Congenitalheartdisease (CHD) is one of the most prevalent developmental anomalies and the leading cause of noninfectious morbidity and mortality in newborns. Despite its prevalence and clinical significance, the etiology of CHD remains largely unknown. GATA4 is a highly conserved transcription factor that regulates a variety of physiological processes and has been extensively studied, particularly on its role inheart development. With the combination of TBX5 and MEF2C, GATA4 can reprogram postnatal fibroblasts into functional cardiomyocytes directly. In the past decade, a variety ofGATA4mutations were identified and these findings originally came from familial CHD pedigree studies. Given that familial and sporadic CHD cases allegedly share a basic genetic basis, we explore the GATA4mutationsin different types of CHD. In this study, via direct sequencing of the GATA4 coding region and exon-intron boundaries in 384 sporadic Chinese CHD patients, we identified 12 heterozygous non-synonymous mutations, among which 8 mutations were only found in CHD patients when compared with 957 controls. Six of these non-synonymous mutations have not been previously reported. Subsequent functional analyses revealed that the transcriptional activity, subcellular localization and DNA binding affinity of some mutant GATA4 proteins were significantly altered. Our results expand the spectrum ofGATA4mutations linked to cardiac defects. Together with the newly reported mutations, approximately 110 non-synonymous mutations have currently been identified inGATA4. Our future analysis will explore why the evolutionarily conserved GATA4 appears to be hypermutable.
Two out of the seven patientswith isolated up- per-limb anomalies had mutationsin TBX5 and/or GATA4. Patient HH2 had radioulnar synostosis of the left arm and hypoplasia of the distal and middle phalanges of the first and fifth digits (Figure 1F). At right, she had synostosis of ulna and humerus, a hypoplastic radius, and only two digi- tal rays of the right hand (the first-finger phalanges were hypoplastic, and only two metacarpal bones were ob- served), as shown in Figure F. No congenitalheartdisease was diagnosed in the patient or in her family (ECO and ECG were normal). Mutational analysis of the GATA4 gene showed two novel de novo intron mutations, c.[997+23A > T; 997+56C > A], in the same allele. These mutations were located far from the splicing sites, and the potential instabil- ity of the mRNA remains to be accessed. They were not found in the 100 control chromosomes tested. The patient also presented the TBX5 c.309C > T polymorphism in homozygosis.
ventricle undergoes a remodeling process, eventually leading to right ventricular failure. The right ventricular chamber dilates and the concentric hypertrophy transits to eccentric hypertrophy, resulting in increased wall stress and systolic dysfunction. Increased heart rate and right ventricular wall stress lead to significant increase of oxygen consumption in right ventricular myocardium. This, in combination with reduced right ventricular endomyocardial coronary perfusion (due to reduced right coronary artery pressure, increased right ventricular mass, and rising right ventricular end diastolic pressure), leads to right ventricular ischemia, and worsening right ventricular diastolic and systolic function. The right ventricular ischemia may be clinically manifested as chest pain. Subsequently the right ventricle and the tricuspid valve annulus dilate and functional tricuspid regurgitation will occur. 22-24 Most patientsin this study affected to pulmonary hypertension although the mean of pulmonary pressure was significantly higher in cyanotic patients compared with acyanotic patients (69±22 vs 39±13 mmHg) but GFR levels does not have any relationship to cyanosis (104.7 ± 50 vs 101.5 ± 32.5), in other word although pulmonary hypertension and hypoxia besides high hematocrit, are more severe in cyanotic groups but these will not guarantee to decrease GFR in childhood period.
