Nicoletta Gallo

CEA mRNA identification in peripheral blood is feasible for colorectal, but not for gastric or pancreatic cancer staging.

Objective: It has been suggested that the molecular identification of cancer cells in the circulation may be useful in predicting the presence of micrometastasis in several cancer types. The aim of the present study was therefore to assess the feasibility of CEA mRNA identification in blood for diagnosing and staging colorectal, gastric and pancreatic cancer. Methods: We studied 16 control subjects, 69 patients with colorectal (CRC), 30 with gastric (GC), 27 with pancreatic cancer (PC) and 8 with benign diseases of the pancreatobiliary tree. At diagnosis CEA mRNA was identified in peripheral blood by means of a RT-PCR procedure. Results: The specificity of this test in control subjects was 94%, and its sensitivity in identifying CRC, GC and PC were 34, 37 and 41%, respectively. False-positive findings were recorded in 25% patients with benign diseases. No association was found between CEA mRNA and stage in patients with GC or PC. In CRC patients, positive CEA mRNA findings were correlated with local spread (χ2 = 14.6, p < 0.01), lymph node (χ2 = 18.95, p < 0.001) and distant metastasis (χ2 = 11.3, p < 0.001). In these cases, CEA mRNA, but not CEA, was entered in stepwise discriminant analysis to classify the presence of lymph node metastasis. Conclusions: The molecular detection of micrometastasis in the blood by means of CEA mRNA identification is feasible for colorectal, but not for gastric or pancreatic cancer staging. Further studies are needed in order to define the clinical utility of this marker also in follow-up protocols.

Helicobacter pylori Infection in Children and Adults: A Single Pathogen But a Different Pathology

Background. The aims of this retrospective study were to ascertain in large series of children and adults: the relationship of the infecting strain to gastric mucosal lesions; and the relationship of the infecting strain to its duodenal localization.

Interleukin 12 gene polymorphisms enhance gastric cancer risk in H pylori infected individuals

Bacterial, environmental, population related, and individual host factors are major determinants of the outcome of H pylori infection.1,2 Many bacterial virulence genes—including the pathogenicity island cagA , the s1m1 vacA alleles, babA2 , sabA , and oipA —have been associated with a higher degree of gastric mucosal inflammation, intestinal metaplasia, gastric or duodenal ulcer, gastric adenocarcinoma, and MALToma.1,3–7 H pylori triggers and maintains gastric mucosal inflammation by different mechanisms, which are partly strain dependent and partly strain independent.1 T and B lymphocyte activation and infiltration of the gastric mucosa depend on H pylori antigen processing. The number of infiltrating polymorphonuclear cells varies depending on the virulence of the infecting strain, being much greater when infections are caused by cagA positive strains.3,5,7–9 The inflammatory cells infiltrating H pylori infected gastric mucosa produce a pattern of proinflammatory cytokines.10,11 High mucosal levels of mononuclear cytokines (IL8, IL6, IL1β, tumour necrosis factor α (TNFα), and interferon γ (IFNγ)) and lymphocytic derived cytokines (IL2, IL2R) have been described in H pylori infected patients.10–13 H pylori infection also induces the production of IL12,14–16 a heterodimeric proinflammatory protein that triggers the production of IFNγ and favours the differentiation of T helper 1 (Th1) cells,17,18 which, in H pylori infected mucosa, prevail over Th2 cells.15,16,19 The ability of IL12 to induce Th1 is one of the biological bases of the importance of this cytokine in resisting most bacteria, including H pylori , and also intracellular protozoa and fungal pathogens.18,20,21 Cellular sources of IL12 in response to infections are mainly dendritic cells and phagocytes.16–21 The two subunits of IL12—p35 and p40—are encoded by different genes, named IL12A and IL12B respectively, which are unrelated …

Retrovirus-mediated herpes simplex virus thymidine kinase gene transfer in pancreatic cancer cell lines: an incomplete antitumor effect.

Introduction The transfer of drug-susceptible (suicide) genes to tumor cells by retroviral or adenoviral vectors is a novel approach to the treatment of human tumors. Aims To ascertain the antitumor effect of retroviral transduction of the pancreatic cancer cell lines MIA PaCa 2, CAPAN-1, PANC1, and PSN1 with the herpes simplex virus thymidine kinase (HSV-TK) gene. Methodology The vector carried a neoselectable marker gene, the human interleukin-2 gene, an internal ribosome entry coding site, and the region coding HSV-TK. Results Twenty micromoles or less of ganciclovir did not modify nontransduced TK− cell growth, whereas ≥100 &mgr;mol completely inhibited TK− cell growth, indicating that this dosage is cytotoxic per se. The 4 TK− and the 4 transduced cell lines were treated daily with 0.001, 0.01, 0.1, 1, 10, and 20 &mgr;mol of ganciclovir for 13 days. CAPAN-1 cell growth was completely inhibited by 0.1 &mgr;mol of ganciclovir; higher doses were required to kill PANC1 (10 &mgr;mol) and PSN1 (20 &mgr;mol). MIA PaCa 2 cell growth decreased following a 20-&mgr;mol ganciclovir dosing. The bystander effect was great in the CAPAN-1 cell line and moderate in PANC1; no bystander effect was recorded in MIA PaCa 2 and PSN1 cell lines. Conclusion Gene therapy with HSV-TK for pancreatic cancer seems effective in only a limited number of tumor-derived cell lines, and this limits its application in vivo.

