients with MI carrying the CC genotype than in GG patients (CC carriers, IL-6 = 2.97 pg mL(-1) vs. GG carriers = 1.81 pg mL(-1), P = 0.016). A positive correlation of IL-6 levels with those of CRP in serum from patients with MI was also found. Data from this study suggest that the C allele of the promoter polymorphism in the IL-6 gene is a risk factor for MI in the elderly, and the production of the IL-6 is differentially affected by different genotypes of the IL-6 -174 promoter polymorphism.

Evaluation of cytokine polymorphisms (TNFalpha, IFNgamma and IL-10) in Down patients with coeliac disease.

BACKGROUND: In Down syndrome there is an increased prevalence of coeliac disease, but the reasons for this association are yet unknown. AIMS: To evaluate a possible correlation between TNFalpha, IFNgamma and IL-10 genotype polymorphisms with the susceptibility to coeliac disease in Down syndrome patients. METHODS: Single nucleotide polymorphisms of TNFalpha (-308G-->A promoter region), IFNgamma (+874T-->A promoter region) and IL-10 (-1082G-->A promoter region) have been studied in 10 Down patients with coeliac disease, in 40 Down patients without coeliac disease and in 220 healthy controls. Clinical features were also studied in coeliac disease-Down syndrome patients. RESULTS: The 10 coeliac disease-Down syndrome patients had a biopsy proven coeliac disease afterward a serological testing positive to antigliadin, antiendomysium and antitransglutaminase antibodies. Intestinal biopsy showed total atrophy in 6/10 and partial villous atrophy in 4/10 of them. All coeliac disease-Down syndrome patients had silent forms of coeliac disease and classical trisomy 21. No significant differences were observed for the IFNgamma and IL-10 polymorphisms in the studied groups. A significant trend for increase of TNFalpha -308A positive frequency was observed in coeliac disease-Down syndrome patients compared to healthy controls (p=0.043). CONCLUSIONS: Single nucleotide polymorphisms of IFNgamma and IL-10 do not play a role in predisposing Down syndrome patients to coeliac disease, while the TNFalpha -308 allele could be an additional genetic risk factor for coeliac disease in trisomy 21.

TNFalpha, IFNgamma and IL-10 gene polymorphisms in a sample of Sicilian patients with coeliac disease.

BACKGROUND: Coeliac disease is associated with DQ2 and DQ8 alleles, but other genes also confer an additional genetic risk. AIMS: Defining whether the genetic profiles of interleukin-10, tumour necrosis factor alpha and interferon gamma are associated with an increased coeliac disease risk. PATIENTS AND METHODS: The functionally gene polymorphisms of tumour necrosis factor alpha (-308G/A), interferon gamma (+874T/A) and interleukin-10 (-1082G/A) were typed using sequence specific primer-polymerase chain reaction in 110 Sicilian coeliac disease patients and in 220 Sicilian healthy controls. RESULTS: No differences in genotype frequencies of interleukin-10 polymorphisms were found between coeliac disease patients and healthy controls. A significant increase of -308A (p<0.033; OR: 1.72; CI: 1.27-2.33) and of +874T (p: 0.0045; OR: 3.02; CI: 1.47-6.21) allele frequencies, both in hetero- and homozygosis, was observed in coeliac patients in comparison with healthy controls. In addition, simultaneous significant higher percentages of -308A and +874T alleles (p: 0.0066; OR: 2.33; CI: 1.42-3.82) as well as simultaneous significant lower percentages of -308A and +874T alleles (p: 0.003; OR: 0.23; CI: 0.10-0.60) were observed in coeliac patients compared with healthy controls. CONCLUSIONS: Genetically determined higher frequencies of -308A tumour necrosis factor alpha and +874T interferon gamma alleles, both in hetero and in homozygosis and mostly whether simultaneous, may play a role in predisposing to gluten intolerance. Subjects positive for -308A tumour necrosis factor alpha and +874T interferon gamma alleles have an increased risk for coeliac disease.

The -308G/A polymorphism of TNF-alpha influences immunological parameters in old subjects affected by infectious diseases.

Abnormal increments of pro-inflammatory cytokines (IL-6 and TNF-alpha) characterize the outbreak of infectious diseases, which are the major cause of death in the elderly. A counterbalance to the inflammation is exerted by IL-10 with an inhibitory role on TNF-alpha production. As is well known, some cytokine gene polymorphisms influence the cytokine production, playing a role as susceptibility or resistance factors against immune-mediated and infectious disease. Genetic variations in the -308A/G locus for TNF-alpha seems to affect the clinical outcome of some infectious diseases. In fact, the -308A allele is associated with severe septic shock and death. On this basis, we have screened healthy old subjects, nonagenarians and old patients affected by the acute phase of chronic obstructive bronchitis and bronchopneumonia of bacteria origin for the -308G/A locus (PCR-RFLP). Subjects are grouped in A+ (AG, AA genotypes) and A- (GG genotype) and data on IL-6, TNF-alpha, IL-10, NK cell cytotoxicity, zinc and metallothioneins (MTs) gene expression (RT-PCR) were stratified according to different TNF-alpha genotypes. The frequency of the A allele was increased in infected patients in comparison with healthy old controls. No differences existed between A+ and A- young adult, old and nonagenarian controls in tested parameters. Conversely, A+-infected patients displayed elevated IL-6, TNF-alpha and MTmRNA, low IL-10 coupled with impaired NK cell cytotoxicity and lower zinc ion than A- patients. However, the data reported are gender independent. Therefore, the -308A polymorphism at the locus of TNF-alpha may be one of the susceptibility factor for infectious diseases in old persons, particularly considering its association to the increased release of pro-inflammatory cytokines and to the reduction of zinc release and MTs synthesis involved in the control of the inflammatory response. These data strongly suggest that the genetic screening of the -308G/A polymorphism may be a valid tool for identification of subjects needing a more appropriate therapy when affected by acute and/or recurrent infectious diseases.