The majority ofcongenitalheartdisease (CHD) is sporadic, with a minority of cases associated with a known genetic abnormality. Combinations of genetic- environmental factors are implicated in the etiology of the disease. Recently, several studies, using mostly animal models, unraveled that perturbations in the molecular processes that precede the beginning ofheart development might also be at the origin of CHD. In fact, some of the most complex CHDs are found associated with laterality defects, a disorder resulting from abnormal Left-Right axis formation. In our laboratory, the identified mouse Cerberus-like2 (Cerl2 – human DAND5), a protein that inhibit Nodal signaling, prompt us to study cardiac and laterality diseases, since the generated Cerl2 KO mice display a wide range of laterality defects and CHD. Considering the high conservation of genetic pathways regulating cardiac development in mouse and human, the main objective of the present thesis was the study of human genes involved in the Nodal pathway, focusing mostly in DAND5, in a CHD and/or laterality defects patients cohort. The sequence analysis of DAND5 revealed two patients displaying the same p.R152H variant, resulting in a substantial decreased in the function of the protein. We propose that p.R152H acts as a risk allele for CHD and/or laterality defects. In addition, we found two alterations in NODAL, two alterations in PITX2C and one alteration in CFC1. We hypothesized that the NODAL p.H165R variant can act as a common modifier and the intronic variants in NODAL and PITX2C might cause alterations in the splicing pattern of the mRNA molecules. Moreover, we generated patient-specific iPSCs to understand the molecular mechanisms ofdisease behind the DAND5 nucleotide variant. Although we cannot make a clearly genotype-phenotype correlation, the variants here identified probably increase the disease susceptibility due to the resulting abnormal Nodal signaling. Because most of the patients presented more than one alteration, the cumulative effect of each variant within the pathway seems to enhance even more these risk. Therefore, the imbalance in dosage-sensitive Nodal signaling is a common denominator for laterality defects and associated CHDs.
two patients each. Other heart defects in this group, included coarctation of the aorta, bicuspid aortic valve, aneurysm of the sinus of Valsalva and aortic stenosis/regurgitation, affecting one individual each. In the cyanotic CHD group, the most common lesion was tetralogy of Fallot (ToF), affecting seven individuals, followed by truncus arteriosus, double outlet right ventricle, transposition of the great arteries and double inlet left ventricle, affecting one patient each. All seven patientswith ToF under- went total surgical correction; one needed a Blalock-Thomas- Taussig shunt before total correction. The patient with trans- position of the great arteries underwent arterial switch (Jatene procedure). One of the patientswith pulmonary stenosis needed an aortic homograft. The two patientswith univentricular heart underwent Glenn procedure followed by Fontan operation.
Narod et al. speculated that gene mutationsin embryogenesis may be related to birth defects, cancer type, or both . The mechanism that explains the association between CHD and cancer remains elusive and it is not known whether the concurrence of CHD and various tumors is just a coincidence or whether they share common etiologic factors. Dysregulation dur- ing early human development may play an important role in cancer occurrence . Changes in cardiovascular structure or function, lifestyle adaptation related to CHD, or environmental exposures, may mediate the risk of cancer development among patientswith CHD. Results of the current study indicate that CHD and cancer may occur in the same individual, through some possible communal underlying factors. Further investigations are warranted to elucidate the as- sociation between CHD and malignancy, and to unravel the mechanisms that are involved.
