Comparison of Immune Profiles in Fetal Hearts with Idiopathic Dilated Cardiomyopathy, Maternal Autoimmune-Associated Dilated Cardiomyopathy and the Normal Fetus.

The etiology of idiopathic dilated cardiomyopathy (iDCM) remains unknown. Immune therapies have improved outcome in fetuses with DCM born to mothers with autoimmune disease (aDCM). The purpose of this retrospective study was to compare the myocardial B and T cell profiles in fetuses and neonates with idiopathic DCM (iDCM) versus autoimmune-mediated DCM (aDCM) and to describe the normal cell maturation within the human fetal myocardium. Of 60 fetal autopsy cases identified from institutional databases, 10 had aDCM (18-38 weeks), 12 iDCM (19-37 weeks) and 38 had normal hearts (11-40 weeks). Paraffin-embedded myocardium sections were stained for all lymphocyte (CD45), B cells (CD20, CD79a), T cells (CD3, CD4, CD7, CD8) and monocyte (CD68) surface markers. Two independent, blinded cell counts were performed. Normal hearts expressed all B and T cell markers in a bimodal fashion, with peaks at 22 and 37 weeks of gestation. The aDCM cohort was most distinct from normal hearts, with less overall T cell markers [EST -9.1 (2.6) cells/mm(2), p = 0.001], CD4 [EST -2.0 (0.6), p = 0.001], CD3 [EST -3.9 (1.0), p < 0.001], CD7 [EST -3.0 (1.1), p = 0.01] overall B cell markers [EST -4.9 (1.8), p = 0.01] and CD79a counts [EST -2.3 (0.9), p = 0.01]. The iDCM group had less overall B cell markers [EST -4.0 (1.8), p = 0.03] and CD79a [EST -1.7 (0.9), p = 0.05], but no difference in T cell markers. Autoimmune-mediated DCM fetuses have less B and T cell markers, whereas iDCM fetuses have less B cell markers compared with normal fetal hearts. The fetal immune system may play a role in the normal development of the heart and evolution of dilated cardiomyopathy.

Combined hereditary and somatic mutations of replication error repair genes result in rapid onset of ultra-hypermutated cancers.

DNA replication-associated mutations are repaired by two components: polymerase proofreading and mismatch repair. The mutation consequences of disruption to both repair components in humans are not well studied. We sequenced cancer genomes from children with inherited biallelic mismatch repair deficiency (bMMRD). High-grade bMMRD brain tumors exhibited massive numbers of substitution mutations (>250/Mb), which was greater than all childhood and most cancers (>7,000 analyzed). All ultra-hypermutated bMMRD cancers acquired early somatic driver mutations in DNA polymerase ɛ or δ. The ensuing mutation signatures and numbers are unique and diagnostic of childhood germ-line bMMRD (P < 10(-13)). Sequential tumor biopsy analysis revealed that bMMRD/polymerase-mutant cancers rapidly amass an excess of simultaneous mutations (∼600 mutations/cell division), reaching but not exceeding ∼20,000 exonic mutations in <6 months. This implies a threshold compatible with cancer-cell survival. We suggest a new mechanism of cancer progression in which mutations develop in a rapid burst after ablation of replication repair.

Undiagnosed heart disease leading to sudden unexpected death in childhood: a retrospective study.

OBJECTIVES: Heart disease accounts for a significant proportion of sudden unexpected deaths among children. We describe here demographic features, pathological conditions, and the frequency of premonitory symptoms in a retrospective series of cases of sudden unexpected cardiac death (SUCD) attributable to undiagnosed structural heart disease. METHODS: A chart review of autopsies involving children 0 to 17 years of age that were performed at the Hospital for Sick Children (Toronto, Ontario, Canada) between 1984 and 2003 was conducted. Cases of sudden unexpected death within 24 hours after clinical presentation with previously undetected fatal heart disease were included. Cases with multiple or thoracic trauma and chronic or multisystem disease were excluded. RESULTS: During the 20-year study period, 4926 autopsies were performed. A total of 103 cases (2.1%), involving 51 male patients and 52 female patients 1 day to 15 years of age (mean: 2.9 ± 4.2 years), were diagnosed as having SUCD. The most common diagnoses were myocarditis (n = 37 [35.9%]), hypoplastic left heart syndrome (HLHS) (n = 19 [18.4%]), dilated cardiomyopathy (DCM) (n = 16 [16.5%]), coronary artery anomalies (n = 6 [5.8%]), and aortic stenosis (n = 5 [4.9%]). There was a significant difference in the mean age of presentation between leading causes of SUCD (6.5 days for HLHS, 1.7 years for DCM, and 5.4 years for myocarditis; P < .0001). Of 103 cases, 27 (26.2%) had premonitory symptoms documented. CONCLUSION: SUCD accounted for 2.1% of all autopsies, and HLHS, DCM, and myocarditis were the 3 most common diagnoses, which presented at increasing ages.

