Nancy L. Glass

Spectrum of CHD7 Mutations in 110 Individuals with CHARGE Syndrome and Genotype-Phenotype Correlation

CHARGE syndrome is a well-established multiple-malformation syndrome with distinctive consensus diagnostic criteria. Characteristic associated anomalies include ocular coloboma, choanal atresia, cranial nerve defects, distinctive external and inner ear abnormalities, hearing loss, cardiovascular malformations, urogenital anomalies, and growth retardation. Recently, mutations of the chromodomain helicase DNA-binding protein gene CHD7 were reported to be a major cause of CHARGE syndrome. We sequenced the CHD7 gene in 110 individuals who had received the clinical diagnosis of CHARGE syndrome, and we detected mutations in 64 (58%). Mutations were distributed throughout the coding exons and conserved splice sites of CHD7. Of the 64 mutations, 47 (73%) predicted premature truncation of the protein. These included nonsense and frameshift mutations, which most likely lead to haploinsufficiency. Phenotypically, the mutation-positive group was more likely to exhibit cardiovascular malformations (54 of 59 in the mutation-positive group vs. 30 of 42 in the mutation-negative group; P=.014), coloboma of the eye (55 of 62 in the mutation-positive group vs. 30 of 43 in the mutation-negative group; P=.022), and facial asymmetry, often caused by seventh cranial nerve abnormalities (36 of 56 in the mutation-positive group vs. 13 of 39 in the mutation-negative group; P=.004). Mouse embryo whole-mount and section in situ hybridization showed the expression of Chd7 in the outflow tract of the heart, optic vesicle, facio-acoustic preganglion complex, brain, olfactory pit, and mandibular component of the first branchial arch. Microarray gene-expression analysis showed a signature pattern of gene-expression differences that distinguished the individuals with CHARGE syndrome with CHD7 mutation from the controls. We conclude that cardiovascular malformations, coloboma, and facial asymmetry are common findings in CHARGE syndrome caused by CHD7 mutation.

Toward a genetic etiology of CHARGE syndrome: I. A systematic scan for submicroscopic deletions

CHARGE syndrome is a distinctive subgroup within the more heterogeneous group of patients with CHARGE association. While significant progress has been made in the clinical delineation of this syndrome, the molecular basis of the disorder remains unknown. Based on the complex phenotype, some overlap with DiGeorge/velocardiofacial syndrome (DGS/VCFS), and its estimated population incidence, we hypothesized that CHARGE syndrome could be caused by an unidentified genomic microdeletion. In order to address this hypothesis, we carried out a genome‐wide screen for loss of expected heterozygosity using 811 microsatellite markers in ten CHARGE syndrome subjects and their unaffected parents. Eight markers gave results suggestive of failure to inherit one parental allele. These loci were tested with fluorescence in situ hybridization (FISH), but none showed evidence of deletion. This screen sets upper limits on the length of a CHARGE‐related microdeletion, should that be the genetic mechanism underlying the phenotype.

Congenital sucrase-isomaltase deficiency presenting with failure to thrive, hypercalcemia, and nephrocalcinosis

BackgroundDisaccharide Intolerance Type I (Mendelian Interance in Man database: *222900) is a rare inborn error of metabolism resulting from mutation in sucrase-isomaltase (Enzyme Catalyzed 3.2.1.48). Usually, infants with SI deficiency come to attention because of chronic diarrhea and nutritional evidence of malabsorption.Case PresentationWe describe an atypical presentation of this disorder in a 10-month-old infant. In addition to chronic diarrhea, the child displayed severe and chronic hypercalcemia, the evaluation of which was negative. An apparently coincidental right orbital hemangioma was detected. Following identification of the SI deficiency, an appropriately sucrose-restricted, but normal calcium diet regimen was instituted which led to cessation of diarrhea, substantial weight gain, and resolution of hypercalcemia.ConclusionsThis case illustrates that, similar to congenital lactase deficiency (Mendelian Interance in Man database: *223000, Alactasia, Hereditary Disaccharide Intolerance Type II), hypercalcemia may complicate neonatal Sucrase-Isomaltase deficiency. Hypercalcemia in the presence of chronic diarrhea should suggest disaccharide intolerance in young infants.

Successful implementation of a protocol for photorefractive keratectomy in children requiring anesthesia

Purpose: To describe a protocol for treating children with photorefractive keratectomy (PRK) under general anesthesia and to review intraoperative and postoperative complications. Setting: Institutional academic practice. Methods: Nine patients between 3 years and 9 years of age were treated with PRK under general anesthesia for anisometropia with unilateral high myopia or high hyperopia and amblyopia of the affected eye. Induction of anesthesia and the surgical procedure were carried out in separate rooms. The laser beam was centered on the entrance pupil, and eye position was monitored throughout the procedure. Specific precautions were taken before and during the procedure to prevent unwanted effects of inhalational anesthetic agents on laser performance. Results: All children did well, with no anesthesia‐related or treatment‐related complications. Conclusions: Our protocol for PRK under general anesthesia was effective and efficient in children who were unable to cooperate for the procedure using local anesthesia. It can be adapted for laser in situ keratomileusis and other refractive surgical procedures in children and uncooperative adults.

