Malformations of the heart, kidney, palate, and skeleton in alpha-MHC-Hoxb-7 transgenic mice.

To begin to define the genetic network involved in cardiogenesis, we generated mice bearing the alpha-myosin heavy chain (MHC)-Hoxb-7 transgene. We hypothesized that using the cardiac-specific alpha-MHC promoter, we can direct ectopic expression of Hoxb-7 in the heart and perturb its normal development. Both whole mount in situ hybridization and northern analyses showed that this alpha-MHC promoter resulted in transgene expression in the developing heart. Severe ventricular septal defects (VSD) were found in several mutant mice. Interestingly, transgenic mice were observed to have other malformations as well, including cleft palate, renal anomalies, and skeletal abnormalities in the craniocervical and costosternal regions. The kidney defect consisted of double ureter and pelvis. In summary, we have shown that a dominant gain-of-function mutation of Hoxb-7 using the murine alpha-MHC promoter results in perturbation of the genetic circuitry underlying multiple developmental processes, including cardiogenesis. Misexpression of Hoxb-7 during heart development may be involved in the pathogenesis of VSD.

Homeobox genes and heart development.

Genes encoding transcription factors are fundamental to developmental processes because their DNA-binding proteins can control expression of a multitude of genes. Therefore, activation of a single transcription factor gene can act as a genetic switch that controls the developmental destinies of groups of cells. Results of expression studies and mutational analyses in mice and Drosophila suggest an essential role in heart development for the homeobox class of transcription factor genes. Understanding the genetic circuitry of cardiogenesis will facilitate the identification of individuals with inherited cardiac diseases, and the possible prevention of these diseases with drug therapy or lifestyle modification. It may also allow for genetic counseling of affected individuals and family members and enable gene therapy aimed at correcting these defects.

Bone mineral content in infants and children with chronic cholestatic liver disease.

OBJECTIVE: To assess bone mineral content (BMC) status and serum vitamin D metabolite levels of infants and children with chronic cholestatic liver disease. To determine if severity of bone disease in these patients correlates with serum vitamin D metabolite levels. METHODOLOGY: We measured radial BMC with the use of a single-beam photon absorptiometer and serum vitamin D metabolite levels in 56 patients with chronic cholestasis seen at our institution from 1985 through 1991. Patients were divided into two groups according to age. RESULTS: In group 1 (n = 37; age 2 to 22 months), decreased levels of BMC were seen as early as the first few months of life, with sharp decline observed with increasing age (approaching 3 to 5 standard deviations [SD] below the mean, P < .0003). Older patients (group 2, n = 19; age 2 to 20 years) had BMC values which clustered between 2 and 4 SD below the mean throughout the age range. Although a downward trend also was noted with increasing age, this was not statistically significant. Despite correction for weight-age or height-age, BMC was decreased in most of these patients. No correlation between severity of osteopenia and serum levels of 25(OH)-vitamin D and 1,25(OH)2-vitamin D was observed in either infants or older children. CONCLUSIONS: Decreased bone mineralization, as a complication of chronic cholestatic conditions, is a disease process that begins early in infancy, rapidly worsens with increasing age and hepatic dysfunction, and remains relatively stable in children with more stable liver disease.

The pharmacokinetics of oral ranitidine in children and adolescents with cystic fibrosis.

The pharmacokinetics of oral ranitidine were studied in 9 patients (ages 9.9 to 19.6 years) with cystic fibrosis (CF). Patients were evaluated at steady-state conditions, and the mean maximum serum concentration (Cmax) was 845.7 +/- 448.1 ng/mL. To adjust for the variable drug dosing used among study patients, both Cmax and area under the concentration curve (AUC) were standardized to dose (CmaxST and AUCST, respectively) and were 217.9 +/- 87.9 ng/mL and 1038.0 +/- 242.2 ng/mL.h. The elimination half-life (t1/2) was 2.7 +/- 1.4 hours, and the apparent steady-state volume of distribution (Vdss) was 4.6 +/- 1.7 L/kg. The plasma clearance was 1.022 +/- 0.290 L/kg/h. The Vdss in this study was greater than that previously reported in children with peptic ulcer disease. Statistically significant relationships between pharmacokinetic parameters and measures of disease severity were not observed in the study population. The pharmacokinetics of ranitidine in children and adolescents with CF may differ from those in children and adolescents without CF.

Fat-soluble vitamin deficiency in infants and children.

