Genetics of Congenital Heart Disease: Past and Present

Congenital heart disease is the most common congenital anomaly, representing an important cause of infant morbidity and mortality. Congenital heart disease represents a group of heart anomalies that include septal defects, valve defects, and outflow tract anomalies. The exact genetic, epigenetic, or environmental basis of congenital heart disease remains poorly understood, although the exact mechanism is likely multifactorial. However, the development of new technologies including copy number variants, single-nucleotide polymorphism, next-generation sequencing are accelerating the detection of genetic causes of heart anomalies. Recent studies suggest a role of small non-coding RNAs, micro RNA, in congenital heart disease. The recently described epigenetic factors have also been found to contribute to cardiac morphogenesis. In this review, we present past and recent genetic discoveries in congenital heart disease.     

Synthesis and characterization of a pyrrole-alginate conjugate and its application in a biosensor construction

N-(3-Aminopropyl)pyrrole was covalently coupled with alginate in an aqueous-phase reaction by means of carbodiimide-mediated activation chemistry to provide a pyrrole-alginate conjugate for subsequent use in biosensor applications. The pyrrole-alginate conjugate was quantified by UV spectroscopy at 230 nm, by an HPSEC-MALLS analytical method, as well as by FTIR and 13C NMR spectroscopies. The new pyrrole-alginate conjugate was used for the immobilization of polyphenol oxidase (PPO) onto an electrode surface by physical entrapment resulting from the gellification process and electrochemical polymerization of the pyrrole groups. The efficiency of this cross-linking approach (chemical and electrochemical) was investigated by comparing the amount of enzyme released from polypyrrole-alginate and regular alginate. In addition, biosensors were prepared by entrapment of the PPO in polypyrrole-alginate and regular alginate matrixes and their performance for the amperometric determination of catechol chosen as a model analyte was examined, yielding a sensitivity of 350 and 80 microA M(-1) cm(-2), respectively, for polypyrrole-alginate and alginate biosensors.     

Association between the 2008–09 Seasonal Influenza Vaccine and Pandemic H1N1 Illness during Spring–Summer 2009: Four Observational Studies from Canada

Background

In late spring 2009, concern was raised in Canada that prior vaccination with the 2008–09 trivalent inactivated influenza vaccine (TIV) was associated with increased risk of pandemic influenza A (H1N1) (pH1N1) illness. Several epidemiologic investigations were conducted through the summer to assess this putative association.

Methods and Findings

Studies included: (1) test-negative case-control design based on Canada''s sentinel vaccine effectiveness monitoring system in British Columbia, Alberta, Ontario, and Quebec; (2) conventional case-control design using population controls in Quebec; (3) test-negative case-control design in Ontario; and (4) prospective household transmission (cohort) study in Quebec. Logistic regression was used to estimate odds ratios for TIV effect on community- or hospital-based laboratory-confirmed seasonal or pH1N1 influenza cases compared to controls with restriction, stratification, and adjustment for covariates including combinations of age, sex, comorbidity, timeliness of medical visit, prior physician visits, and/or health care worker (HCW) status. For the prospective study risk ratios were computed. Based on the sentinel study of 672 cases and 857 controls, 2008–09 TIV was associated with statistically significant protection against seasonal influenza (odds ratio 0.44, 95% CI 0.33–0.59). In contrast, estimates from the sentinel and three other observational studies, involving a total of 1,226 laboratory-confirmed pH1N1 cases and 1,505 controls, indicated that prior receipt of 2008–09 TIV was associated with increased risk of medically attended pH1N1 illness during the spring–summer 2009, with estimated risk or odds ratios ranging from 1.4 to 2.5. Risk of pH1N1 hospitalization was not further increased among vaccinated people when comparing hospitalized to community cases.

Conclusions

Prior receipt of 2008–09 TIV was associated with increased risk of medically attended pH1N1 illness during the spring–summer 2009 in Canada. The occurrence of bias (selection, information) or confounding cannot be ruled out. Further experimental and epidemiological assessment is warranted. Possible biological mechanisms and immunoepidemiologic implications are considered. Please see later in the article for the Editors'' Summary     

The Src-Family Kinase Inhibitor PP2 Rescues Inducible Differentiation Events in Emergent Retinoic Acid-Resistant Myeloblastic Leukemia Cells

Retinoic acid is an embryonic morphogen and dietary factor that demonstrates chemotherapeutic efficacy in inducing maturation in leukemia cells. Using HL60 model human myeloid leukemia cells, where all-trans retinoic acid (RA) induces granulocytic differentiation, we developed two emergent RA-resistant HL60 cell lines which are characterized by loss of RA-inducible G1/G0 arrest, CD11b expression, inducible oxidative metabolism and p47^phox expression. However, RA-treated RA-resistant HL60 continue to exhibit sustained MEK/ERK activation, and one of the two sequentially emergent resistant lines retains RA-inducible CD38 expression. Other signaling events that define the wild-type (WT) response are compromised, including c-Raf phosphorylation and increased expression of c-Cbl, Vav1, and the Src-family kinases (SFKs) Lyn and Fgr. As shown previously in WT HL60 cells, we found that the SFK inhibitor PP2 significantly increases G1/G0 cell cycle arrest, CD38 and CD11b expression, c-Raf phosphorylation and expression of the aforementioned regulators in RA-resistant HL60. The resistant cells were potentially incapable of developing inducible oxidative metabolism. These results motivate the concept that RA resistance can occur in steps, wherein growth arrest and other differentiation events may be recovered in both emergent lines. Investigating the mechanistic anomalies in resistant cell lines is of therapeutic significance and helps to mechanistically understand the response to retinoic acid’s biological effects in WT HL60 cells.     