The myocardium of fetuses with endocardial fibroelastosis contains fewer B and T lymphocytes than normal control myocardium.

The observation that endocardial fibroelastosis (EFE) can result from an immune response to maternal autoantibody deposition in the fetal myocardium raises the possibility that the fetal immune system may contribute to the pathogenesis of idiopathic EFE and dilated cardiomyopathy (DCM). This study sought to characterize myocardial immune cell presence in fetuses and neonates with idiopathic EFE + DCM, in those with EFE + structural heart disease, and in normal control subjects. Paraffin tissue sections from fetuses identified from the pathology database were stained for B cell, T cell, macrophage, and general hematopoietic cell surface markers. Of the 14 fetuses included in the study, 5 had EFE + DCM, 4 had EFE + structural heart disease, and 5 were normal control fetuses. The EFE + DCM group had fewer B cells than the control group (0.15 vs. 0.44 cells/mm(2); p = 0.005). The EFE + heart disease group had both fewer B cells (0.18 vs. 0.44 cells/mm(2); p = 0.08) and T cells (0.29 vs. 0.80 cells/mm(2); p = 0.04) than the control group. The CD4/CD8 ratio was similar in the EFE + DCM and EFE + heart disease groups (1.0 vs. 0.9; p = 0.17) but higher in the EFE + DCM group than in the control group (0.9 vs. 0.3; p = 0.03). The myocardium of fetuses with EFE contains fewer B and T lymphocytes than normal control fetuses.

Mitogenic cardiomyopathy: a lethal neonatal familial dilated cardiomyopathy characterized by myocyte hyperplasia and proliferation.

Pediatric cardiomyopathies are a heterogenous group of conditions of which dilated cardiomyopathies are the most common clinicomorphologic subtype. However, the etiology and pathogenesis of many cases of dilated cardiomyopathies remain unknown. We describe a series of 5 cases of a rare but clinically and histologically distinctive dilated cardiomyopathy that was uniformly lethal in early infancy. The 5 cases include 2 pairs of siblings. There was parental consanguinity in 1 of the 2 pairs of siblings. Death occurred in early infancy (range, 22-67 days; mean, 42 days) after a short history of general lethargy, decreased feeding, respiratory distress, or cyanosis. There was no specific birth or early neonatal problems. Autopsy revealed congestive cardiac failure and enlarged, dilated hearts with ventricular dilatation more pronounced than atrial dilatation, and endocardial fibroelastosis. Histology showed prominent hypertrophic nuclear changes of cardiac myofibers and markedly increased myocyte mitotic activity including occasional atypical mitoses. Immunohistochemical staining for Mib1 showed a markedly increased proliferative index of 10% to 20%. Ancillary investigations, including molecular studies, did not reveal a primary cause for the cardiomyopathies. This distinctive dilated cardiomyopathy characterized by unusual histologic features of myocyte nuclear hypertrophy and marked mitotic activity is lethal in early infancy. Its occurrence in 2 pairs of siblings suggests familial inheritance. Although the underlying molecular pathogenesis remains to be elucidated, it is important to recognize this distinctive entity for purposes of genetic counseling.

Abnormal connexin43 in arrhythmogenic right ventricular cardiomyopathy caused by plakophilin-2 mutations.

Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a disorder of cardiomyocyte intercalated disk proteins causing sudden death. Heterozygous mutations of the desmosomal protein plakophilin-2 (PKP-2) are the commonest genetic cause of ARVC. Abnormal gap junction connexin43 expression has been reported in autosomal dominant forms of ARVC (Naxos and Carvajal disease) caused by homozygous mutations of desmosomal plakoglobin and desmoplakin. In tissue culture, suppression of PKP-2 results in decreased expression of connexin43. We sought to characterize the expression and localization of connexin43 in patients with ARVC secondary to heterozygous PKP-2 mutations. Complete PKP-2 gene sequencing of 27 ARVC patients was utilized to identify mutant genotypes. Endomyocardial biopsies of identified carriers were then assessed by immunofluorescence to visualize intercalated disk proteins. N-cadherin was targeted to highlight intercalated disks, followed by counterstaining for PKP-2 or connexin43 using confocal double immunofluorescence microscopy. Immunofluorescence was quantified using an AdobeA Photoshop protocol, and colocalization coefficients were determined. PKP-2 siRNA experiments were performed in mouse cardiomyocyte (HL1) cell culture with Western blot analysis to assess connexin43 expression following PKP-2 suppression. Missense and frameshift mutations of the PKP-2 gene were found in four patients with biopsy material available for analysis. Immunofluorescent studies showed PKP-2 localization to the intercalated disk despite mutations, but associated with decreased connexin43 expression and abnormal colocalization. PKP-2 siRNA in HL1 culture confirmed decreased connexin43 expression. Reduced connexin43 expression and localization to the intercalated disk occurs in heterozygous human PKP-2 mutations, potentially explaining the delayed conduction and propensity to develop arrhythmias seen in this disease.

Scurvy diagnosed in a pediatric liver transplant patient awaiting combined kidney and liver retransplantation.

First described in the 1500 s, scurvy is infrequently seen in industrialized countries today, although vulnerable patient groups remain. A 15-yr-old girl underwent liver transplantation at age 26 months for a primary diagnosis of biliary hypoplasia, and subsequently developed late allograft failure and progressive renal insufficiency culminating in listing for combined liver retransplantation and kidney transplantation at age 13 yr. She required regular hemodialysis treatment for 12 months prior to deceased donor organ availability, with a complicated clinical course including recurrent septic episodes and severe cachexia. Ten months after initiation of hemodialysis, she presented with severe bone pain, purpura, ecchymoses, gingival hyperplasia, mucosal bleeding, and subconjunctival hemorrhages. Serial serum ascorbic acid levels were found to be extremely low (<10 micromol/L) despite routine supplementation both in her dialysate and via regular oral supplementation. Histopathology from skin biopsy revealed purpura, hyper- and parakeratosis, and follicular plugging. She had ECG and 2D echocardiogram disturbances, as well as osteopenia and sclerosis of the extremities on radiological evaluations. Therapy with high-dose ascorbic acid (1 g/day orally) led to complete resolution of skin lesions. This case highlights the importance of awareness and recognition of this historic diagnosis, and particularly in children with end-stage organ disease with severely compromised nutrition.

Decreased left ventricular coronary artery density in pulmonary atresia and intact ventricular septum.

BACKGROUND: The prognosis for pulmonary atresia and intact ventricular septum (PAIVS) has been poor. Our hypothesis is that intrinsic abnormal left ventricular (LV) intramyocardial circulation might be related to the poor outcomes of these patients. METHODS: Neonatal heart specimens were examined microscopically in four groups of 6 cases each. Group I had PAIVS with ventriculocoronary artery connections (VCAC), group II had PAIVS without VCAC, group III had normal hearts, and group IV had LV hypertrophy. A projection microscope with grid overlay was used to count the LV intramyocardial coronary artery density (IMCAD), which was expressed as the number of profiles/mm(2). RESULTS: The LV IMCAD of groups I (0.40 +/- 0.14/mm(2)) and II (0.45 +/- 0.15/mm(2)) were significantly lower than those of groups III (0.77 +/- 0.11/mm(2)) and IV (0.76 +/- 0.09/mm(2); all with p = 0.002). There was no significant difference between either groups I and II (p = 0.394) or groups III and IV (p = 0.818). CONCLUSIONS: This study demonstrates lower LV IMCAD in a widely heterogeneous spectrum of neonatal hearts with PAIVS, which might potentially predispose these patients to myocardial ischemia and in turn contribute to the poor prognosis of this disease.

Presence of periorbital and conjunctival petechial hemorrhages in accidental pediatric drowning.

