ADP-ribosylation factor 6 mediates E-cadherin recovery by chemical chaperones

E-cadherin plays a powerful tumor suppressor role. Germline E-cadherin mutations justify 30% of Hereditary Diffuse Gastric Cancer (HDGC) and missense mutations are found in 30% of these families. We found possible to restore in vitro mutant E-cadherin associated to HDGC syndrome by using Chemical Chaperones (CCs). Herein, our aim was to disclose the molecular mechanisms underlying the CCs effects in E-cadherin regulation. Using cells stably expressing WT E-cadherin or two HDGC-associated missense mutations, we show that upon DMSO treatment, not only mutant E-cadherin is restored and stabilized at the plasma membrane (PM), but also Arf6 and PIPKIγ expressions are altered. We show that modulation of Arf6 expression partially mimics the effect of CCs, suggesting that the cellular effects observed upon CCs treatment are mediated by Arf6. Further, we show that E-cadherin expression recovery is specifically linked to Arf6 due to its role on endocytosis and recycling pathways. Finally, we demonstrated that, as DMSO, several others CCs are able to modulate the trafficking machinery through an Arf6 dependent mechanism. Interestingly, the more effective compounds in E-cadherin recovery to PM are those that simultaneously inhibit Arf6 and stimulate PIPKIγ expression and binding to E-cadherin. Here, we present the first evidence of a direct influence of CCs in cellular trafficking machinery and we show that this effect is of crucial importance in the context of juxtamembrane E-cadherin missense mutations associated to HDGC. We propose that this influence should be taken into account when exploring the therapeutic potential of this type of chemicals in genetic diseases associated to protein-misfolding.     

McKusick-Kaufman or Bardet-Biedl syndrome? A new borderline case in an Italian nonconsanguineous healthy family

McKusick-Kaufman syndrome (MKS, OMIM #236700) is a rare syndrome inherited in an autosomal recessive pattern with a phenotypic triad comprising hydrometrocolpos (HMC), postaxial polydactyly (PAP), and congenital cardiac disease (CHD). The syndrome is caused by mutations in the MKKS gene mapped onto chromosome 20p12 between D20S162 and D20S894 markers. Mutations in the same gene causes Bardet-Biedl-6 syndrome (BBS-6, OMIM #209900) inherited in an autosomal recessive pattern. BBS-6 comprises retinitis pigmentosa, polydactyly, obesity, mental retardation, renal and genital anomalies. HMC, CHD, and PAP defects can also occur in BBS-6, and there is a significant clinical overlap between MKS and BBS-6 in childhood. We describe a new borderline case of MKS and BBS syndrome and suggest insights for understanding correlation between MKKS gene mutations and clinical phenotype. Here, we report the results of molecular analysis of MKKS in a female proband born in an Italian nonconsanguineous healthy family that presents HMC and PAP. The mutational screening revealed the presence of two different heterozygous missense variants (p.242A>S in exon 3, p.339 I>V in exon 4) in the MKKS gene, and a nucleotide variation in 5'UTR region in exon 2 (-417 A>C).     

HELP LDL Apheresis Reduces Plasma Pentraxin 3 in Familial Hypercholesterolemia

Background

Pentraxin 3 (PTX3), a key component of the humoral arm of innate immunity, is secreted by vascular cells in response to injury, possibly aiming at tuning arterial activation associated with vascular damage. Severe hypercholesterolemia as in familial hypercholesterolemia (FH) promotes vascular inflammation and atherosclerosis; low-density lipoprotein (LDL) apheresis is currently the treatment of choice to reduce plasma lipids in FH. HELP LDL apheresis affects pro- and antiinflammatory biomarkers, however its effects on PTX3 levels are unknown. We assessed the impact of FH and of LDL removal by HELP apheresis on PTX3.

Methods

Plasma lipids, PTX3, and CRP were measured in 19 patients with FH undergoing chronic HELP LDL apheresis before and after treatment and in 20 control subjects. In the patients assessment of inflammation and oxidative stress markers included also plasma TNFα, fibrinogen and TBARS.