IgA anticardiolipin and IgA anti-β2 glycoprotein I antibody positivity determined by fluorescence enzyme immunoassay in primary antiphospholipid syndrome.

Abstract Background: Primary antiphospholipid syndrome (PAPS) is an autoimmune disease characterized by thrombosis and/or pregnancy morbidity as well as blood antiphospholipid (aPL) antibodies such as anticardiolipin (aCL), anti-β2 glycoprotein I (anti-β2GPI) antibodies of the IgG/IgM isotype and lupus anticoagulant (LA). The clinical significance of aCL and anti-β2GPI antibodies of the IgA isotype in PAPS is still a controversial issue. Methods: Sera and plasma were collected from 84 PAPS patients (54 with thrombosis and/or pregnancy morbidity and 30 with pregnancy morbidity alone), 66 seronegative patients (subjects with clinical manifestations of PAPS although with negative results on conventional antiphospholipid antibody testing), and 78 healthy blood donors. IgA aCL and IgA anti-β2GPI were determined using fluorescence enzyme immunoassay (FEIA), (EliATM, Phadia AB, Uppsala, Sweden). For comparison purposes, the sera were also tested for IgG/IgM aCL/anti-β2GPI antibodies using the same immunoassay method. LA was assayed following internationally accepted guidelines. Results: Present respectively in 19% and 50% of the PAPS patients studied, IgA aCL and IgA anti-β2GPI antibody frequencies were both statistically significant (p=0.001 and p<0.001, respectively). The mean titers of both IgA aCL and IgA anti-β2GPI antibodies were higher in the thrombotic patients, but only the latter were significantly associated with thrombosis. Isolated IgA anti-β2GPI antibody positivity was significantly prevalent (p=0.04) in seven (10.6%) of the seronegative patients. Conclusions: Positivity to IgA anti-β2GPI antibody detected using FEIA was found to be clinically relevant in PAPS patients. Moreover the prevalence of isolated IgA anti-β2GPI antibody positivity was significant in the seronegative patients.

Effect of cagA status on the sensitivity of enzyme immunoassay in diagnosing Helicobacter pylori-infected children.

Background. The aims of our study were twofold. First, we sought to evaluate in symptomatic children the influence of the Helicobacter pylori genotype on gastritis, abdominal pain, and circulating anti–H. pylori IgG antibodies (anti–H. pylori IgG) or pepsinogen A (PGA) and C (PGC). Additionally, we sought to assess anti–H. pylori IgG, PGA, and PGC patterns in a large cohort (N = 921) of asymptomatic children.

Polymorphonuclear oxidative burst after Helicobacter pylori water extract stimulation is not influenced by the cytotoxic genotype but indicates infection and gastritis grade.

Abstract H. pylori-associated gastric mucosal inflammation is characterized by the presence of polymorphonuclear (PMN) leukocyte infiltrate, which is more severe when the infecting strain is cagA positive. After appropriate stimuli, such as bacterial products, PMN release large amounts of oxygen derived free radicals and proteases, to kill the bacterium. H. pylori seems to be particularly resistant to the oxidative machinery of PMN, which can in turn damage the host gastric mucosa. We evaluated peripheral PMN oxidative burst response after stimulation with water extracts from cagA positive (WEcagA+) or negative (WEcagA−) H. pylori strains in infected (n=31) and non-infected patients (n=32) in comparison with healthy controls (n=16); the influence of gastric mucosal inflammatory infiltrate and activity grade on PMN oxidative burst were also assessed. PMN oxidative burst was measured by FACS analysis. H. pylori water extracts were obtained from bacterial culture. H. pylori genotype was determined by means of the polymerase chain reaction. The PMN oxidative burst in H. pylori infected patients was significantly higher than that in H. pylori negative or healthy controls, no differences being found when the results following WEcagA+ and WEcagA-stimulation were compared. The difference in PMN oxidative burst obtained after WEcagA− and E. coli (standard stimulus for PMN oxidative burst) stimulation discriminated H. pylori infected from non-infected patients with a sensitivity of 90 % and a specificity of 97 %. The grade of PMN oxidative burst correlated with PMN infiltration grade of the gastric mucosa. Our findings allow to conclude that PMN oxidative burst activation by H. pylori WE is species-but not strain-correlated. PMN priming, probably consequent to the action of soluble mediators released by mononuclear cells, makes PMN hyper-responsive to H. pylori products, thus favoring the release in the gastric mucosa of infected patients of large amounts of oxygen-derived free radicals, which are not enough to eliminate the infection, but may contribute to damaging the gastric mucosa itself. Peripheral PMN oxidative burst response to H. pylori WE might furthermore be of help in diagnosing H. pylori infection.