Fat-soluble vitamin deficiency is known to result in various complications that may be prevented if the problem is recognized and managed appropriately. In infants and children with chronic cholestasis, replacement therapy of the fat-soluble vitamins, vitamins A, D, E, and K, may prove extremely difficult because low concentrations of intraluminal bile acids lead to malabsorption of these compounds and other fat-soluble substances. Recent progress in the use of a water-soluble form of vitamin E, d-alpha-tocopheryl polyethylene glycol-1000 succinate, has enabled correction of vitamin E-deficiency states in these patients. It has also allowed for the admixture and coadministration of other fat-soluble vitamins and compounds in d-alpha-tocopheryl polyethylene glycol-1000 succinate to enhance their absorption. For managing vitamin K deficiency, similar success has been achieved using a vitamin K compound solubilized in glycocholate and lecithin. Vitamin A deficiency has been implicated in the higher incidence of childhood mortality and morbidity in Third World countries. Increased risk of childhood cancer has recently been associated with intramuscular injection of vitamin K to newborns. Finally, it is worth noting that among the pediatric population, exclusively breastfed infants, in general, are at risk for hypovitaminosis D, and at even greater risk in the absence of adequate exposure to sunlight or when the maternal diet is not sufficient to provide for vitamin D requirements.

d-Alpha-tocopheryl polyethylene glycol-1000 succinate enhances the absorption of vitamin D in chronic cholestatic liver disease of infancy and childhood.

Rickets and osteopenia, common problems in chronic childhood cholestasis, have been attributed to vitamin D malabsorption leading to reduced serum levels of 25(OH)-vitamin D. d-alpha-Tocopheryl polyethylene glycol-1000 succinate (TPGS), a water-soluble form of vitamin E, forms micelles at low concentration. We evaluated the potential role of TPGS in enhancing vitamin D absorption in eight children (aged 5 mo to 19 y) with severe chronic cholestasis (three extrahepatic biliary atresia, three nonsyndromic intrahepatic cholestasis, and two Alagille syndrome). To evaluate vitamin D absorption, the subjects received vitamin D3 1000 IU/kg (maximum dose of 50,000 IU); they then received the same dose of vitamin D3 mixed with TPGS (25 IU/kg). Serial serum vitamin D3 levels and areas under the curve were measured. All patients had enhanced absorption of vitamin D when it was administered in a mixture with TPGS. Mean area under the curve for serum vitamin D3 was 403.0 +/- 83.1 nmol x h/L (155.6 +/- 32.1 ng x h/mL), with a mean rise above baseline of 13.5 +/- 1.8 nmol/L (5.2 +/- 0.7 ng/mL) with vitamin D/TPGS compared with no rise when vitamin D was given alone (both p less than 0.001). Seven patients have been followed for at least 3 mo while receiving the vitamin D/TPGS combination. Those with initially low serum 25(OH)-vitamin D levels (less than 37.5 nmol/L or 15 ng/mL) had normalization (range 37.5-146 nmol/L) within 1 mo, whereas those with initially normal levels remained normal. While the patients were receiving vitamin D/TPGS, serum vitamin E to total lipid ratio either normalized or remained normal.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of orthotopic liver transplantation on bone mineral content and serum vitamin D metabolites in infants and children with chronic cholestasis.

Almost all infants and children with chronic cholestasis have osteopenia. We evaluated the effect of orthotopic liver transplantation on bone mineral content and serum 25(OH)-vitamin D-[25(OH)D]-of nine infants and children (five girls; age, 6 to 21 mo at the time of orthotopic liver transplantation) with end-stage liver disease resulting from chronic cholestasis. We hypothesized that after orthotopic liver transplantation, decreased bone mineral content will recover and the serum 25(OH)D level will either normalize or remain normal in those who were previously vitamin D deficient or sufficient, respectively. All had subnormal bone mineral content before transplant. On long-term follow-up (> 4 mo) of seven patients, bone mineral content normalized in all between 6.5 and 19 mo after transplant, with a mean of 11.2 +/- 4.5 mo. In six patients with normal serum 25(OH)D levels before orthotopic liver transplantation, the serum 25(OH)D levels had declined markedly 1 to 2 mo after transplant, followed by return to normal by 3 to 6 mo. Low serum 25(OH)D levels (< 15 ng/ml) in three patients before orthotopic liver transplantation normalized after transplant. Although there was no correlation between bone mineral content and serum 25(OH)D level before transplant, sustained normal serum 25(OH)D and 1,25(OH)2D levels preceded or accompanied normalization of bone mineral content in the seven patients available for long-term follow-up. We conclude that (a) in infants and children younger than 2 yr with chronic cholestasis, bone mineral content normalizes approximately 11 mo after orthotopic liver transplantation. This normalization is preceded by a sustained period of normal serum 25(OH)D levels.(ABSTRACT TRUNCATED AT 250 WORDS)

Genomic organization and chromosome localization of the murine homeobox gene Pmx.

