Jean M. Silvestri

Idiopathic congenital central hypoventilation syndrome: analysis of genes pertinent to early autonomic nervous system embryologic development and identification of mutations in PHOX2b.

Idiopathic congenital central hypoventilation syndrome (CCHS) has been linked to autonomic nervous system dysregulation and/or dysfunction (ANSD) since it was first described in 1970. A genetic basis of CCHS has been proposed because of the reports of four families with two affected children, because of mother–child transmission, and because of a recent report of a polyalanine expansion mutation in PHOX2b in a subset of CCHS subjects. We, therefore, studied genes pertinent to early embryologic development of the ANS including mammalian achaete‐scute homolog‐1 (MASH1), bone morphogenic protein‐2 (BMP2), engrailed‐1 (EN1), TLX3, endothelin converting enzyme‐1 (ECE1), endothelin‐1 (EDN1), PHOX2a, and PHOX2b in 67 probands with CCHS, and gender‐ and ethnicity‐matched controls. No disease‐defining mutations were identified in MASH1, BMP2, EN1, TLX3, ECE1, EDN1, or PHOX2a.The 65/67 CCHS probands (97%) were found to be heterozygous for the exon 3 polyalanine expansion mutation identified previously in PHOX2b. Further, there was an association between repeat mutation length and severity of the CCHS/ANSD phenotype. Of the two probands who did not carry the expansion mutation, one had a nonsense mutation in exon 3 which truncated the protein and the other had no mutation in PHOX2b but had a previously reported EDN3 frameshift point mutation. The polyalanine expansion mutation was not found in any of 67 matched controls. Of 54 available families (including 97 unaffected parents), whose child carried the PHOX2b mutation, 4 parents demonstrated mosaicism for an expansion mutation identical to that seen in the CCHS cases, suggesting that not all mutations in affected probands with unaffected parents are de novo. We also studied four women with CCHS who were heterozygous for the PHOX2b mutation, each with one child. Three of the four children were also affected and had the same mutation, demonstrating autosomal dominant inheritance of the mutation. Assay of the PHOX2b polyalanine repeat mutation represents a highly sensitive and specific technique for confirming the diagnosis of CCHS. Identification of the CCHS mutation will lead to clarification of the phenotype, allow for prenatal diagnosis for parents of CCHS probands and adults with CCHS in future pregnancies, and potentially direct intervention strategies for the treatment of CCHS.

Congenital central hypoventilation syndrome: Diagnosis, management, and long-term outcome in thirty-two children

The diagnosis, management, and long-term outcome of 32 patients with congenital central hypoventilation syndrome are summarized. Sleep hypoventilation was severe in all cases, resulting in an alveolar carbon dioxide pressure (mean +/- SEM) of 62 +/- 2.5 mm Hg and a hemoglobin saturation of 65% +/- 3.3% without ventilatory or arousal response. Awake hypoventilation on initial assessment was present in 12 of the 32 patients, resulting in an alveolar carbon dioxide pressure of 58 +/- 2.2 mm Hg and a hemoglobin saturation of 59% +/- 7%. Associated conditions included pulmonary hypertension or cor pulmonale or both (78%), heart block and sick sinus syndrome requiring a cardiac pacemaker (two patients), mild atrophy by cranial imaging evidence (40%), seizures (72%), normal brain-stem auditory evoked responses in all but one patient tested, ganglioneuroblastomas (one patient), Hirschsprung disease (16%), and ophthalmologic abnormalities (60%). Growth was deficient in 44% of patients; hypotonia or major motor delay or both were apparent in all. Twenty-two patients are living; 12 of them require continuous ventilatory support and 10 breathe spontaneously while awake and require ventilatory support while asleep. Ten patients have died. Autopsy performed in six cases indicated diffuse central nervous system astrocytosis, gliosis, and atrophy but no primary brain-stem abnormality. Although these data support a diffuse central nervous system process, the specific cause and the mode of inheritance remain unclear. With early diagnosis and careful ventilatory management, the sequelae of hypoxia and morbidity should be minimized and long-term outcome improved.

Sudden infant death syndrome: Association with a promoter polymorphism of the serotonin transporter gene

Serotonergic receptor binding in the arcuate nucleus, n. raphé obscurus, and other medullary regions is decreased in sudden infant death syndrome (SIDS) cases. Further, a variable tandem repeat sequence polymorphism in the promoter region of the serotonin transporter protein (5‐HTT) gene has recently been associated with risk of SIDS in a Japanese cohort. This polymorphism differentially regulates 5‐HTT expression, with the long allele (L), the SIDS‐associated allele, being a more effective promoter than the short allele (S). We therefore investigated the 5‐HTT promoter polymorphism in a cohort of 87 SIDS cases (43 African American and 44 Caucasian) and gender/ethnicity‐matched controls. Significant positive associations were found between SIDS and the 5‐HTT genotype distribution (P = 0.022), specifically with the L/L genotype (P = 0.048), and between SIDS and the 5‐HTT L allele (P = 0.005). There was also a significant negative association between SIDS and the S/S genotype (P = 0.011). The comparisons were repeated in the African American and Caucasian subgroups. The data patterns were consistent in the subgroups, i.e., the L/L genotype and L allele were increased in the cases, but not all subgroup comparisons were statistically significant. These results indicate a relationship between SIDS and the L allele of the 5‐HTT gene in African Americans and Caucasians, and if confirmed, will provide an important tool for identifying at‐risk individuals and estimating the risk of recurrence.

