Deletion of Porcn in Mice Leads to Multiple Developmental Defects and Models Human Focal Dermal Hypoplasia (Goltz Syndrome)

Focal Dermal Hypoplasia (FDH) is a genetic disorder characterized by developmental defects in skin, skeleton and ectodermal appendages. FDH is caused by dominant loss-of-function mutations in X-linked PORCN. PORCN orthologues in Drosophila and mice encode endoplasmic reticulum proteins required for secretion and function of Wnt proteins. Wnt proteins play important roles in embryo development, tissue homeostasis and stem cell maintenance. Since features of FDH overlap with those seen in mouse Wnt pathway mutants, FDH likely results from defective Wnt signaling but molecular mechanisms by which inactivation of PORCN affects Wnt signaling and manifestations of FDH remain to be elucidated.We introduced intronic loxP sites and a neomycin gene in the mouse Porcn locus for conditional inactivation. Porcn-ex3-7flox mice have no apparent developmental defects, but chimeric mice retaining the neomycin gene (Porcn-ex3-7Neo-flox) have limb, skin, and urogenital abnormalities. Conditional Porcn inactivation by EIIa-driven or Hprt-driven Cre recombinase results in increased early embryonic lethality. Mesenchyme-specific Prx-Cre-driven inactivation of Porcn produces FDH-like limb defects, while ectodermal Krt14-Cre-driven inactivation produces thin skin, alopecia, and abnormal dentition. Furthermore, cell-based assays confirm that human PORCN mutations reduce WNT3A secretion.These data indicate that Porcn inactivation in the mouse produces a model for human FDH and that phenotypic features result from defective WNT signaling in ectodermal- and mesenchymal-derived structures

Inflammatory Markers and Immune Response to Pneumococcal Vaccination in HIV-Positive and -Negative Adults.

BACKGROUND:Members of the Tumor Necrosis Factor (TNF)-superfamily have speculated roles in the response against T-independent type II antigens (TI-II) including pneumococcal polysaccharides (PPS). Dysregulation in their expression is associated with an enhanced risk for pneumococcal disease in neonates but their expression in other high-risk populations including HIV-positive individuals remains to be elucidated. OBJECTIVE:To investigate signals that contribute towards PPS-response and identify potential anomalies that may account for diminished serological response in HIV-positive individuals post Pneumovax (PPV23) immunization. METHODS:Markers of inflammation, C-reactive protein (CRP), IL-6, sCD27 and sCD30, were assessed in HIV-positive and -negative individuals as potential predictors of PPV23 response. Serum levels of B cell activating factor (BAFF), transmembrane activator and calcium-modulator and cytophilin ligand interactor (TACI), B cell maturation antigen (BCMA) and B cell expression of BAFF-R, TACI, BCMA, CD40 and CD21 were assessed in total (unselected) and PPS23F (antigen)-specific B cells of PPV23 immunized HIV-positive and -negative individuals. RESULTS:CRP, sCD27, sCD30 and BAFF were significantly elevated in the serum of HIV-positive individuals but did not adversely affect PPV23 response. Assessment of PPS-specific B cells revealed enhanced TACI and reduced BAFF-R expression compared to unselected B cells in HIV-positive and -negative individuals. Surface TACI was similar but soluble TACI was significantly lower in HIV-positive compared to HIV-negative individuals. CONCLUSION:Current studies highlight a potential role for TACI in PPV23 response based on its enhanced expression on PPS-specific B cells. Although surface levels of TACI were similar, diminished soluble TACI (sTACI) in HIV-positive compared to HIV-negative individuals could potentially decrease BAFF responsiveness and Ig response. A better understanding of the role of TNF receptors could contribute to the design of improved pneumococcal vaccines. TRIAL REGISTRATION:ClinicalTrials.gov NCT02515240

PPS23F-specific memory B cells showed enhanced expression of TACI in HIV-positive and HIV-negative individuals.

<p>Surface expression of BAFF-R (A), CD40 (D) and CD21 (E) on day 7 PPS23F-selected B cells of HIV-positive (grey bars with slanted stripes) and HIV-negative (white bars slanted stripes) individuals compared to day 0 unselected B cells in HIV-positive (plain grey bars) and HIV-negative (plain white bars) individuals. Data is represented as mean ± SEM. *p<0.05, **p<0.01, ***p<0.001.</p

PPS23F-specific B cells showed enhanced expression of CD40 in HIV-positive compared to HIV-negative individuals.

<p>Post-vaccination (Day7) analysis of PPS-specific B cells against the tested serotype 23F showed similar levels of BAFF-R, BCMA, TACI, CD21 but higher levels of CD40 in HIV-positive (black bars) compared to HIV-negative (white bars) individuals. Data is represented as mean ± SEM. *p<0.05, **p<0.01, ***p<0.001.</p

Pro-inflammatory markers are elevated in HIV-positive compared to HIV-negative individuals.

<p>Serum levels of CRP ng/ml (A), IL-6 pg/ml (B), sCD27 U/ml (C) and sCD30 ng/ml (D) were tested in HIV-positive (black bars) and -negative (white bars) individuals. Data is represented as mean ± SEM. *p<0.05, **p<0.01, ***p<0.001.</p

TNF and complement receptor CD21 expression is similar in the total B cells of HIV-positive and HIV-negative individuals.

<p>Expression of receptors BAFF-R, BCMA, TACI, CD40 and CD21 were analyzed in total B cells of HIV-positive (black bars) and -negative (white bars) individuals. Data is represented as mean ± SEM. *p<0.05, **p<0.01, ***p<0.001.</p

Baseline Characteristics of HIV-negative, HIV-positive HAART naïve and experienced individuals enrolled in the current study.

<p>Baseline Characteristics of HIV-negative, HIV-positive HAART naïve and experienced individuals enrolled in the current study.</p

Serum BAFF and sTACI are dysregulated in HIV-positive individuals.

<p>Serum levels of soluble sBAFF pg/ml (A), sTACI pg/ml (B) and sBCMA pg/ml (C) were tested in HIV-positive (black bars) and -negative (white bars) individuals. Data is represented as mean ± SEM. *p<0.05, **p<0.01, ***p<0.001.</p