No abstract

The mammalian gastrointestinal tract harbors a microbial community with metabolic activity critical for host health, including metabolites that can modulate effector functions of immune cells. Mice treated with vancomycin have an altered microbiome and metabolite profile, exhibit exacerbated T helper type 2 cell (Th2) responses, and are more susceptible to allergic lung inflammation. Here we show that dietary supplementation with short-chain fatty acids (SCFAs) ameliorates this enhanced asthma susceptibility by modulating the activity of T cells and dendritic cells (DCs). Dysbiotic mice treated with SCFAs have fewer interleukin-4 (IL4)-producing CD4 T cells and decreased levels of circulating immunoglobulin E (IgE). In addition, DCs exposed to SCFAs activate T cells less robustly, are less motile in response to CCL19 in vitro, and exhibit a dampened ability to transport inhaled allergens to lung draining nodes. Our data thus demonstrate that gut dysbiosis can exacerbate allergic lung inflammation through both T cell- and DC-dependent mechanisms that are inhibited by SCFAs.

The prognosis of fludarabine (F)-refractory chronic lymphocytic leukemia (CLL) is very poor, and underlying mechanisms are only partly understood. To assess the contribution of p53 abnormalities to F-refractory CLL, we studied TP53 mutations in the CLL2H trial (subcutaneous alemtuzumab; n ‫؍‬ 99). We found TP53 mutations in 37% of patients. Twelve of 67 (18%) patients without the 17p deletion showed a TP53 mutation and 50% showed evidence of uniparental disomy. A total of 75% of cases with TP53 mutation (without 17p؊) showed clonal evolution/expansion. TP53 mutations had no impact on overall survival (P ‫؍‬ .48). CLL with the 17p deletion or TP53 mutation showed very low miR-34a expression. To investigate the mechanisms underlying refractory CLL beyond p53, we studied cases without 17p؊/TP53 mutation in detail. In several paired samples before and after F-refractory disease, no change in p21/ p53 induction was observed after DNA damage. Although TP53 mutations and 17p deletions are found in a high proportion of F-refractory CLL, more than half of the cases cannot be explained by p53 defects (deletion or mutation), and alternative mechanisms need to be investigated. Alemtuzumab is effective irrespective of genetic high-risk subgroups with TP53 mutations. These clinical trials are registered at www.clinicaltrials. gov as #NCT00274976. (Blood. 2009;114: 2589-2597)

17p (TP53) deletion identifies patients with chronic lymphocytic leukemia (CLL) who are resistant to chemotherapy. The members of the miR-34 family have been discovered to be direct p53 targets and mediate some of the p53-dependent effects. We studied miR-34a and miR-34b/c expression in a large cohort to define their potential role in refractory CLL. While no expression of miR-34b/c could be detected, we found variable expression levels of miR-34a. miR-34a levels were upregulated after DNA damage in the presence of functional p53, but not in cases with 17p deletion (P < .001). We found a strong correlation of low miR-34a levels with impaired DNA damage response, TP53 mutations (without 17p deletion), and fludarabine-refractory disease (also in the absence of 17p deletion). Up-regulation of miR-34a after irradiation was associated with induction of Bax and p21, but not Puma. CLL cells with reduced miR-34a expression showed increased viability after DNA damage independently of 17p status. Therefore, low expression of miR-34a in CLL is associated with p53 inactivation but also chemotherapy-refractory disease, impaired DNA damage response, and apoptosis resistance irrespective of 17p deletion/TP53 mutation. The elucidation of mechanisms underlying miR-34a regulation and overcoming its role in chemotherapy resistance warrant further study. IntroductionChronic lymphocytic leukemia (CLL) is the most frequent type of leukemia in the Western world and is characterized by a highly variable clinical course. 1-3 Traditionally, therapy has been used for advanced stage or symptomatic disease and consists of chemotherapy with alkylating agents, purine analogs, and more recently antibody-chemotherapy combinations. Different molecular prognostic factors have been used to predict time to treatment, likelihood of responding to chemotherapy and survival time. [4][5][6] A central role of the DNA damage-response pathway and particularly p53 has been suggested by the prognostic role of 17p and 11q deletions in CLL. In the critical regions, 2 prominent genes are located that are involved in the cells' response to DNA damage (eg, induced by chemotherapy). ATM and TP53 have been shown to be deleted in virtually all cases with deletion 11q and 17p, respectively. In contrast, TP53 and ATM are mutated in different proportions. [7][8][9][10][11] There is growing evidence that mutations of TP53 or ATM also in the absence of deletion of 17p or 11q are associated with poor prognosis as a result of impaired response to chemotherapy. 12 Particularly loss of 17p has been associated with failure to respond to chemotherapy and short event-free and overall survival. 8,13,14 However, in chemotherapy-refractory CLL, only 30% to 40% percent of cases will have a deletion or mutation of TP53, whereas approximately one-third of the remaining cases have a deletion of 11q. 15 This suggests that almost half of the refractory cases cannot be explained by a direct defect of p53 or loss of 11q. A hypothesis explaining resistance in these cases might be defects ...

Hereditary retinal dystrophies (RD) constitute a group of blinding diseases that are characterized by clinical variability and pronounced genetic heterogeneity. The different forms of RD can be caused by mutations in 4100 genes, including 41600 exons. Consequently, next generation sequencing (NGS) technologies are among the most promising approaches to identify mutations in RD. So far, NGS is not routinely used in gene diagnostics. We developed a diagnostic NGS pipeline to identify mutations in 170 genetically and clinically unselected RD patients. NGS was applied to 105 RD-associated genes. Underrepresented regions were examined by Sanger sequencing. The NGS approach was successfully established using cases with known sequence alterations. Depending on the initial clinical diagnosis, we identified likely causative mutations in 55% of retinitis pigmentosa and 80% of Bardet-Biedl or Usher syndrome cases. Seventy-one novel mutations in 40 genes were newly associated with RD. The genes USH2A, EYS, ABCA4, and RHO were more frequently affected than others. Occasionally, cases carried mutations in more than one RD-associated gene. In addition, we found possible dominant de-novo mutations in cases with sporadic RD, which implies consequences for counseling of patients and families. NGS-based mutation analyses are reliable and cost-efficient approaches in gene diagnostics of genetically heterogeneous diseases like RD.