ExAC database describes DAND5 p.R152H as a SNV, detected in the heterozygous state in more than 1200 in- dividuals, and present in 16 apparently normal homozy- gotes. Indeed, the mother of patient 2 is phenotypically normal but displays the DAND5 p.R152H variant in hererozygosity. Nevertheless, we cannot exclude the presence of a mild or undiagnosed heart defect on the mother of patient 2, or on the subjects of the genome projects, since detailed examination of these subjects is rarely available. In addition, the ExAC database and the American College of Medical Genetics and Genomics warns to the fact that in these population databases the presence of individuals with subclinical diseases among the supposed healthy individuals leading to minor dis- crepancies in some variant frequencies . For ex- ample, in a study conducted in Newcastle, 30% of the patients (198/669 infants) were undiagnosed at the time of discharge from the hospital, and in 30/198 the diag- nosis was only made after death . Moreover, some phenotypes of CHD such as small atrial septal defects or ventricular defects may be asymptomatic and undetected throughout life . Also, there are cases of apparently normal individuals whose laterality defects were only de- tected during adulthood . Despite that, differences in the overall level offunctional Nodal pathway might be explained by factors like differential allelic expressivity, an incomplete penetrance, the presence of genetic mo- difiers, and/or environmental factors. Interestingly, all Cerl2 heterozygous mice are phenotypically normal, and ~40% of the Cerl2 homozygous KO mice become
Human GATA4 maps to a region of chromosome 8 and consists of 7 exons encoding 442 amino acids. The functionalGATA4 protein comprises of two independent transcriptional activation domains (TAD) required for transcriptional activity, two highly conserved zinc fin- ger domains (ZnF) essential for DNA sequence identification and protein-DNA binding, and one nuclear localization signal (NLS) region associated with the subcellular trafficking and dis- tribution ofGATA4 . The GATA4 mutation, p.R311W, found in our study is identified in NLS region and may disrupt the normal subcellular localization ofGATA4. We performed immunofluorescence staining to determine whether the p.R311 was necessary for the nuclear import ofGATA4. The result showed that the mutant GATA4 protein translocated normally into the nucleus. Several mutationsin NLS region had been identified in CHD patients (S1 Table) and some of the mutant proteins still accumulated to the nuclei (S2 Fig) . All these findings suggested that not all the mutationsin NLS region were sufficient to inhibit GATA4 nuclear targeting. This was consistent with the results revealed by Philips, et al. 2007  that four amino acids Arg283, Arg284, Arg318, Arg320 were essential for GATA4 nuclear localiza- tion and proteins with individual mutated site in this region all showed normal nuclear distribution.
Group 2 comprised 12 patientswith different degrees of cardiac impairment. Most patients were infunctional classes I or II, because 58.4% had few symptoms, and, supposedly, had lower indices ofheart chamber dilation and, probably, lower BNP levels. It is worth noting that distension of the cardiac cavities stimulates the release of that peptide. If the study had comprised only patientswith the congestive form of chronic chagasic heartdisease, greater differences might have been observed between the 2 groups. On the other hand, the fact that the study comprised patientswith different degrees of cardiac involvement and in different stages of the disease allowed the inference of the importance of BNP not only inpatientswith congestive heart failure, but also in individuals without signs or symptoms, or both, of left ventricular dysfunction. The NYHA functional class was directly related to serum BNP levels, which is in accordance with the literature, except for func- tional class III, which did not show values greater than those infunctional class II. It is worth noting that only one individual was infunctional class III, which represents a bias in the sample. Ho- wever, if functional classes III and IV are combined, the BNP levels are statistically different from those infunctional classes I and II.
13. Konstantinov VV, Deev AD, Kapustina AV, et al. Prevalence of overweight and its relationship with mortality from cardiovascular and other chronic non-communicable diseases among the male population in cities of different regions. Кardiologiia 2002; 10: 45–54. Russian (Константинов В. В., Деев А. Д.,.Капустина А. В. и др. Распространенность избы- точной массы тела и ее связь со смертностью от сердеч- но-сосудистых и других хронических неинфекционных за- болеваний среди мужского населения в городах различных регионов. Кардиология 2002; 10: 45–54).
consensus in the literature regarding this issue could be ex- plained by the observations that clinical manifestations of WD are heterogeneous even inpatients carrying the same mutation. It is, therefore, hypothesized that other addition- al genetic and/or environmental factors could inluence the phenotypes of WD, such as dietary cooper intake, metallo- thionein inducibility, the individual capacity to handle cop- per overload, the ApoE genotype, human prion gene poly- morphism and mutationsin COMMD1 6,17,19-22 .