The pathological findings of drowning are variable and non-specific. Petechial hemorrhages involving the periorbital region and the conjunctiva have been described in many causes of death, but are thought to be exceedingly uncommon in cases of drowning. However, such studies have not specifically addressed the pediatric population. The current study retrospectively examined 79 cases of accidental pediatric drowning for the presence of periorbital/conjunctival hemorrhages and analyzed factors that may have affected their presence. Ten victims had periorbital/conjunctival petechial hemorrhages (13%), with five having periorbital petechiae, three having conjunctival petechiae, and two having both periorbital and conjunctival petechiae. The age and gender of the victim, site of drowning, resuscitation history and the presence of other pathological findings were not significantly associated with the presence of periorbital/conjunctival petechiae. However, as the interval between the drowning episode and autopsy increased, the incidence of periorbital/conjunctival petechiae decreased (28% for <24h; 7% for >24h). The presence of periorbital/conjunctival hemorrhages in a significant proportion of pediatric drowning victims confirms that the pathologist must add this finding to the spectrum of changes seen in pediatric drowning.

Piloting a novel porcine model for endolaryngeal injury following prolonged intubation.

OBJECTIVES: (1) To develop a practical animal model of endolaryngeal damage secondary to prolonged endotracheal intubation. (2) To demonstrate the contribution of chronic hypoxia to laryngeal injury in the context of this model. METHODS: Four Sus scrofa piglets were anaesthetized and intubated for 24h. Two animals were maintained in a state of constant hypoxia (pO(2)<60 mmHg) while two others were ventilated with 100% oxygen. Prior to sacrifice, fluorescein dye was infused intravenously to highlight areas of hypoperfusion. The larynx and trachea were then harvested for gross and histological examination. RESULTS: All four specimens demonstrated areas of edema, erythema, and ulceration on gross examination. Areas of significant histological inflammation, ulceration, and necrosis involved tube-mucosa contact, in particular, the arytenoids, the interarytenoid area, and the subglottis. CONCLUSIONS: This animal model represents a practical and novel means for the investigation and treatment of laryngeal injury secondary to prolonged endotracheal intubation. Significant injury to the endolarynx was evident after only 24h of intubation and the injury involved similar areas within the larynx as described in human studies. Although clinical experience suggests that chronic hypoxia is a risk factor for endolaryngeal injury, this model did not provide experimental evidence to support this observation, most likely due to the small study size.

Sudden unexpected death in infancy and childhood due to undiagnosed neoplasia: an autopsy study.

Sudden unexpected death due to clinically undiagnosed neoplasia in infancy and childhood (SUDNIC) is a rare phenomenon, with only small numbers of cases reported in the literature. In the majority of instances, the tumors involve critical structures within the heart or central nervous system and include gliomas, medulloblastomas, rhabdomyomas, and neoplasms of stromal elements. A 20-year retrospective review of autopsy records from the Hospital for Sick Children, Toronto, was performed (1984-2003, n = 4926), and 7 cases of SUDNIC were identified (0.14%). In addition, 1 case was obtained from the files of the Children's and Women's Health Centre of British Columbia, Vancouver. Diagnoses included 2 cases of acute leukemia (1 myelogenous, 1 lymphoblastic), 2 cases of mediastinal lymphoblastic lymphoma (pre-T cell type), 1 papillary fibroelastoma of the mitral valve prolapsing into and totally occluding the left anterior descending coronary artery, 1 medulloblastoma, 1 Wilms tumor associated with fatal intraperitoneal hemorrhage, and 1 widely disseminated gastric carcinoma. These cases demonstrate that infants and children may have minimal or no symptoms in the presence of significant disease and highlight the need for a thorough autopsy examination in cases of sudden unexpected death in infancy and childhood.

Association of drowning and myocarditis in a pediatric population: an autopsy-based study.