Results

At baseline, FH patients had higher (p = 0.0002) plasma PTX3 than matched control subjects. In FH PTX3 correlated positively (p≤0.05) with age, gender and CRP and negatively (p = 0.01) with HELP LDL apheresis vintage. The latter association was confirmed after correction for age, gender and CRP. HELP LDL apheresis acutely reduced (p≤0.04) plasma PTX3, CRP, fibrinogen, TBARS and lipids, but not TNFα. No association was observed between mean decrease in PTX3 and in LDL cholesterol. PTX3 paralleled lipids, oxidative stress and inflammation markers in time-course study.

Conclusion

FH is associated with increased plasma PTX3, which is acutely reduced by HELP LDL apheresis independently of LDL cholesterol, as reflected by the lack of association between change in PTX3 and in LDL levels. These results, together with the finding of a negative relationship between PTX3 and duration of treatment suggest that HELP LDL apheresis may influence both acutely and chronically cardiovascular outcomes in FH by modulating PTX3.     

Quantifying the re-exposure process to an infectious agent. Measles and Varicella as examples

Background and objectives

For viral infections conferring what is usually considered as permanent immunity, re-exposure to the pathogen due to contacts with infectious individuals might be critical for immunity boosting. A major example is represented by the varicella-zoster virus (VZV) where re-exposure is thought to lead to boosting of cell mediated immunity (CMI), which plays a protective role against the development of herpes zoster (HZ). Similar concerns have recently been raised also in relation to measles. However, while the first effective exposure, i.e. infection, has been the object of many studies, both theoretical and epidemiological, there has been no corresponding investigation of the re-exposure process.

Methodology and data

By combining basic concepts from deterministic and stochastic modelling of infection, we develop a basic model for quantifying the timing and number of re-exposures and, consequently, the potential for immune boosting at any given age. The model is then applied to measles, mumps and rubella (MMR) in the UK, and to varicella in Italy, using literature estimates of the pre-vaccination forces of infection.

Results

We supply analytical expressions for the expected number of lifetime re-exposures and for underlying age-patterns, including the average age at which the last re-exposure occurs. Based on updated estimates of the force of VZV infection, we show that the expected number of boosting opportunities of CMI might be in the range 2–3, which is consistent with recent findings about the development of herpes zoster. We also show that the estimate of the age at which the last re-exposure to VZV occurs is highly sensitive to the underlying form of age dependence of the force of infection.

Conclusions

Our results contribute to the study of the potential immunity boosting effect of re-exposures to an infective agent by quantifying the re-exposure process.     

Helicobacter pylori induces gastric epithelial cell invasion in a c-Met and type IV secretion system-dependent manner

Helicobacter pylori interacts with gastric epithelial cells, activating signaling pathways important for carcinogenesis. In this study we examined the role of H. pylori on cell invasion and the molecular mechanisms underlying this process. The relevance of H. pylori cag pathogenicity island-encoded type IV secretion system (T4SS), CagA, and VacA for cell invasion was also investigated. We found that H. pylori induces AGS cell invasion in collagen type I and in Matrigel invasion assays. H. pylori-induced cell invasion requires the direct contact between bacteria and cancer cells. H. pylori-mediated cell invasion was dependent on the activation of the c-Met receptor and on increased MMP-2 and MMP-9 activity. The abrogation of the c-Met receptor using the specific NK4 inhibitor or the silencing of c-Met expression with small interference RNA suppressed both cell invasion and MMP activity. Studies with different H. pylori strains revealed that cell invasion, c-Met tyrosine phosphorylation, and increased MMP-2 and MMP-9 activity were all dependent on the presence of a functional bacterial T4SS, but not on VacA cytotoxicity. Our findings demonstrate that H. pylori strains with a functional T4SS stimulate gastric epithelial cell invasion through a c-Met-dependent signaling pathway that comprises an increase in MMP-2 and MMP-9 activity.     