Autonomic Nervous System Dysregulation: Breathing and Heart Rate Perturbation During Wakefulness in Young Girls with Rett Syndrome

This study characterizes cardiorespiratory dysregulation in young girls with MECP2 mutation–confirmed Rett syndrome (RS). Respiratory inductance plethysmography of chest/abdomen and ECG was obtained during daytime wakefulness in 47 girls with MECP2 mutation–confirmed RS and 47 age-, gender-, and ethnicity-matched controls (ages 2–7 y). An in-home breath-to-breath and beat-to-beat characterization was conducted and revealed that breathing was more irregular, with an increased breathing frequency, mean airflow, and heart rate in RS versus controls. There was a decreased correlation between normal breathing and heart rate variability, and an exaggerated increase in heart rate response to breathholds in RS versus controls. We conclude that girls with RS have cardiorespiratory dysregulation during breathholds as well as during “normal” breaths and during breaths before and subsequent to breathholds. This dysregulation may offer insight into the mechanisms that render girls with RS more vulnerable to sudden death.

Association of the serotonin transporter gene with sudden infant death syndrome: A haplotype analysis

Serotonergic receptor binding in the arcuate nucleus, n. raphé obscurus, and other medullary regions is decreased in sudden infant death syndrome (SIDS) cases. Further, an insertion/deletion polymorphism in the promoter region of the serotonin transporter protein (5‐HTT) gene has recently been associated with risk of SIDS. This polymorphism differentially regulates 5‐HTT expression, with the long allele (L), the SIDS‐associated allele, being a more effective promoter than the short allele (S). To further elucidate the role of the 5‐HTT gene in SIDS, we investigated the 5‐HTT intron 2 polymorphism, which also differentially regulates 5‐HTT expression with the 12 repeat allele being the more effective promoter. In a cohort of 90 SIDS cases (44 African‐American and 46 Caucasian) and gender/ethnicity‐matched controls, significant positive associations were found between SIDS and the intron 2 genotype distribution (P‐value = 0.041) among African‐American SIDS vs. African‐American controls, specifically with the 12/12 genotype (P‐value = 0.03), and with the 12 repeat allele (P‐value=0.018). The frequency of the 12/12 genotype and 12‐repeat allele was significantly different (P < 0.001) between the African‐American and Caucasian SIDS cases. Furthermore, the promoter and intron 2 loci were in significant linkage disequilibrium, and the L‐12 haplotype was significantly associated with SIDS in the African‐American (P = 0.002) but not Caucasian (P = 0.117) subgroups. These results indicate a relationship between SIDS and the 12‐repeat allele of the intron 2 variable number tandem repeat of the 5‐HTT gene in African‐Americans, and a significant role of the haplotype containing the 12‐repeat allele and the promoter L‐allele in defining SIDS risk in African‐Americans. These data, if confirmed in larger studies, may begin to explain the differences in SIDS incidence by ethnicity, suggest a role for levels of 5‐HTT expression in generation of SIDS susceptibility, and provide an important tool for identifying at‐risk individuals and estimating the risk of recurrence.

Sudden infant death syndrome: Case-control frequency differences at genes pertinent to early autonomic nervous system embryologic development

We have previously identified polymorphisms in the serotonin transporter gene promoter region and in intron 2 that were more common among sudden infant death syndrome (SIDS) cases compared with control subjects. To elucidate further the genetic profile that might increase an infants vulnerability to SIDS, we focused on the recognized relationship between autonomic nervous system (ANS) dysregulation and SIDS. We therefore studied genes pertinent to early embryologic development of the ANS, including MASH1, BMP2, PHOX2a, PHOX2b, RET, ECE1, EDN1, TLX3, and EN1 in 92 probands with SIDS and 92 gender- and ethnicity-matched control subjects. Eleven protein-changing rare mutations were identified in 14 of 92 SIDS cases among the PHOX2a, RET, ECE1, TLX3, and EN1 genes. Only 1 of these mutations (TLX3) was identified in 2 of 92 control subjects. Black infants accounted for 10 of these mutations in SIDS cases and 2 control subjects. Four protein-changing common polymorphisms were identified in BMP2, RET, ECE1, and EDN1, but the allele frequency did not differ between SIDS cases and control subjects. However, among SIDS cases, the allele frequency for the BMP2 common polymorphism demonstrated ethnic differences; among control subjects, the allele frequency for the BMP2 and the ECE1 common polymorphisms also demonstrated ethnic differences. These data represent further refinement of the genetic profile that might place an infant at risk for SIDS.