One carbon metabolism disturbances and the C677T MTHFR gene polymorphism in children with autism spectrum disorders

Autism spectrum disorders (ASDs), which include the prototypic autistic disorder (AD), Asperger’s syndrome (AS) and pervasive developmental disorders not otherwise specified (PDD‐NOS), are complex neurodevelopmental conditions of unknown aetiology. The current study investigated the metabolites in the methionine cycle, the transsulphuration pathway, folate, vitamin B₁₂ and the C677T polymorphism of the MTHFR gene in three groups of children diagnosed with AD (n= 15), AS (n= 5) and PDD‐NOS (n= 19) and their age‐ and sex‐matched controls (n= 25). No metabolic disturbances were seen in the AS patients, while in the AD and PDD‐NOS groups, lower plasma levels of methionine (P= 0.01 and P= 0.03, respectively) and α‐aminobutyrate were observed (P= 0.01 and P= 0.001, respectively). Only in the AD group, plasma cysteine (P= 0.02) and total blood glutathione (P= 0.02) were found to be reduced. Although there was a trend towards lower levels of serine, glycine, N, N‐dimethylglycine in AD patients, the plasma levels of these metabolites as well as the levels of homocysteine and cystathionine were not statistically different in any of the ASDs groups. The serum levels of vitamin B₁₂ and folate were in the normal range. The results of the MTHFR gene analysis showed a normal distribution of the C677T polymorphism in children with ASDs, but the frequency of the 677T allele was slightly more prevalent in AD patients. Our study indicates a possible role for the alterations in one carbon metabolism in the pathophysiology of ASDs and provides, for the first time, preliminary evidence for metabolic and genetic differences between clinical subtypes of ASDs.     

Paraoxonase 1 activities and polymorphisms in autism spectrum disorders

Autism spectrum disorders (ASD) comprise a complex and heterogeneous group of conditions of unknown aetiology, characterized by significant disturbances in social, communicative and behavioural functioning. Recent studies suggested a possible implication of the high-density lipoprotein associated esterase/lactonase paraoxonase 1 (PON1) in ASD. In the present study, we aimed at investigating the PON1 status in a group of 50 children with ASD as compared to healthy age and sex matched control participants. We evaluated PON1 bioavailability (i.e. arylesterase activity) and catalytic activity (i.e. paraoxonase activity) in plasma using spectrophotometric methods and the two common polymorphisms in the PON1 coding region (Q192R, L55M) by employing Light Cycler real-time PCR. We found that both PON1 arylesterase and PON1 paraoxonase activities were decreased in autistic patients (respectively, P < 0.001, P < 0.05), but no association with less active variants of the PON1 gene was found. The PON1 phenotype, inferred from the two-dimensional enzyme analysis, had a similar distribution in the ASD group and the control group. In conclusion, both the bioavailability and the catalytic activity of PON1 are impaired in ASD, despite no association with the Q192R and L55M polymorphisms in the PON1 gene and a normal distribution of the PON1 phenotype.     

Isolation and characterization of mesenchymal stem cells from human umbilical cord blood: reevaluation of critical factors for successful isolation and high ability to proliferate and differentiate to chondrocytes as compared to mesenchymal stem cells from bone marrow and adipose tissue

Human umbilical cord blood (CB) is a potential source for mesenchymal stem cells (MSC) capable of forming specific tissues, for example, bone, cartilage, or muscle. However, difficulty isolating MSC from CB (CB‐MSC) has impeded their clinical application. Using more than 450 CB units donated to two public CB banks, we found that successful cell recovery fits a hyper‐exponential function of time since birth with very high fidelity. Additionally, significant improvement in the isolation of CB‐MSC was achieved by selecting cord blood units having a volume ≥90 ml and time ≤2 h after donor's birth. This resulted in 90% success in isolation of CB‐MSC by density gradient purification and without a requirement for immunoaffinity methods as previously reported. Using MSC isolated from bone marrow (BM‐MSC) and adipose tissue (AT‐MSC) as reference controls, we observed that CB‐MSC exhibited a higher proliferation rate and expanded to the order of the 1 × 10⁹ cells required for cell therapies. CB‐MSC showed karyotype stability after prolonged expansion. Functionally, CB‐MSC could be more readily induced to differentiate into chondrocytes than could BM‐MSC and AT‐MSC. CB‐MSC showed immunosuppressive activity equal to that of BM‐MSC and AT‐MSC. Collectively, our data indicate that viable CB‐MSC could be obtained consistently and that CB should be reconsidered as a practical source of MSC for cell therapy and regenerative medicine using the well established CB banking system. J. Cell. Biochem. 112: 1206–1218, 2011. © 2011 Wiley‐Liss, Inc.     

Relationship of Pax6 activity levels to the extent of eye development in the mouse, Mus musculus

In this study we extend the mouse Pax6 mutant allelic series to include a homozygous and hemizygous viable hypomorph allele. The Pax6(132-14Neu) allele is a Phe272Ile missense mutation within the third helix of the homeodomain. The mutant Pax6 homeodomain shows greatly reduced binding activity to the P3 DNA binding target. Glucagon-promoter activation by the entire mutant Pax6 product of a reporter gene driven by the G1 paired and homeodomain DNA binding target was slightly increased. We constructed mutant Pax6 genotypes such that Pax6 activity ranged between 100 and 0% and show that the extent of eye development is progressively reduced as Pax6 activity decreased. Two apparent thresholds identify three groups in which the extent of eye development abruptly shifted from complete eye at the highest levels of Pax6 to a rudimentary eye at intermediate levels of Pax6 to very early termination of eye development at the lowest levels of Pax6. Of the two Pax6-positive regions that participate in eye development, the surface ectoderm, which develops into the lens vesicle and the cornea, is more sensitive to reduced levels of Pax6 activity than the optic vesicle, which develops into the inner and outer retinal layers.