Mauriac syndrome still exists

Background/objectiveMauriac syndrome (MS) is a rare complication of type 1 diabetes mellitus (DM1). It is related to low insulin concentrations and is less common since longer-acting insulins became available. It is characterized by hepatomegaly, growth and puberty delay, and the presence of elevated transaminases and serum lipids. The aim of this study was to describe the patients from a pediatric diabetic population that fulfill the criteria of MS.Materials and methodsA retrospective analysis of the pediatric diabetic population with diagnostic criteria of MS currently followed at Hospital de Braga, was performed.ResultsFrom a population of 91 patients with DM1 18 years, 6 patients with the criteria for MS were identified: 5 girls, and 1 boy. The age at presentation was 13–17 years, with a minimum interval between DM1 diagnosis and MS criteria of 4 years. All the patients were prescribed intensive insulin therapy (median daily insulin dose: 0.88 U/kg). All had a previous history of poor glycemic control before the diagnosis of MS with glycated hemoglobin (HbA1c) between 8.8 and 12.9%. Increase of hepatic enzymes was present in all the patients; 4 of them had associated hepatomegaly. All the girls presented puberty delay and cushingoid features. None of the patients presented short stature and 5 of them presented mixed dyslipidemia.ConclusionsAlthough MS is an ancient entity described in DM1, it still exists, particularly in adolescent females. Being aware of MS is of extreme importance since most of the clinical features are reversible with better glycemic control.     

1H, 13C and 15N resonance assignment of the soluble form of the Lipid-modified Azurin from Neisseria gonorrhoeae

Lipid-modified azurin (Laz) from Neisseria gonorrhoeae is a type 1 copper protein proposed to be the electron donor to several enzymes involved in the resistance mechanism to reactive oxygen and nitrogen species. Here we report the backbone and side-chain resonance assignment of Laz in the reduced form, which has been complete at 97 %. The predicted secondary structure indicates that this protein belongs to the azurin subfamily of type 1 copper proteins.     

The Brazilian Preference: Cesarean Delivery among Immigrants in Portugal

Objective

To evaluate how the country of origin affects the probability of being delivered by cesarean section when giving birth at public Portuguese hospitals.

Study Design

Women delivered of a singleton birth (n = 8228), recruited from five public level III maternities (April 2005–August 2006) during the procedure of assembling a birth cohort, were classified according to the country of origin and her migration status as Portuguese (n = 7908), non-Portuguese European (n = 84), African (n = 77) and Brazilian (n = 159). A Poisson model was used to evaluate the association between country of birth and cesarean section that was measured by adjusted prevalence ratio (PR) and respective 95% confidence intervals (95%CI).

Results

The cesarean section rate varied from 32.1% in non-Portuguese European to 48.4% in Brazilian women (p = 0.008). After adjustment for potential confounders and compared to Portuguese women as a reference, Brazilian women presented significantly higher prevalence of cesarean section (PR = 1.26; 95%CI: 1.08–1.47). The effect was more evident among multiparous women (PR = 1.39; 95%CI: 1.12–1.73) and it was observed when cesarean section was performed either before labor (PR = 1.43; 95%CI: 0.99–2.06) or during labor (PR = 1.30; 95%CI: 1.07–1.58).

Conclusions

The rate of cesarean section was significantly higher among Brazilian women and it was independent of the presence of any known risk factors or usual clinical indications, suggesting that cultural background influences the mode of delivery overcoming the expected standard of care and outcomes in public health services.     

The Bacterial Protein Azurin Impairs Invasion and FAK/Src Signaling in P-Cadherin-Overexpressing Breast Cancer Cell Models

P-cadherin overexpression occurs in about 30% of all breast carcinomas, being a poor prognostic factor for breast cancer patients. In a cellular background of wild-type E-cadherin, we have previously shown that its expression promotes invasion, motility and migration of breast cancer cells due to the induced secretion of metalloproteases (MMPs) to the extracellular medium and to the concomitant shedding of a pro-invasive soluble form of this protein (sP-cad). Azurin is secreted by Pseudomonas aeruginosa and induces in vitro and in vivo cytotoxicity after its preferential penetration in human cancer cells relative to normal cells. Three different breast cancer cell lines, MCF-7/AZ.Mock, MCF-7/AZ.Pcad and SUM149 were treated with sub-killing doses of azurin. Invasion of these cells was measured using Matrigel Invasion Assays and MTT assays were performed to determine cell viability upon treatment and the effects on cadherins expression was determined by Western blot and Immunofluorescence. Gelatin Zymography was used to determine activity of MMP2 in the conditioned media of azurin treated and untreated cells and the phosphorylation levels of intracellular signaling proteins were determined by Western blot. The invasive phenotype of these breast cancer cells was significantly reduced by azurin. Azurin (50–100 µM) also caused a specific decrease on P-cadherin protein levels from 30–50% in MCF-7/AZ.Pcad and SUM149 breast cancer cell lines, but the levels of E-cadherin remain unaltered. More, the levels of sP-cad and the activity of MMP2 were reduced in the extracellular media of azurin treated cells and we also observed a decrease in the phosphorylation levels of both FAK and Src proteins. Our data show that azurin specifically targets P-cadherin, not E-cadherin, abrogating P-cadherin-mediated invasive effects and signaling. Therefore, azurin could possibly be considered a therapeutic tool to treat poor-prognosis breast carcinomas overexpressing P-cadherin in a wild type E-cadherin context.     

