Non-coding variation in disorders of sex development.

Genetic studies in Disorders of Sex Development (DSD), representing a wide spectrum of developmental or functional conditions of the gonad, have mainly been oriented towards the coding genome. Application of genomic technologies, such as whole-exome sequencing, result in a molecular genetic diagnosis in ∼50% of cases with DSD. Many of the genes mutated in DSD encode transcription factors such as SRY, SOX9, NR5A1, and FOXL2, characterized by a strictly regulated spatiotemporal expression. Hence, it can be hypothesized that at least part of the missing genetic variation in DSD can be explained by non-coding mutations in regulatory elements that alter gene expression, either by reduced, mis- or overexpression of their target genes. In addition, structural variations such as translocations, deletions, duplications or inversions can affect the normal chromatin conformation by different mechanisms. Here, we review non-coding defects in human DSD phenotypes and in animal models. The wide variety of non-coding defects found in DSD emphasizes that the regulatory landscape of known and to be discovered DSD genes has to be taken into consideration when investigating the molecular pathogenesis of DSD.

Interleukin-12 regulates the production of Bacille Calmette-Guérin-induced interferon-gamma from human cells in a CD40-dependent manner.

Interferon (IFN)-gamma is a cytokine which plays a critical role in the host defence against human tuberculosis infection. There is evidence that interleukin (IL)-12 can exert a potent effect in stimulating the production of IFN-gamma, and it is well known that a costimulatory signal provided by CD40 may enhance IL-12 production by monocytes/macrophages. However, it is unclear whether CD40-CD40L stimulation is able to regulate the production of mycobacterial-induced IFN-gamma through an IL-12-dependent pathway. In this study, we investigated the capacity of the Bacille Calmette-Guérin (BCG) strain of Mycobacterium bovis to induce the production of interferon-gamma through IL-12 and/or CD40 costimulation from human cells. Our data demonstrate that anti-IL-12 antibodies markedly reduced the levels of IFN-gamma produced by the BCG-stimulated human cells, while exogenous recombinant (r)IL-12 up-regulated the production of IFN-gamma. In addition, the stimulatory effect of IL-12 on BCG-induced IFN-gamma secretion was specific, as it was significantly abolished in the presence of anti-IL-12 antibodies. We also observed that the presence of an anti-CD40L monoclonal antibody significantly inhibited the production of IL-12 and IFN-gamma by human cells activated with BCG. In contrast, an isotype control antibody showed no effect on cytokine production. Furthermore, the presence of a trimeric soluble CD40L agonist (CD40T) in cultures increased the production of IL-12 and IFN-gamma. Importantly, the stimulatory capacity of CD40T on BCG-induced IFN-gamma secretion was blocked by a monoclonal antibody against IL-12, indicating that the effect of CD40T on T cells was mediated through IL-12. Together, these studies are the first to demonstrate that BCG-induced IFN-gamma production by human cells appears to be mediated by IL-12 in a CD40-dependent manner and suggest that CD40-CD40L activation may be a critical mediator in regulating the immune response to stimulation with BCG.

Regulation of interleukin-12 production in human cells stimulated with Mycobacterium bovis BCG.

Interleukin 12 (IL-12) is a monokine which plays a critical role in resistance to Mycobacterium tuberculosis infection. However, little is known about the regulation of IL-12 production by human cells stimulated with BCG. Here we report that in vitro infection of human mononuclear cells with M. bovis BCG induces the release of IL-12 protein. We also observed that the ability of BCG to stimulate release of IL-12 from human cells was significantly inhibited by IL-10. The inhibitory effect of IL-10 on the secretion of IL-12 was specific, as it was significantly abolished in the presence of anti-IL-10 neutralizing monoclonal antibody. These results were further confirmed as anti-IL-10 antibodies markedly increased the levels of IL-12, suggesting that BCG-induced IL-10, as well as exogenous IL-10, can regulate IL-12 production by human cells stimulated with M. bovis BCG. Interestingly, IFN-gamma production in response to BCG had no significant increase by the addition of neutralizing antibodies to IL-10. Moreover, anti-IL-12 antibodies markedly reduced the levels of IFN-gamma produced by BCG-stimulated human cells and abrogated the capacity of anti-IL-10 to increase BCG-induced IFN-gamma. These studies are the first to demonstrate a regulatory effect on IL-12 production by human cells infected with M. bovis BCG and at the same time suggest that IL-12 may play an essential role during the human immune response to M. bovis BCG stimulation.