Dual molecular diagnosis contributes to atypical Prader-Willi phenotype in monozygotic twins.

We describe monozygotic twin girls with genetic variation at two separate loci resulting in a blended phenotype of Prader-Willi syndrome and Pitt-Hopkins syndrome. These girls were diagnosed in early infancy with Prader-Willi syndrome, but developed an atypical phenotype, with apparent intellectual deficiency and lack of obesity. Array-comparative genomic hybridization confirmed a de novo paternal deletion of the 15q11.2q13 region and exome sequencing identified a second mutational event in both girls, which was a novel variant c.145+1G>A affecting a TCF4 canonical splicing site inherited from the mosaic mother. RNA studies showed that the variant abolished the donor splicing site, which was accompanied by activation of an alternative non-canonical splicing-site which then predicts a premature stop codon in the following exon. Clinical re-evaluation of the twins indicated that both variants are likely contributing to the more severe phenotypic presentation. Our data show that atypical clinical presentations may actually be the expression of blended clinical phenotypes arising from independent pathogenic events at two loci.

Antihypertensive therapy in pre-eclampsia: effects of plasma from nonresponsive patients on endothelial gene expression.

AIM: Over 40% of patients with pre-eclampsia are nonresponsive to antihypertensive therapy, but the underlying mechanisms are unknown. We examined the effects of plasma from nonresponsive and responsive patients on endothelial gene expression. PATIENTS & METHODS: PCR array was performed in human umbilical vein endothelial cells (HUVEC) incubated with plasma from nonresponsive (n = 4) and responsive (n = 4) patients. Gene networks and interactions with antihypertensive drugs used in pre-eclampsia were identified by Ingenuity Pathway Analysis. RESULTS: Nifedipine and hydralazine act by upregulate or downregulate genes found to be downregulated or upregulated in HUVEC incubated with plasma from nonresponsive patients. CONCLUSION: Our novel findings suggest that plasma from nonresponsive and responsive patients evoke different responses in HUVEC, and might advance the pharmacogenomics research in pre-eclampsia.

Plasma matrix metalloproteinase-9 levels, MMP-9 gene haplotypes, and cardiovascular risk in obese subjects.

Plasma matrix metalloproteinase (MMP)-9 is a predictor of cardiovascular mortality, and MMP-9 polymorphisms affect plasma MMP-9 levels. However, no study examined whether MMP-9 haplotypes affect MMP-9 levels in obese adults. We examined whether MMP-9 polymorphisms and haplotypes are associated with obesity, and whether they affect MMP-9 levels in obese subjects. We examined the plasma levels of MMP-9 and tissue inhibitor of metalloproteinase (TIMP)-1 in 105 subjects with normal weight (controls), 100 obese subjects, and 156 obese subjects with ≥3 metabolic risk factors (MRFs). We determined genotypes for three polymorphisms: C-1562T (rs3918242), Q279R (A>G, rs17576), and R668Q (G>A, rs17577). MMP-9 levels and activity (MMP-9/TIMP-1 ratio) were higher in obese subjects than in controls (P < 0.05). However, MMP-9 levels were higher in obese subjects with ≥3 MRFs than in obese subjects (P < 0.05). Obese subjects with ≥3 MRFs carrying the GA+AA genotypes for R668Q (G>A) polymorphism had higher MMP-9 levels than subjects carrying the AA genotype (P < 0.05). The "T, G, A" haplotype was more common in both groups of obese subjects than in controls (OR 3.95 and 4.39, respectively; P < 0.01). Notably, obese subjects with ≥3 MRFs carrying the "T, G, A" haplotype had higher MMP-9 levels than subjects carrying the "C, A, G" reference haplotype (P < 0.05). The "T, G, A" haplotype was associated with an increased risk of obesity and affected MMP-9 levels in obese subjects with ≥3 MRFs. Our findings suggest that plasma MMP-9 levels and MMP-9 haplotypes may help to discriminate obese subjects at an increased cardiovascular risk.

NLRP1 L155H Polymorphism is a Risk Factor for Preeclampsia Development.

PROBLEM: Augmented levels of IL-1ß have been pointed out as an important pathogenic factor for preeclampsia development. Inflammasome is the cytoplasmic complex responsible for pro-IL1ß cleavage and IL-1ß secretion. Aim of the study was to evaluate the association between polymorphisms in inflammasome' genes and preeclampsia. METHOD OF STUDY: Selected polymorphisms in inflammasome genes (NLRP1, NLRP3, CARD8, and IL1B) were analyzed in 286 Brazilian women with and 309 without preeclampsia. RESULTS AND CONLCLUSIONS: The NLRP1 variant rs12150220 (L155H) was associated with the development of preeclampsia (OR = 1.58), suggesting a role of this inflammasome receptor in the pathogenesis of this multifactorial disorder.

4G/5G polymorphism modulates PAI-1 circulating levels in obese women.

The increase in plasminogen activator inhibitor type 1 (PAI-1) has been described as a risk factor to thrombosis-related diseases. In addition, it has been demonstrated that the variant 4G of polymorphism 4G/5G located in promoter region of PAI-1 gene is associated with higher PAI-1 levels. We investigate the role of this polymorphism on circulating PAI-1 concentration in a population of 57 obese women (23%, 4G/4G; 49%, 4G/5G and 28%, 5G/5G genotypes). Our results demonstrate a genotype-specific modulation on PAI-1 levels in obese women, thus 5G/5G genotype presented significantly lower levels of plasma PAI-1 when compared to 4G/4G group (46 ± 19 ng/mL vs. 63 ± 13 ng/mL, respectively). Our findings indicate that obese carriers of 4G/4G genotype may have increased risk to develop thrombotic diseases.