Pulsed electromagnetic fields promote repair of focal articular cartilage defects with engineered osteochondral constructs

Articular cartilage injuries are a common source of joint pain and dysfunction. We hypothesized that pulsed electromagnetic fields (PEMFs) would improve growth and healing of tissue-engineered cartilage grafts in a direction-dependent manner. PEMF stimulation of engineered cartilage constructs was first evaluated in vitro using passaged adult canine chondrocytes embedded in an agarose hydrogel scaffold. PEMF coils oriented parallel to the articular surface induced superior repair stiffness compared to both perpendicular PEMF (p = .026) and control (p = .012). This was correlated with increased glycosaminoglycan deposition in both parallel and perpendicular PEMF orientations compared to control (p = .010 and .028, respectively). Following in vitro optimization, the potential clinical translation of PEMF was evaluated in a preliminary in vivo preclinical adult canine model. Engineered osteochondral constructs (∅ 6 mm × 6 mm thick, devitalized bone base) were cultured to maturity and implanted into focal defects created in the stifle (knee) joint. To assess expedited early repair, animals were assessed after a 3-month recovery period, with microfracture repairs serving as an additional clinical control. In vivo, PEMF led to a greater likelihood of normal chondrocyte (odds ratio [OR]: 2.5, p = .051) and proteoglycan (OR: 5.0, p = .013) histological scores in engineered constructs. Interestingly, engineered constructs outperformed microfracture in clinical scoring, regardless of PEMF treatment (p < .05). Overall, the studies provided evidence that PEMF stimulation enhanced engineered cartilage growth and repair, demonstrating a potential low-cost, low-risk, noninvasive treatment modality for expediting early cartilage repair.     

Activation, dysfunction and retention of T cells in vaccine sites after injection of incomplete Freund’s adjuvant, with or without peptide

We conducted a randomized clinical trial in 45 patients with resected AJCC stage IIB-IV melanoma to characterize cellular and molecular events at sites of immunization with incomplete Freund’s adjuvant (IFA) alone, or a melanoma vaccine in IFA. At a primary vaccine site, all patients received a multi-peptide melanoma vaccine in IFA. At a replicate vaccine site, which was biopsied, group 1 received IFA only; group 2 received vaccine in IFA. Lymphocytes isolated from replicate vaccine site microenvironments (VSME) were compared to time-matched peripheral blood mononuclear cells (PBMC) in ELISpot and flow cytometry assays. Compared to PBMC, the VSME had fewer naïve and greater proportions of effector memory CD8⁺ T cells (T_CD8). The vast majority of T_CD8 within the VSME were activated (CD69⁺), with a concentration of antigen-specific (tetramer^pos) cells in the VSME, particularly in vaccine sites with peptide (group 2). CXCR3⁺ lymphocytes were concentrated in the VSME of all patients, suggesting IFA-induced chemokine recruitment. T_CD8 expression of retention integrins αEβ7 and α1β1 was elevated in VSME, with the highest levels observed in antigen-specific cells in VSME containing peptide (group 2). T_CD8 retained in the VSME of both groups were strikingly dysfunctional, with minimal IFN-γ production in response to peptide stimulation and few tetramer^pos cells producing IFN-γ. These data suggest that vaccine-induced selective retention and dysfunction of antigen-specific T_CD8 within VSME may represent a significant mechanism underlying transient immune responses and low clinical response rates to peptide vaccines administered in IFA.     

Unconjugated Bilirubin Restricts Oligodendrocyte Differentiation and Axonal Myelination