CONTEXT: Drowning is a frequent cause of accidental death in childhood, but the association of myocarditis and drowning has only rarely been reported. OBJECTIVE: To report 5 cases of drowning in children with coexistent myocarditis. DESIGN: A retrospective review of autopsy records of patients 0 years to 18 years of age was performed during a 6-year period (1998-2003, total cases reviewed = 1431). RESULTS: Twenty-two drownings were identified, in 14 male and 8 female children. Five patients (23%), 3 female and 2 male children, had coexistent myocarditis. The 5 patients ranged in age from 23 months to 13 years (mean, 7 years 2 months). None of the patients had antecedent symptomatology suggestive of myocarditis. In all patients, the myocarditis was focal mild or moderate, and the inflammatory infiltrate comprised lymphocytes with smaller numbers of neutrophils. All 5 patients had foci of myocyte necrosis. One patient had histologic evidence of myocardial hypertrophy but no evidence of a cardiomyopathy. Microbiologic studies, including culture, immunohistochemistry, polymerase chain reaction, and reverse transcriptase polymerase chain reaction, revealed Mycoplasma pneumoniae DNA in 1 case. CONCLUSIONS: The finding of myocarditis in a significant proportion of drowning victims in this series highlights the importance of a thorough autopsy examination in apparently straightforward cases and has clinicopathologic significance.

Abnormal pericyte recruitment as a cause for pulmonary hypertension in Adams-Oliver syndrome.

Adams-Oliver syndrome (AOS) consists of congenital scalp defects with variable limb defects of unknown pathogenesis. We report on two children with AOS plus additional features including intrauterine growth retardation (IUGR), cutis marmorata telangiectatica congenita (CMTC), pulmonary hypertension (PH), intracranial densities shown in one case to be sites of active bleeding and osteopenia. Autopsy in one case revealed defective vascular smooth muscle cell/pericyte coverage of the vasculature associated with two blood vessel abnormalities. Pericyte absence correlated with vessel dilatation while hyperproliferation of pericytes correlated with vessel stenosis. These findings suggest a unifying pathogenic mechanism for the abnormalities seen in AOS. These and previously reported cases establish that a subset of AOS patients is at high risk for PH.

Intrascrotal lipoblastoma with a complex karyotype: a case report and review of the literature.

Lipoblastoma is a tumor of adipose tissue that usually occurs in young children. Most lipoblastomas occur on the extremities, trunk, and head and neck, and most have rearrangements of the 8q region. We describe a lipoblastoma in a 12-month-old boy who presented with a rapidly enlarging scrotal mass. Electron microscopy revealed features consistent with immature adipocytes, and cytogenetic analysis revealed the following karyotype: 57,XY,+4,+6,+7,der(8)t(8;12) (q22;q13), +der(8)t(8;12) (q22;q13), +9,+10,+12,-16,+17,+der(18)t(8;18)(q22;q23),+19,+20. Interestingly, the breakpoint on chromosome 12 (q13) is the same as that seen in lipoblastomas. To our knowledge, this is the first reported case of such a complex karyotype in lipoblastoma and adds to the expanding list of karyotypic abnormalities seen in such tumors.

Supravalvar aortic stenosis with supravalvar pulmonary stenosis and peripheral vascular stenoses.

A non-dysmorphic 10 month old female was discovered at surgery to have severe vasculopathy of both the systemic and pulmonary arteries. These findings were confirmed by pathologic examination. Follow-up angiography has confirmed multiple sites of vascular obstruction which appear to be worsening. Angioplasty has only partially relieved these obstructions. The pathology and possible etiology are reviewed.

Endocardial fibroelastosis associated with maternal anti-Ro and anti-La antibodies in the absence of atrioventricular block.

OBJECTIVES: This study was designed to document the association of endocardial fibroelastosis (EFE) and maternal autoantibodies. BACKGROUND: Neonatal lupus erythematosus is associated with the transplacental passage of maternal anti-Ro and anti-La antibodies, leading to complete atrioventricular block (CAVB). In some cases, CAVB is associated with EFE. Isolated EFE may be independently related to maternal anti-Ro and anti-La antibodies. METHODS: We identified three cases (one fetus and two infants, all female) of isolated EFE in infants born to autoantibody-positive mothers in the absence of CAVB. Demographics, echocardiograms, and pathology were reviewed. Immunohistochemical analyses for immunoglobulin (Ig)G, IgM, IgA, T-cell, B-cell, and terminal deoxynucleoleotidyl transferase-mediated dUTP-biotin nick end-labeling (TUNEL) (test for cell apoptosis) staining were performed on multiple sections of the heart in each case and compared with negative controls. RESULTS: Two cases died and one received a cardiac transplant. All three cases had histologically confirmed EFE. All cases demonstrated significant diffuse IgG infiltration. To a lesser extent, myocardial tissue was also positive for IgM, CD43, and Granzyme B. None of the specimens were TUNEL positive. CONCLUSIONS: These are the first reported cases of isolated EFE associated with maternal anti-Ro and anti-La antibodies in the absence of CAVB. The diffuse deposition of IgG and the presence of a T-cell infiltrate throughout the myocardium suggest that the transplacental passage of maternal autoantibodies induces an immune reaction within the myocardium, leading to isolated EFE. Autoantibody-mediated EFE may be an etiologic factor in cases of fetal and neonatal "idiopathic" dilated cardiomyopathy.