A eficácia da profilaxia nunca foi comprovada por estudos randomizados. O tema tem despertado muitas discussões e continua controverso [3-7]. Atualmente, a American Heart Association (AHA) recomenda profilaxia antibiótica apenas para pacientes considerados de alto risco para EI. Estão incluídos nesse grupo pacientes portadores de cardiopatias congênitas cianóticas não corrigidas ou com correções paliativas, e pacientes que se submeteram a correção cirúrgica da cardiopatia, mas que persistem com lesões residuais adjacentes a material prostético. Esses critérios incluem grande parte da população pediátrica portadora de cardiopatias. Medidas profiláticas adicionais incluem educação dos pacientes de alto risco e de seus cuidadores sobre a necessidade de higiene oral e cutânea meticulosa, visitas frequentes ao dentista, bem como evitar procedimentos invasivos desnecessários.
eral population: ACEI, ARBs, βblockers, inotropic drugs and diuretics. Hypervolemia and most of the related symptoms can be kept under control effectively by the isolated or ultra filtation, in conjunction with dialysis, during the standard bicar bonate hemodialysis or hemodiafiltration. In the same respect peritoneal dialysis is efficient for the control of hypervolemia symptoms, mainly during the first years of its application and in case of the lower NYHA class (II°/III°). In general, heart support therapy, surgical interventions of the myocardium and valve replacement are rarely used inpatients on dialysis, whereas revascularization procedures are beneficial for associated IHD. In selected cases the application of cardiac resynchronization and/or implantation of a cardioverter defibrillator are advisable. Keywords: congestive heart failure; chronic renal failure; car diovascular pharmacotherapy
Felcar et al.  performed a study with 141 children withcongenitalheartdisease, aged varying between one day old to six years, randomly divided into two groups, whereas one of them received physiotherapy in the pre- and postoperative and the other only postoperatively. The study obtained statistically significant difference regarding the presence of pulmonary complications (pneumonia and atelectasis), being more frequent in the group undergoing physiotherapy only postoperatively. Moreover, when the presence of pulmonary complications was associated with other complications regarding the time of hospital stay, such as sepsis, pneumothorax, pleural effusion and others, surgery associated with cardiopulmonary bypass (CPB)
with CHD and a control group in France. The study inclu- ded fetuses and stillbirths. The diagnosis ofheart defect was conirmed by echocardiogram, cardiac catheterization, surgery or autopsy reports. Among the reported cases, 25.7% showed at least one EM, and such abnormalities were ten times more frequent in this group than in the control group. Among individuals withheart defects, 11.5% exhibited some type of syndrome. The most frequent EM affected the kidneys (21.4%) and the digestive system (19.6%). The most frequent abnormalities of the upper renal system were the urethral anomalies, hydronephrosis and unilateral renal agenesis. The presence of olygohydramnios and polyhydra- mnios and threatened miscarriage (the occurrence of vaginal bleeding before fetal viability) were more frequent among children with CHD and concomitant EM or syndromes.
The randomization was done in two steps: by first generating numbers (using a table of random numbers) and by then concealing the patients’ assignments using opaque sealed envelopes. After signing an Informed Consent, an envelope was opened by an independent professional who declared in which group the patient would be included (I or II). Group I received pre- and postoperative physiotherapy (intervention) and group II received only postoperative physiotherapy (control).
So far, there are 70 different mutations described in the scientific literature and 104 different families reported with one of these mutations (families and mutations reported in this study were not taken into account). In these 70 mutations, 60 correspond to the substitution of a single nucleotide, 3 correspond to the substitution of 2 nucleotides, 2 correspond to the deletion of a single nucleotide, 3 correspond to the deletion of 3 nucleotides and 1 mutation corresponds to an almost complete deletion of the AVP gene (with the exception of exon 1). For one mutation it was not possible to determine the exact nucleotide change due to insufficient data. According to these variants, 54 mutations cause missense changes, 2 mutations cause deletion of a single amino acid residue, 3 mutations cause deletion of 4 amino acids residues, 7 mutations lead to the insertion of a premature stop codon (nonsense), 1 mutation causes a frameshift, 1 mutation causes an indel (insertion/deletion) and two mutations have an unknown effect at protein (p.?). Several families shared the same mutations; the most relevant was the c.55G>A mutation, corresponding to 8% of studied families. Regarding the type of alteration at the protein level, Fig. 10 presents their distribution between all reported families (families reported in this study were not taken into account).