Ecogeographical variation of body size in the newt Triturus carnifex: comparing the hypotheses using an information‐theoretic approach

Aim Ecogeographical variation of body size in vertebrates (e.g. Bergmann's rule) has long been recognized. However, the patterns and causes of intra‐specific ecogeographical variation of body size in ectotherms, and in amphibians in particular, are strongly debated. We identified the relationship between bioclimatic variables and body size predicted a priori by alternative hypotheses (heat balance, endurance, seasonality, starvation resistance, water availability, primary productivity, parental investment) proposed to explain ecogeographical patterns of body size in ectotherms, and we evaluated the relative support of these hypotheses in explaining variation in body size of the Italian crested newt, Triturus carnifex. Location Twenty‐three populations covering the whole range of T. carnifex (Austria, Croatia, Italy and Slovenia). Methods We obtained data on body size (snout–vent length, SVL) of 2639 adult newts from direct measurements and the literature; we obtained high‐resolution environmental data for the sampled localities. We used an information‐theoretic approach to evaluate the support of the data for the different hypotheses. We also integrated information on population genetics in our models. Results We observed strong geographical variation of body size. The best Akaike information criterion (AIC) models showed that populations with larger body size are associated with cold climates and secondarily with high primary productivity. Furthermore, sexual dimorphism increases in cold climates, as the increase in body size was stronger for females. When taking into account population genetics, we did not find support for relationships with the other variables. Main conclusion Our results are consistent with three hypotheses proposed to explain ecogeographical variation in amphibians: heat balance, increased parental investment of females and productivity. Information theory provides the framework for comparing hypotheses rather than looking for patterns. We suggest that evaluating the support for mechanisms can provide better insights than simply assessing whether ecogeographical variation is in agreement with some ‘rule’.     

Macrophage migration inhibitory factor (MIF) is necessary for progression of autoimmune diabetes mellitus

Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine of the innate immune system that plays a major role in the induction of immunoinflammatory responses. To examine the role of endogenous MIF in the pathogenesis of type 1 diabetes (TID) we evaluated the effects of administration of neutralizing anti-MIF antibodies to NOD mice with accelerated forms of diabetes induced by injection of cyclophosphamide or by transfer of diabetogenic spleen cells. Both accelerated forms of diabetes were markedly reduced by anti-MIF antibody. Furthermore, MIF-deficient (MIF(-/-)) mice were less susceptible to the induction of immunoinflammatory diabetes, insulitis and apoptosis within the endocrine pancreas by multiple low doses of streptozotocin (MLD-STZ) than genetically matched wild type (WT) mice. MIF deficiency resulted in lower proliferation and lymphocyte adhesion, as well as reduced production from the spleens and peritoneal cells of a variety of inflammatory mediators typically associated with development of the disease including IL-12, IL-23, TNF-alpha, and IL-1beta. Furthermore, MIF deletion affected the production of IL-18, TNF-alpha, IL-1beta, and iNOS in the islets of Langerhans. These data, along with the higher expression of IL-4 and TGF-beta observed in the periphery and in the pancreas of MLD-STZ-challenged MIF(-/-) mice as compared to WT controls suggest that MIF deficiency has induced an immune deviation towards protective type 2/3 response. These results suggest that MIF participates in T1D by controlling the functional activity of monocytes/macrophages and T cells and modulating their secretory capacity of pro- and anti-inflammatory molecules.     

Tumor-derived MUC1 mucins interact with differentiating monocytes and induce IL-10highIL-12low regulatory dendritic cell

Dendritic cells (DC) initiate immunity by the activation of naive T cells and control immunity through their ability to induce unresponsiveness of lymphocytes by mechanisms that include deletion and induction of regulatory cells. An inadequate presentation to T cells by tumor-induced "regulatory" DC, among several mechanisms, can explain tolerance to tumor-associated Ags. In this study, we show that tumor-derived mucin profoundly affects the cytokine repertoire of monocyte-derived DC and switch them into IL-10(high)IL-12(low) regulatory APCs with a limited capacity to trigger protective Th1 responses. In fact, DC cocultured with pancreatic tumor cell lines in a Transwell system did not reach full maturation, had low immunostimulatory functions, did not produce IL-12, and released high levels of IL-10. The involvement of known tumor-derived immune-suppressive factors (e.g., vascular endothelial growth factor, TGF-beta, IL-6, and IL-10) was considered and excluded. We provide evidence that tumor-derived MUC1 mucins are responsible for the impaired DC maturation and function. DC obtained in the presence of tumor microenvironment preferentially polarized IL-4(+) response. Moreover, T cells primed by these regulatory DC became anergic and behaved as suppressor/regulatory cells. These findings identify mucin secretion as a novel mechanism of tumor escape from immune surveillance and provide the basis for the generation of potentially tolerogenic DC.