Role of TLR4 receptor polymorphisms in Boutonneuse fever.

The genetics of the interaction between host and microbes plays an essential role in the survival of the individual and attainment of longevity. The activation of toll-like receptor (TLR)4 plays a key role in natural and clonotypic immune responses. We evaluated whether TLR4 genotype is a component of genetic background protective versus rickettsiosis and whether this background influences longevity. We genotyped for +896A/G TLR4 polymorphism 78 patients affected with Boutonneuse fever, 78 age-matched controls and 78 advanced age individuals from Sicily. The +869G allele, that attenuates receptor signalling, was significantly overrepresented in patients in comparison with age-matched controls. By analyzing data according to gender, this allele was significantly higher in female patients when compared to advanced age women. Pro-inflammatory responses are programmed to resist fatal infections. So, it is not surprising that the genetic background of people that survive to an advanced age may be protective against infections. However, this seems to occur in women but not in men. In a previous study, the +896G TLR4 allele was overrepresented in advanced age men and underrepresented in men affected by myocardial infarction. Thus, previous and present results tend to agree with the suggestion that men and women may follow different trajectories to reach longevity. For men it might be more important to control atherogenesis, whereas for women it might be more important to control infectious diseases.

Association between platelet endothelial cellular adhesion molecule 1 (PECAM-1/CD31) polymorphisms and acute myocardial infarction: a study in patients from Sicily.

Adhesion of circulating cells to the arterial surface is among the first detectable events in atherogenesis. Cellular adhesion molecules, expressed by the vascular endothelium and by circulating leucocytes, mediate cell recruitment and their transendothelial migration. Platelet endothelial cellular adhesion molecule 1 (PECAM-1/CD31), involved in this migration, has been associated with the developmental course of atherosclerosis. A few studies have investigated an association between coronary heart disease and single nucleotide polymorphisms (SNPs) located in functionally important domains of the PECAM-1/CD31 gene. In particular, Ser563Asn and Gly670Arg SNPs have been described as susceptibility factors involved in acute myocardial infarction (AMI) in the Japanese male population. To confirm these observations, we studied 96 male patients (mean age 40 years; age range 20-46) affected by AMI and 118 healthy male controls (mean age 38 years, age range: 20-55), and analysed for the following PECAM-1/CD31 SNPs: Val125Leu, Asn563Ser and Gly670Arg. The frequency of the Gly670Arg polymorphism was significantly higher in patients with AMI (58.9% vs. 48.3%; P = 0.019), whereas the frequencies of the other two SNPs (Leu125Val and Ser563Asn) were not significantly different between patients and controls. By comparing the observed number of 670Arg/Arg genotypes in the patients with the expected number, calculated from the allele frequency in a healthy population, a significance of P = 0.02 (odds ratio, 2.04; 95% CI: 1.1-3.7) was obtained, supporting a recessive model of inheritance. Hence, the differences between patients and controls are significant, but relatively small. However, as AMI is a multifactorial disease, any single mutation will only provide a small or modest contribution to the risk, which also depends on environmental interaction. All in all, we believe that the results of the present study would add support to the role of pro/anti-inflammatory genotypes in determining susceptibility or resistance to immune-inflammatory diseases, including atherosclerosis.

Cytokine genotyping (TNF and IL-10) in patients with celiac disease and selective IgA deficiency.

OBJECTIVE: Selective IgA deficiency (IgAD) and celiac disease (CD) are frequently associated and share the ancestral haplotype human leukocyte antigen (HLA)-8.1, which is characterized by a peculiar cytokine profile. The aim of this study was to evaluate the role of tumor necrosis factor (TNF) and interleukin (IL)-10 alleles in CD and CD-IgAD. METHODS: The distribution of some biallelic polymorphisms of both cytokine promoters (-308G-->A and -863C-->A at TNF promoter sequence and -1082G-->A, -819C-->A, and -592C-->T at IL-10 promoter) were typed using biotilinated specific probes in 32 celiac patients, in 34 CD-IgAD patients, and in 96 healthy controls. RESULTS: In CD and CD-IgAD, the -308A allele was significantly more frequent than in controls, whereas no significant differences were observed for the biallelic polymorphisms at the -863 and for the three IL-10 promoter polymorphisms. The evaluation of combined TNF and IL-10 genotypes showed in CD-IgAD a significant reduction of -308G/-1082G homozygous subjects and both in CD and CD-IgAD groups an increase of 308AA/1082GG. Accordingly, CD-IgAD patients positive both for -308A TNF and -1082A IL-10 showed an increase of TNF-alpha and a reduction of IL-10 serum levels. CONCLUSIONS: Genetically determined increased production of TNF-alpha and reduction of IL-10 may be relevant for susceptibility to CD, mainly in IgAD, as the different allele expression at TNF and IL-10 loci seems to influence cytokine production profile.