Fetal cardiomyopathies: pathogenic mechanisms, hemodynamic findings, and clinical outcome.

BACKGROUND: Although the prenatal diagnosis of most fetal structural heart defects and dysrhythmias has been described, there is a paucity of information about cardiomyopathies (CMs) in prenatal life. METHODS AND RESULTS: To determine the pathogenic mechanisms, hemodynamic findings, and outcome of fetal CM, we reviewed the fetal echocardiograms and perinatal histories of 55 affected fetuses. Dilated CM was diagnosed in 22 cases, including 2 with congenital infections, 5 familial cases, 6 with endocardial fibroelastosis related to maternal anti-Ro/La antibodies, and 9 idiopathic cases. Thirty-three had hypertrophic CM, 7 associated with maternal diabetes, 2 with Noonan's syndrome, 2 with alpha-thalassemia, 18 with twin-twin transfusion syndrome, 1 with familial hypertrophy, and 3 with idiopathic hypertrophy. Systolic dysfunction was present in all cases of dilated CM and 15 cases of hypertrophic CM. Diastolic dysfunction was present in 19 of 30 fetuses with assessment of diastolic function parameters. Significant mitral or tricuspid valve regurgitation was seen in 32 cases. Eight fetuses were hydropic and 23 had signs of early hydrops. Seven pregnancies were terminated. Of 46 continued pregnancies with follow-up, 29 (63%) died perinatally. The presence of systolic dysfunction, diastolic dysfunction, and significant atrioventricular valve regurgitation were identified as risk factors for mortality. By multiple logistic regression, diastolic dysfunction was associated with an 8-fold increased risk relative to the other parameters. CONCLUSIONS: Fetal CM has a broad spectrum of intrinsic and extrinsic causes. A poor outcome is observed in many affected fetuses. Diastolic dysfunction in fetal CM is associated with the highest risk of mortality.

Maternal anti-Ro and anti-La antibody-associated endocardial fibroelastosis.

BACKGROUND: Maternal anti-Ro and anti-La antibodies are associated with congenital heart block (CHB). Although endocardial fibroelastosis (EFE) has been described in isolated cases of autoantibody-mediated CHB, the natural history and pathogenesis of this disease are poorly understood. METHODS AND RESULTS: We retrospectively reviewed the clinical history, echocardiography, and pathology of fetuses and children with EFE associated with CHB born to mothers positive for anti-Ro or anti-La antibodies at 5 centers. Thirteen patients were identified, 6 with a prenatal and 7 with a postnatal diagnosis. Six mothers were positive for anti-Ro and anti-La antibodies, and 7 were positive for anti-Ro antibodies only. Only 1 mother had autoimmune disease. Severe ventricular dysfunction was seen in all fetal and postnatal cases. Four fetal and 3 postnatal cases had EFE at initial presentation. However, 2 fetal and 4 postnatal cases developed EFE 6 to 12 weeks and 7 months to 5 years from CHB diagnosis, respectively, even despite ventricular pacing in 6 postnatal cases. Eleven (85%) either died (n=9) or underwent cardiac transplantation (n=2) secondary to the EFE. Pathologic assessment of the explanted heart, available in 10 cases, revealed moderate to severe EFE in 7 and mild EFE in 3 cases, predominantly involving the left ventricle. Immunohistochemistry in 4 cases (including 3 fetuses) demonstrated deposition of IgG in 4 and IgM in 3 and T-cell infiltrates in 3 cases, suggesting an immune response by the affected fetus or child. CONCLUSIONS: EFE occurs in the presence of autoantibody-mediated CHB despite adequate ventricular pacing. Autoantibody-associated EFE has a very high mortality rate, whether developing in fetal or postnatal life.