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American Journal of Medical Genetics 85:476–478 (1999)
Prenatal Evaluation of a De Novo X;9 Translocation
Baruch Feldman,1 Ralph L. Kramer,1 Salah A.D. Ebrahim,2 Dayna J. Wolff,3 and Mark I. Evans1,2,4*
1
Division of Reproductive Genetics, Department of Obstetrics and Gynecology, Wayne State University,
Detroit, Michigan
2
Division of Reproductive Genetics, Department of Pathology, Wayne State University, Detroit, Michigan
3
Center for Human Genetics, Case Western Reserve University, Cleveland, Ohio
4
Division of Reproductive Genetics, Department of Molecular Medicine and Genetics, Wayne State University,
Detroit, Michigan
A case of X-autosome translocation was diagnosed prenatally [46,X,t(X;9)(p21.3∼
22.1;q22]. We describe the use of fluorescence in situ hybridization (FISH) to estimate the integrity of the Duchenne muscular dystrophy (DMD) gene. X-inactivation
studies were used as well to assess the probability of phenotypic abnormalities associated with functional partial disomy X and
monosomy 9. Am. J. Med. Genet. 85:476–478,
1999. © 1999 Wiley-Liss, Inc.
KEY WORDS: X;9 translocation; prenatal
diagnosis; X inactivation;
FISH
INTRODUCTION
X-autosome translocations have generally been
shown to be of parental origin or arise de novo. The
phenotypic consequences of the translocation will depend on the break point on the X chromosome and the
autosome as well as the pattern of X inactivation. Most
of these patients are either phenotypically normal or
had gonadal dysgenesis, whereas approximately 9%
had multiple anomalies and/or mental retardation
[Schmidt and Du Sart, 1992]. When the break point
occurs within the proper gene, a “classic” X-linked disorder can be phenotypically expressed in a female [Zatz
et al., 1981]. We describe a case of fetal X-autosome
translocation in which molecular studies were used in
order to estimate the X-inactivation pattern and the
integrity of the Duchenne muscular dystrophy (DMD)
gene.
CASE REPORT
A 38-year-old patient was seen at 17 weeks of gestation for both elevated maternal serum alpha-
*Correspondence to: Mark I. Evans, M.D., Division of Reproductive Genetics, Department of Obstetrics and Gynecology, Hutzel Hospital/Wayne State University, 4707 St. Antoine Boulevard, Detroit, MI 48201. E-mail: [email protected]
Received 17 November 1998; Accepted 23 March 1999
© 1999 Wiley-Liss, Inc.
fetoprotein (MSAFP) level and advanced maternal age.
The patient’s medical, obstetrical, and family history
was unremarkable. Ultrasound examination demonstrated no anomalies other than a 3-mm choroid plexus
cyst.
Cytogenetic analysis of cultured amniocytes revealed
the fetal karyotype to be 46,X,t(X;9)(p21.3∼22.1;q22).
The translocation arose de novo because both parents
were tested and shown to be karyotypically normal.
There was concern, however, that the break point on
the X chromosome could involve the DMD gene at
Xp21.3.
In order to obtain a more precise estimate of fetal
risk for DMD, assessment of the specific break point
was determined with a specific fluorescence in situ hybridization (FISH) probe (Quint-Essential X-Specific
DNA Probe, Oncor, Gaithersburg, MD). This probe hybridizes to the Xp21.2-p21.3 region and specifically
identifies sequences containing the DMD locus. FISH
analysis showed that the Xp21.2-p21.3 locus was present on the normal and the abnormal X-chromosome. No
other chromosomes demonstrated a hybridization signal (Fig. 1). This observation suggests that the DMD
region was not disrupted, translocated, or deleted by
the rearrangement and significantly reduced the likelihood of the fetus affected with DMD. The patient was
counseled that the results of the FISH studies are reassuring, however, normal gene function cannot be determined by this technique. Fetal muscle biopsy for
dystrophin studies was offered for confirmation of normal gene function, but the couple declined any further
invasive procedures.
In order to assess the risk of functional partial disomy X and concomitant monosomy 9, an assay to
analyze methylation at the fragile X mental retardation gene (FMR1) was performed, as described by Carrel and Willard [1996]. DNA from the patient and appropriate controls were digested with the methylation-sensitive enzyme HpaII. This enzyme cleaves at
two restriction sites near the CGG repeat of the unmethylated FMR1 gene on the active X chromosome
but does not digest these sites on the inactive (methylated) X chromosome. Digested and undigested DNA
samples were amplified by polymerase chain reaction.
Samples were separated by gel electrophoresis, and X-
Prenatal Evaluation of X;9 Translocation
Fig. 1.