Diaphragm Pacing with a Quadripolar Phrenic Nerve Electrode: An International Study

We sought to determine the international experience with the quadripolar diaphragm pacer system and to test two hypotheses: the incidence of pacer complications would be (1) increased among pediatric as compared to adult patients; and (2) highest among active pediatric patients with idiopathic congenital central hypoventilation syndrome (CCHS). Data were collected via a questionnaire coupled with the Atrotech Registry data for a total of 64 patients (35 children and 29 adults) from 14 countries. Thoracic implantation of electrodes and bilateral pacer use each occurred in 94% of all subjects. Tetraplegic (vs pediatric CCHS) patients were more typically paced 24 hours/day (P = 0.001). Pacing duration averaged 2.0 ± 1.0 years among children and 2.2 ± 1.1 years among adults. Infections occurred among 2.9% of surgical procedures, all in pediatric CCHS patients (vs pediatric tetraplegic patients, P = 0.01). The incidence of mechanical trauma was 3.8%, without significant differences among patient groups. The incidence of presumed electrode and receiver failure were 3.1% and 5.9%, respectively, with internal component failure greater among pediatric CCHS than pediatric tetraplegic patients (P < 0.01). Intermittent or absent function of 0–4 electrode combinations occurred among 19% of all patients, with increased frequency among pediatric CCHS than pediatric tetraplegic patients (P < 0.03). Complication‐ free successful pacing occurred in 60% of pediatric and 52% of adult patients. In all, 94 % of the pediatric and 86% of the adult patients paced successfully after the necessary intervention. Although pacer complications were not increased among pediatric as compared to adult patients, the incidence of complications was highest among the active pediatric patients with CCHS. Longitudinal study of these patients will provide invaluable information for modification and improvement of the quadripolar system.

Effect of auditory, tactile, visual, and vestibular intervention on length of stay, alertness, and feeding progression in preterm infants

This study determined whether an auditory, tactile, visual, and vestibular intervention (ATVV) reduced the length of hospitalization of 37 preterm infants by increasing the proportion of alert behavioral states, thereby improving their feeding progression. Participants comprised 12 infants born between 23 and 26 weeks’gestation with normal head ultrasounds and 25 CNS‐injured infants born between 23 and 31 weeks’gestation. Infants were randomly assigned to the control group (11 males, five females) or study group (seven males, 14 females) at 32 weeks’postconceptional age. ATVV intervention was administered to the study group for 15 minutes, twice daily, 5 days per week, from 33 weeks of age until discharge. The study group demonstrated increased alertness during the first 5 minutes of intervention, which was significantly correlated to length of stay (p<0.05). The proportion of nippled (teat) intake increased significantly faster for the study group (p=0.0001). Infants in the study group were discharged at a mean of 36.54 weeks, 1.6 weeks earlier than control infants (p<0.05). ATVV intervention facilitated increased alertness, faster transition to complete nipple feeding, and decreased length of hospitalization.

Diaphragm pacing in infants and children

Since 1976 we have implanted bilateral diaphragm pacers in 34 infants and children: 26 with central hypoventilation syndrome (CHS), three with myelomeningocele, and five with quadriplegia. Compared to adults, several modifications have been necessary to achieve effective ventilation in infants and younger children. In all instances, a tracheostomy has been necessary due to impaired neuromuscular control of upper airway patency during pacing. Bilateral pacing has been necessary to achieve adequate ventilation; in the CHS children with normal awake ventilation, bilateral pacing during sleep eliminates the need for positive pressure ventilation. For the remaining children, adequate awake ventilation is achieved with bilateral pacing, thus permitting substantially greater mobility and limiting use of the ventilator to sleep time. Our longest survivor has now been paced for 10.7 years, and in no instance has phrenic nerve damage occurred secondary to electrical stimulation. Our current pacing regime is characterized by moderate respiratory rates (21 breaths/min), long interpulse intervals (95 ms), and short inspiratory times (0.6 sec), resulting in 50%-75% fewer stimuli/min compared to our previous regime. For all infants and children requiring 24-hour ventilatory assistance, our recent successes in maintaining ventilation using significantly fewer stimuli suggest that long-term continuous pacing is a realistic future goal.

Cardiac rhythm disturbances among children with idiopathic congenital central hypoventilation syndrome

The objective of this study was to determine whether subjects with congenital central hypoventilation syndrome (CCHS) had an increased frequency of cardiac arrhythmias and decreased heart rate variability when compared to subjects without a known deficit in control of breathing, and that these abnormalities would be exaggerated by anesthesia. Continuous ambulatory Holter recordings were obtained in patients with CCHS and compared to two otherwise healthy control groups without a deficit in control of breathing: one with an intact airway (n = 11) and a second group with a tracheostomy (n = 6). Holter recordings were obtained before, during (under general anesthesia), and after bronchoscopy. Fourteen children with CCHS (age: 9.3 ± 4.4 years mean ± S.D.) were studied, and 7 underwent bronchoscopy. Seventeen control children were studied (age 6.6 ± 3.6 years): 11 without a tracheostomy, and 6 with a tracheostomy who also underwent bronchoscopy.