High levels of serum unconjugated bilirubin (UCB) in newborns are associated with axonal damage and glial reactivity that may contribute to subsequent neurologic injury and encephalopathy (kernicterus). Impairments in myelination and white matter damage were observed at autopsy in kernicteric infants. We have recently reported that UCB reduces oligodendrocyte progenitor cell (OPC) survival in a pure OPC in vitro proliferative culture. Here, we hypothesized that neonatal hyperbilirubinemia may also impair oligodendrocyte (OL) maturation and myelination. We used an experimental model of hyperbilirubinemia that has been shown to mimic the pathophysiological conditions leading to brain dysfunction by unbound (free) UCB. Using primary cultures of OL, we demonstrated that UCB delays cell differentiation by increasing the OPC number and reducing the number of mature OL. This finding was combined with a downregulation of Olig1 mRNA levels and upregulation of Olig2 mRNA levels. Addition of UCB, prior to or during differentiation, impaired OL morphological maturation, extension of processes and cell diameter. Both conditions reduced active guanosine triphosphate (GTP)-bound Rac1 fraction. In myelinating co-cultures of dorsal root ganglia neurons and OL, UCB treatment prior to the onset of myelination decreased oligodendroglial differentiation and the number of myelinating OL, also observed when UCB was added after the onset of myelination. In both circumstances, UCB decreased the number of myelin internodes per OL, as well as the myelin internode length. Our studies demonstrate that increased concentrations of UCB compromise myelinogenesis, thereby elucidating a potential deleterious consequence of elevated UCB.     

Distinct signaling pathways regulate TLR2 co-stimulatory function in human T cells

Toll-like receptor 2 (TLR2) serves as a co-stimulatory receptor for human T cells by enhancing T cell receptor (TCR)-induced cytokine production and proliferation. However, it is unknown where signals from the TCR and TLR2 converge to enhance T cell activation. To address this gap, we examined changes in TCR-induced signaling following concurrent TLR2 activation in human T cells. Both proximal TCR-mediated signaling and early NFκB activation were not enhanced by TCR andTLR2 co-activation, potentially due to the association of TLR2 with TLR10. Instead, TLR2 co-induction did augment Akt and Erk1/Erk2 activation in human T cells. These findings demonstrate that TLR2 activates distinct signaling pathways in human T cells and suggest that alterations in expression of TLR2 co-receptors may contribute to aberrant T cell responses.     

Characterization of the 2012 Highly Pathogenic Avian Influenza H7N3 Virus Isolated from Poultry in an Outbreak in Mexico: Pathobiology and Vaccine Protection

In June of 2012, an H7N3 highly pathogenic avian influenza (HPAI) virus was identified as the cause of a severe disease outbreak in commercial laying chicken farms in Mexico. The purpose of this study was to characterize the Mexican 2012 H7N3 HPAI virus (A/chicken/Jalisco/CPA1/2012) and determine the protection against the virus conferred by different H7 inactivated vaccines in chickens. Both adult and young chickens intranasally inoculated with the virus became infected and died at between 2 and 4 days postinoculation (p.i.). High virus titers and viral replication in many tissues were demonstrated at 2 days p.i. in infected birds. The virus from Jalisco, Mexico, had high sequence similarity of greater than 97% to the sequences of wild bird viruses from North America in all eight gene segments. The hemagglutinin gene of the virus contained a 24-nucleotide insert at the hemagglutinin cleavage site which had 100% sequence identity to chicken 28S rRNA, suggesting that the insert was the result of nonhomologous recombination with the host genome. For vaccine protection studies, both U.S. H7 low-pathogenic avian influenza (LPAI) viruses and a 2006 Mexican H7 LPAI virus were tested as antigens in experimental oil emulsion vaccines and injected into chickens 3 weeks prior to challenge. All H7 vaccines tested provided ≥90% protection against clinical disease after challenge and decreased the number of birds shedding virus and the titers of virus shed. This study demonstrates the pathological consequences of the infection of chickens with the 2012 Mexican lineage H7N3 HPAI virus and provides support for effective programs of vaccination against this virus in poultry.     

Aging bone marrow mesenchymal stromal cells have altered membrane glycerophospholipid composition and functionality

Human mesenchymal stem/stromal cells (hMSC) are increasingly used in advanced cellular therapies. The clinical use of hMSCs demands sequential cell expansions. As it is well established that membrane glycerophospholipids (GPL) provide precursors for signaling lipids that modulate cellular functions, we studied the effect of the donor''s age and cell doublings on the GPL profile of human bone marrow MSC (hBMSC). The hBMSCs, which were harvested from five young and five old adults, showed clear compositional changes during expansion seen at the level of lipid classes, lipid species, and acyl chains. The ratio of phosphatidylinositol to phosphatidylserine increased toward the late-passage samples. Furthermore, 20:4n-6-containing species of phosphatidylcholine and phosphatidylethanolamine accumulated while the species containing monounsaturated fatty acids (FA) decreased during passaging. Additionally, in the total FA pool of the cells, 20:4n-6 increased, which happened at the expense of n-3 polyunsaturated FAs, especially 22:6n-3. The GPL and FA correlated with the decreased immunosuppressive capacity of hBMSCs during expansion. Our observations were further supported by alterations in the gene expression levels of several enzymes involved in lipid metabolism and immunomodulation. The results show that extensive expansion of hBMSCs harmfully modulates membrane GPLs, affecting lipid signaling and eventually impairing functionality.