Acute immunomodulatory changes during controlled ovarian stimulation: evidence from the first trial investigating the short-term effects of estradiol on biomarkers and B cells involved in autoimmunity

PurposeThe aim of the present study was to evaluate the in vivo immunomodulatory effects of an acute short-term estradiol (E2) increase on serum levels of B cell-activating factor (BAFF), immunoglobulins (Ig), anti-nuclear antibodies (ANA), and the peripheral B cell phenotype.MethodsWe conducted, at the Infertility Center of the University of Padua, a prospective case–control study on a cohort of infertile normo-responder women (group-A, 63 patients) undergoing controlled ovarian stimulation (COS) compared with an age-matched cohort of normo-ovulatory healthy women (group-B, 39 patients). Three serial blood sample assays were conducted in both groups, at T0, hypothalamic suppression; T1, ovulation induction; and T2, βhCG test in group A, and at T0, 2nd day; T1, 14th day; and T2, 21st day of cycle in group B, and serum levels of E2 and BAFF, BAFF/E2 ratio, circulating IgM, IgG, and IgA, ANA titer, and peripheral B cell phenotype were measured. We compared group-A versus group-B in terms of absolute and E2 normalized values of BAFF at baseline (T0) to verify for possible differences between healthy and infertile women, at T1 to verify for possible differences occurring after spontaneous ovulation versus COS, and at T2 to evaluate differences in serum BAFF levels between pregnant versus non-pregnant patients (considering only group-A) and between non-pregnant women after spontaneous versus COS cycles (group-B versus group-A). In group-A, we also evaluated IgM, IgG, IgA levels, ANA titer, and peripheral B cell phenotype at T0 versus T1 versus T2.ResultsWith the exception of E2 levels at T1 (as expected), no significant differences were found between the two groups for all outcome measures. In group-A, BAFF at T0 positively correlated with IgM levels; marginal zone CD19+/CD27+/IgD+ memory B cell compartment tended to be expanded at T1 when compared with T0.ConclusionsDespite several mechanistic and clinical studies supporting a stimulatory role of E2 on autoimmunity, the acute increase of E2 during COS for infertility treatment does not seem to have a major impact on the immune system.

Insulin autoimmune syndrome (Hirata’s disease) in an Italian patient: a case report and review of the literature

Abstract We describe the case of a 54-year-old Caucasian Italian male experiencing episodes of hypoglycemia, occurring mainly after meals. He had never been exposed to insulin and was taking ramipril, flecainide and acetylsalicylic acid. An oral glucose tolerance test (OGTT) showed high blood glucose levels diagnostic for diabetes mellitus at 120 min and hypoglycemia with inappropriately high insulin levels at 240 min. The 72-h fasting test, abdominal computed tomography (CT) and positron emission tomography-CT were normal. Insulin autoantibodies were positive at high titers, prompting a diagnosis of insulin autoimmune syndrome (IAS). The patient was advised to take frequent, small meals and thus achieved a good control of his hypoglycemic symptoms. After 18 months of this dietary management, his insulin autoantibody levels decreased considerably but remained detectable. During an OGTT, his blood glucose levels at 120 min were now indicative of an impaired glucose tolerance rather than diabetes, and there was improvement in the glucose nadir. The patient had no other clinical or latent autoimmune diseases. Here we discuss the main features of IAS (also known as Hirata’s disease) and review the cases of IAS reported in Italy to date.

The natural history of autoimmune Addison’s disease with a non-classical presentation: a case report and review of literature

Abstract Autoimmune Addison’s disease (AAD) is the most frequent cause of adrenocortical insufficiency. The natural history of AAD usually comprises five consecutive stages with the first stage characterized by the increase of plasma renin consistent with the impairment of pars glomerulosa, which is usually the first affected layer of the adrenal cortex. We describe a 19-year-old female with Hashimoto’s thyroiditis (HT) who underwent an autoantibody screening due to having the personal and family history of other autoimmune diseases in the absence of relevant clinical manifestations. She was positive for adrenal cortex autoantibodies (ACA) and steroid 21-hydroxylase autoantibodies (21-OH Ab) at high titers. She had increased basal levels of ACTH with normal basal cortisol not responding to ACTH stimulation, reduced levels of dehydroepiandrosterone-sulfate but normal levels of orthostatic renin and aldosterone. This scenario was consistent with a subclinical AAD presenting with first impairments in pars fasciculata and reticularis and conserved pars glomerulosa function. Only subsequently, progressive deficiency in pars glomerulosa function has become evident. Review of the literature showed that there was only one case, reported to date, with a similar atypical natural history of AAD. The strategies for screening for ACA/21-OH Ab in patients with HT are discussed.