Homeobox genes are expressed in very specific temporal and spatial patterns and function as transcriptional regulators of developmental processes. The murine homeobox gene, Pmx (paired mesoderm homeobox), is expressed in a mesodermally restricted pattern in embryos and most abundantly in cardiac, skeletal, and smooth muscle tissues in adults. Previously, this murine gene was named K-2 and mHox, while the human homolog was named Phox1. In this report, the localization of Pmx has been determined by interspecific backcross analysis. The Pmx gene is located on Chromosome 1, approximately 3.3 cM distal to the Gsh-4 homeobox locus. The sequence of the Pmx transcript has been extended toward the 5' end and corresponds in size to one of the transcripts previously detected by Northern blot analysis. Sequence analysis indicates that Pmx is the first characterized mammalian gene to encode a paired type homeodomain, but not a paired domain. The Pmx gene includes at least five exons spanning a minimum of 60 kb of genomic DNA, making this the largest known murine homeobox gene.

The role of magnesium in the pathogenesis of bone disease in childhood cholestatic liver disease: a preliminary report.

BACKGROUND: Magnesium deficiency may contribute to the metabolic bone disease that complicates chronic cholestatic liver disease. We hypothesized that magnesium deficiency alters vitamin D metabolism by decreasing parathyroid hormone (PTH) response, resulting in decreased serum osteocalcin and decreased bone accretion. METHODS: Nine subjects, age 3-22 years, with cholestatic liver disease were evaluated with the magnesium retention test. The response of PTH, 1,25(OH)2 vitamin D, and osteocalcin to provocative stimuli and dual x-ray absorptiometry measurement of bone mineral density (BMD) of the lumbar spine were assessed. Thereafter, subjects were treated with oral magnesium supplements. RESULTS: All nine subjects were magnesium depleted. Repletion with magnesium was successful in seven subjects, and required 4 to 31 (median 14) months with doses of 6 to 34 (median 11) mg/kg/day. Baseline serum PTH was significantly reduced in the cholestatic subjects compared to 15 age-matched controls. Comparison of baseline to repleted provocative testing was performed in six Mg-repleted subjects. Osteocalcin response increased significantly (p = 0.048) with repletion, while PTH response increased (p = 0.061). Lumbar spine BMD increased modestly with repletion (p = 0.093). CONCLUSIONS: This preliminary report suggests that magnesium depletion is extremely common in children with chronic cholestasis. We speculate that magnesium supplementation may be warranted to forestall the progression of metabolic bone disease in chronic cholestasis.

Paired-related homeobox genes cooperate in handplate and hindlimb zeugopod morphogenesis.

The closely related homeobox genes prx-1 and prx-2 are expressed in lateral plate and limb bud mesoderm, but targeted inactivation of these genes failed to demonstrate a limb phenotype. Here we report that mice carrying compound mutations in prx-1 and prx-2 have severe limb deformities. In the forelimb autopod, pre- and postaxial polydactyly were found most commonly, but also syndactyly, oligodactyly, and abnormal digit placement affecting posterior elements were observed. In the hindlimb, preaxial polydactyly with variable expressivity was seen in all cases. Extreme distal digit duplications were seen in both the fore- and hindlimbs. prx-1; prx-2 double-mutant mice also displayed extreme shortening and impaired ossification of the hindlimb zeugopods. Integrity of the forelimb apical ectodermal ridge was abnormal as determined by expression of FGF8 and BMP4. Expression of msx-1 and msx-2, markers for BMP signaling pathways, was absent in regions of the posterior handplates, while expression of Shh and patched was unaffected. The mutant phenotypes were dosage dependent, since prx-1 -/-; prx-2 +/- mice also displayed severe limb abnormalities. These data suggest that prx-1 and prx-2 cooperatively regulate handplate and hindlimb zeugopod morphogenesis through BMP-mediated signaling pathways.