477
FISH of metaphase chromosomes probed for the DMD region, Xp21.2-p21.3. Positive hybridization is detected on both X chromosomes.
inactivation ratios were determined by visual comparison of the cut and uncut fragments obtained from the
radiograph. The results of the X-inactivation study
strongly suggest nonrandom inactivation of the normal
X chromosome (Fig. 2). This significantly reduced the
risk of functional partial disomy X and partial monosomy 9. Based on these results the couple decided to
continue the pregnancy.
Unfortunately, at 33 weeks of gestation, preterm
premature rupture of membranes occurred and the patient was hospitalized. At 34 weeks of gestation, chorioamnionitis was diagnosed, but prior to induction of
labor, intrauterine fetal death was confirmed. Labor
was induced and the patient delivered a stillborn female infant. There were no apparent fetal anomalies,
however, a true knot was noted in the umbilical cord.
Histological examination of the placenta confirmed the
diagnosis of chorioamnionitis but autopsy did not show
any other fetal anomalies. Normal dystrophin gene
function was demonstrated by positive immunofluorescence staining for dystrophin [Arahata et al, 1989] in
tissue obtained from postnatal fetal muscle biopsy.
DISCUSSION
Fig. 2. Results of FMR1 methylation analysis. For the fetus with the
X;9 translocation (lanes 3 and 4), only allele “2” was amplified following
digestion. Controls: normal female with random X inactivation (lanes 1
and 2); female with skewed X inactivation pattern (lanes 5 and 6); normal
male was included to control for complete digestion (lanes 7 and 8). u,
uncut; c, cut.
The phenotypic consequences of X-autosome translocation will depend on the breakpoint on the X chromosome and the autosome as well as the pattern of X
inactivation.
Duchenne muscular dystrophy (DMD) is an X-linked
muscle wasting disorder observed with a frequency of
approximately 1 in 3,500 newborn males. DMD has
also been described in females in which de novo Xautosome translocations have been shown to be the
cause [van Bakel et al., 1995]. Expression of the disease
in these females is the result of nonrandom inactivation of the normal X chromosome as well as a breakpoint inside or very close to the DMD gene on the translocated chromosome.
478
Feldman et al.
X inactivation is the process by which females
achieve dosage compensation by silencing one X chromosome. In chromosomally normal females the process
is random. However, most females with one abnormal
X chromosome demonstrate nonrandom inactivation.
Nonrandom inactivation of the normal X chromosome
will lead to phenotypically normal fetus. In 10–15% of
structurally balanced X-autosome heterozygotes, however, the mechanism fails and some cells with partial
disomy of the X chromosome and partial monosomy of
the autosomal translocated segment survive and cause
phenotypic abnormality [Schmidt and Du Sart, 1992;
Wolff et al., 1998].
Our observations on a case of X;9 translocation diagnosed prenatally illustrate the clinical dilemmas associated with X-autosome translocations. We believe that
the use of FISH and X-inactivation studies is an important adjunct to conventional cytogenetic techniques
in such cases. The use of molecular studies may determine the break points more precisely. It can also provide a more precise estimate of the phenotypic consequences of X-autosome translocation.
Unfortunately in the case presented, the fetus was
stillborn. This appeared to have resulted from causes
unrelated to the X-autosome translocation. The normal
dystrophin gene function observed postnatally con-
firmed that the DMD gene region was not disrupted as
suggested by the prenatal FISH studies. Although the
fetus appeared phenotypically normal, we obviously
cannot comment on postnatal development or ovarian
function.
REFERENCES
Arahata K, Hoffman EP, Kunkel LM, Ishiura S, Tsukahara T, Ishihara T,
Sunohara N, Nonaka I, Ozawa E, Sugita H. 1989. Dystrophin diagnosis: comparison of dystrophin abnormalities by immunofluorescence
and immunoblot analyses. Proc Natl Acad Sci USA 86:7154–7158.
Carrel L, Willard HF. 1996. An assay for X inactivation based on differential methylation at the fragile X locus, FMR1. Am J Med Genet 64:27–
30.
Schmidt M, Du Sart D. 1992. Functional disomies of the X chromosome
influence the cell selection and hence the X inactivation pattern in
females with balanced X-autosome translocations: a review of 122
cases. Am J Med Genet 42:161–169.
van Bakel I, Holt S, Craig I, Boyd Y. 1995. Sequence analysis of the breakpoint regions of an X;5 translocation in a female with Duchenne muscular dystrophy. Am J Hum Genet 57:329–336.
Wolff DJ, Schwartz S, Montgomery T, Zackowski JL. 1998. Random X
inactivation in a girl with a balanced t(X;9) and an abnormal phenotype. Am J Med Genet 77:401–404.
Zatz M, Vianna-Morgante AM, Campos P, Diament AJ. 1981. Translocation (X;6) in a female with Duchenne muscular dystrophy: implications
for the localization of the DMD locus. J Med Genet 18:442–447.
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