Technical communication: new teaching model for practicing ultrasound-guided regional anesthesia techniques: no perishable food products!

BACKGROUND:There is a pronounced learning curve for the technique of ultrasound-guided regional anesthesia. Practicing with a simulator model has been shown to speed the acquisition of these skills for various ultrasound-guided procedures. However, commercial models for ultrasound-guided regional anesthesia may be too costly or not readily available. Models using turkey breasts or tofu blocks have the disadvantage of containing perishable food products that can be a source for infection. We describe an alternative inexpensive model that is made from nonperishable components readily available in the operating room. METHODS:The materials required include 1 clean used 500-mL bag of IV fluids, a bottle of Premisorb® (TYCO Healthcare Group, Mansfield, MA), and a piece of foam material approximately 0.3 cm in diameter and 5 cm in length trimmed from operating room foam pads. After filling the IV bag with tap water and inserting the foam into the IV bag from the outlet port of the IV bag, one-third of a bottle of Premisorb (approximately 15 g) is poured into the IV bag. The outlet port of the bag is then sealed by taping the rubber stopper that originally came with the bag. RESULTS:Premisorb, a solidifying agent frequently used to absorb irrigating fluids or blood in operating room suction canisters, produces a gel-like material in the IV bag. The foam inserted into the bag creates a relatively hyperechoic target. This gel-like substance in the bag will seal the holes created after multiple practice needle insertions, resulting in minimal leakage. The semitransparent nature of the gel allows the trainee to visualize the target directly and on the ultrasound screen. CONCLUSION:The model we describe is inexpensive and easy to make from materials readily available in the operating room with the advantages of being nonperishable, easy to carry, and reusable.

Comparison of the development of performance skills in ultrasound-guided regional anesthesia simulations with different phantom models.

Background Ultrasound-guided regional anesthesia (UGRA) skills are traditionally obtained by supervised performance on patients, but practice on phantom models improves success. Currently available models are expensive or use perishable products, for example, olive-in-chicken breasts (OCB). We constructed 2 inexpensive phantom (transparent and opaque) models with readily available nonperishable products and compared the process of learning UGRA skills by novice practitioners on these models with the OCB model. Methods Three experts first established criteria for a satisfactory completion of the simulated UGRA task in the 3 models. Thirty-six novice trainees (<20 previous UGRA experience) were randomly assigned to perform a UGRA task on 1 of 3 models—the transparent, opaque, and OCB models, where the hyperechoic target was identified, a needle was advanced to it under ultrasound guidance, fluid was injected, and images were saved. We recorded the errors during task completion, number of attempts and needle passes, and the time for target identification and needle placement until the predetermined benchmark of 3 consecutive successful UGRA simulations was accomplished. Results The number of errors, needle passes, and time for task completion per attempt progressively decreased in all 3 groups. However, failure to identify the target and to visualize the needle on the ultrasound image occurred more frequently with the OCB model. The time to complete simulator training was shortest with the transparent model, owing to shorter target identification times. However, trainees were less likely to agree strongly that this model was realistic for teaching UGRA skills. Conclusions Training on inexpensive synthetic simulation models with no perishable products permits learning of UGRA skills by novices. The OCB model has disadvantages of containing potentially infective material, requires refrigeration, cannot be used after multiple needle punctures, and is associated with more failures during simulated UGRA. Direct visualization of the target in the transparent model allows the trainee to focus on needle insertion skills, but the opaque model may be more realistic for learning target identification skills required when UGRA is performed on real patients in the operating room.

SNP genotyping to screen for a common deletion in CHARGE Syndrome

BackgroundCHARGE syndrome is a complex of birth defects including coloboma, choanal atresia, ear malformations and deafness, cardiac defects, and growth delay. We have previously hypothesized that CHARGE syndrome could be caused by unidentified genomic microdeletion, but no such deletion was detected using short tandem repeat (STR) markers spaced an average of 5 cM apart. Recently, microdeletion at 8q12 locus was reported in two patients with CHARGE, although point mutation in CHD7 on chromosome 8 was the underlying etiology in most of the affected patients.MethodsWe have extended our previous study by employing a much higher density of SNP markers (3258) with an average spacing of approximately 800 kb. These SNP markers are diallelic and, therefore, have much different properties for detection of deletions than STRs.ResultsA global error rate estimate was produced based on Mendelian inconsistency. One marker, rs431722 exceeded the expected frequency of inconsistencies, but no deletion could be demonstrated after retesting the 4 inconsistent pedigrees with local flanking markers or by FISH with the corresponding BAC clone. Expected deletion detection (EDD) was used to assess the coverage of specific intervals over the genome by deriving the probability of detecting a common loss of heterozygosity event over each genomic interval. This analysis estimated the fraction of unobserved deletions, taking into account the allele frequencies at the SNPs, the known marker spacing and sample size.ConclusionsThe results of our genotyping indicate that more than 35% of the genome is included in regions with very low probability of a deletion of at least 2 Mb.