Matrix metalloproteinase-9 genotypes and haplotypes are associated with multiple sclerosis and with the degree of disability of the disease.

Multiple sclerosis (MS) is an autoimmune disease causing severe neurological disability. This study was carried out in order to determine whether the MMP-9 C(-1562)T and (CA)(13-25) polymorphisms are associated with MS. A total of 165 patients (92 whites/73 mulattos) and 191 controls (96 whites/95 mulattos) were enrolled in the study. While no difference in C(-1562)T polymorphism was observed between MS and healthy subjects, (CA)(n) genotypes and alleles were associated with MS. Moreover, the haplotypes are not associated with MS but seem to be relevant to the clinical status of MS. Thus the (CA)(n) polymorphism may contribute to MS susceptibility, but C(-1562)T and (CA)(n) haplotypes may modulate disease severity.

Prostaglandin mediates IL-23/IL-17-induced neutrophil migration in inflammation by inhibiting IL-12 and IFNgamma production.

IL-23/IL-17-induced neutrophil recruitment plays a pivotal role in rheumatoid arthritis (RA). However, the mechanism of the neutrophil recruitment is obscure. Here we report that prostaglandin enhances the IL-23/IL-17-induced neutrophil migration in a murine model of RA by inhibiting IL-12 and IFN gamma production. Methylated BSA (mBSA) and IL-23-induced neutrophil migration was inhibited by anti-IL-23 and anti-IL-17 antibodies, COX inhibitors, IL-12, or IFNgamma but was enhanced by prostaglandin E(2) (PGE(2)). IL-23-induced IL-17 production was increased by PGE(2) and suppressed by COX-inhibition or IL-12. Furthermore, COX inhibition failed to reduce IL-23-induced neutrophil migration in IL-12- or IFNgamma-deficient mice. IL-17-induced neutrophil migration was not affected by COX inhibitors, IL-12, or IFNgamma but was inhibited by MK886 (a leukotriene synthesis inhibitor), anti-TNFalpha, anti-CXCL1, and anti-CXCL5 antibodies and by repertaxin (a CXCR1/2 antagonist). These treatments all inhibited mBSA- or IL-23-induced neutrophil migration. IL-17 induced neutrophil chemotaxis through a CXC chemokines-dependent pathway. Our results suggest that prostaglandin plays an important role in IL-23-induced neutrophil migration in arthritis by enhancing IL-17 synthesis and by inhibiting IL-12 and IFNgamma production. We thus provide a mechanism for the pathogenic role of the IL-23/IL-17 axis in RA and also suggest an additional mechanism of action for nonsteroidal anti-inflammatory drugs.

Neutrophil recruitment in immunized mice depends on MIP-2 inducing the sequential release of MIP-1alpha, TNF-alpha and LTB(4).

Neutrophils are thought to play an important role in the tissue damage observed in various autoimmune diseases. Chemokines, cytokines and leukotrienes have recognized roles in the orchestration of neutrophil migration. We have recently shown that antigen-induced neutrophil migration into the peritoneum of immunized mice is mediated by macrophage-inflammatory protein (MIP)-1alpha which interacts with CCR1 and induces the sequential release of TNF-alpha and leukotriene B(4) (LTB(4)). The present study investigates the role of MIP-2 and CXCR2 in the cascade of events leading to mediator generation and neutrophil influx. Antigen challenge of immunized mice induced the expression of CXCR2 and the production of KC and MIP-2 proteins. Antigen-induced neutrophil migration was inhibited by a CXCR2 receptor antagonist (repertaxin) or an anti-MIP-2 antibody, but not by an anti-KC antibody. Administration of MIP-2 promoted a dose-dependent neutrophil migration in naive mice which was inhibited by repertaxin, anti-TNF-alpha, anti-MIP-1alpha antibodies or by MK886 (leukotriene synthesis inhibitor). MIP-2 administration induced the release of MIP-1alpha, TNF-alpha and LTB(4), and the release of the latter two was inhibited by anti-MIP-1alpha antibody treatment. Our studies highlight the intricate balance between mediator production and action during an immune-mediated inflammatory response and suggest a mediator cascade leading to neutrophil influx following antigen challenge of immunized mice: MIP-2 --> MIP-1alpha --> TNF-alpha --> LTB(4).

Differences in virulence of Sporothrix schenckii conidia related to culture conditions and cell-wall components.

A murine model was used to evaluate the virulence of Sporothrix schenckii conidia cultured for 4, 7, 10 or 12 days in Sabouraud's dextrose broth (SDB). A correlation was observed between length of culture and virulence. Mice infected intravenously with S. schenckii conidia cultured for 4 or 7 days showed 40-100% cumulative mortality. In contrast, mice infected with conidia from cultures grown for 10 or 12 days in SDB showed no mortality (100% survival). A much greater accumulation of fungal colony forming units (cfu) was observed in the lungs, livers and spleens of mice inoculated with conidia of S. schenckii cultured for 7 days than in mice infected with conidia cultured for 12 days. The livers of mice from the former group showed a widespread granulomatous reaction whereas mice inoculated with S. schenckii cultured for 12 days showed a more limited response with fewer granulomas. No difference in viability or replicative capacity was discerned for these S. schenckii cultured cells. However, the more virulent forms of the fungus showed differences in cell-wall sugar composition with rhamnose:mannose molar ratios of 1.7:1.0 for cells cultured for 4 days and 1.0:1.7 for conidia cultured for 12 days. These results suggest that the virulence of S. schenckii conidia may be determined by their cell-wall composition.