Results: The prevalence ofcongenitalheartdiseasein this study was 5/1000 live-births (339 live-births, 20% with critical defects). The most common defects were ventricular septal defect (38%), atrial septal defect (15%), aortic coarctation (7%), tetralogy of Fallot (7%) and pulmonary stenosis (5%). One third of the live births had antenatal diagnosis ofcongenitalheartdisease. In the live-births with critical congenitalheartdisease, 54% had antenatal diagnosis and 14% were diagnosed at birth. There were records of 84 pregnancy terminations; 49% had critical defects, 75% had non-cardiac congenital anomalies and 40% had chromosomal abnormalities. There were 15 postnatal deaths recorded (3.4% mortality rate), associated with prematurity/low birthweight, critical congenitalheartdisease, other non-cardiac congenital anomalies and chromosomal abnormalities.
device was studied. The study took into account the effects of machine scale, wear surface structure of the rolls, grinding pressures and rolls speed, gap settings, feed size distribution and moisture content for a range of ores. The authors proposed a prevailing wear mechanism and a methodology for minimising wear of the grinding rolls, specific to the high pressure grinding device only. An example of a direct method, Bond (1964) and Buchi (1995) developed testing apparatus that determine rock abrasiveness in a low abrasion/medium impact mode of wear where rock abrasiveness is measured as the amount of material lost by a standard steel paddle which rotates on a shaft in a sample of loose rock particles of a certain specified size range. As can be seen from the above examples, the indirect methods of rock abrasivity assessment have the advantage of using data which is either readily available or relatively straightforward to obtain. However, they do not take into account process variables for specific modes of wear. Hence, they are normally not used in isolation, but rather in combination with direct methods, or holistic approaches, to supplement or confirm other more relevant direct measures. However, there is no universally accepted one standard test to determine the rock abrasivity although a large number of different tests are in use. All the studies about rock abrasiveness are concentrated on the amount of quartz, grain size and cementation degree of quartz, the geometry of the abrasive mineral and mechanical strength of rock.
to determine protein content. Tissue lysate (100–200μg) was subjected to SDS–polyacrylamide gel electrophoresis, and protein was subsequently transferred to a PVDF membrane (Whatman Westran, UK). The membrane was pre-incubated in a buffer containing 10mM Tris (pH 7.4), 150mM NaCl, and 0.1% Tween supplemented with 5% skim milk for 1h at room temperature. Following pre-incubation, the membrane was incubated in the Tris–NaCl–Tween buffer supplemented with 5% bovine serum albumin and antibody for 1–2h at room temperature. Antibodies employed were a mouse monoclonal antibody against cytochrome c oxidase I, a rabbit polyclonal antibody against nNOS, a rabbit polyclonal antibody against iNOS, a rabbit polyclonal antibody against eNOS(all of them 1:1000, Santa Cruz, USA) and a rabbit polyclonal antibody against β-actin(1:1,000; Biosynthesis Biotechnology, Beijing, China). Following incubation, the membrane was washed with buffer for 1h, and re-probed with the appropriate secondary antibody conjugated to horseradish peroxidase (1:1,000; Biosynthesis Biotechnology, Beijing, China) in Tris–NaCl–Tween buffer supplemented with 5% skim milk for 1h at room temperature. Following incubation, the membrane was washed three times with the Tris–NaCl– Tween buffer and bands visualized by an ECL detection kit (ShineGene, Shanghai, China). Films were scanned with a laser densitometer utilizing the program Quantity One (Bio- Rad Laboratories, Canada). Relative levels of protein were standardized using β-actin as the internal control.