Abnormal Skin Fibroblast Cytogenetics in Four Dysmorphic Patients with Normal Lymphocyte Chromosomes
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AmJHum Genet 31:54-61, 1979 Abnormal Skin Fibroblast Cytogenetics in Four Dysmorphic Patients with Normal Lymphocyte Chromosomes ROBERTA A. PAGON,' JUDITH G. HALL,' SANDRA L. H. DAVENPORT,' JON AASE,' THOMAS H. NORWOOD,' AND HOLGER W. HOEHN' SUMMARY Four patients with features suggestive of chromosome disorders but with normal lymphocyte karyotypes were found to have chromosome aberrations in skin fibroblast karyotypes. Although mosaicism for chromosome abnor- malities in lymphocyte cultures is common, apparent restriction of mosai- cism to one tissue is unusual. We suggest that after examination of lymphocyte karyotypes, certain patients warrant cytogenetic evaluation of a second tissue, usually cultured skin fibroblasts. INTRODUCTION Many physicians dealing with genetic disorders have encountered patients whose clinical features strongly suggest a chromosome disorder, but who have apparently normal lymphocyte karyotypes. With chromosome banding, some of this discrepancy has been resolved through identification of previously unrecognizable chromosome changes. Where even careful banding of lymphocyte metaphases is negative, examina- tion of fibroblast cultures may be diagnostic. In a 12 month period, four of 12 patients with features suggestive of chromosome disorders (but with normal karyotypes in at least one lymphocyte culture) showed chromosome aberrations in cultured skin fibroblasts. Received August 15, 1978; revised September 18, 1978. This research was supported by National Institutes of Health grant GM 15253, Fellowship Training grant CA-90, and National Foundation-March of Dimes grant 6-44. ' Departments of Medicine, Pediatrics, and Pathology, University of Washington Medical School, Division of Medical Genetics and Children's Orthopedic Hospital and Medical Center, Seattle, Washington 98195. 2 Department of Pediatrics, University of New Mexico, Albuquerque, New Mexico. © 1979 by the American Society of Human Genetics. 0002-9297/79/3 lo1-0009$00.75 54 FIBROBLAST VERSUS LYMPHOCYTE CHROMOSOMES s5 CASE REPORTS Patient I The first case was referred at age 16, when the diagnosis of Down syndrome was first considered. He was born at term to a 30-year-old mother and a 32-year-old unrelated father. Birth weight was 3350 gm and length, 53 cm. The diagnosis of arthrogryposis multiplex congenita was made at birth, and surgical correction of dislocated hips and equinovarus deformities was done. Progressive scoliosis required bracing. Additional problems were hypospadias and herniation of an ectopic spleen into the scrotum. IQ was estimated to be 54. One brother and three sisters were normal. The mother had one first trimester spontaneous abortion. There was no family history of congenital malformation or mental retardation. Height was 153 cm (below the third percentile), weight was 76 kg (greater than the 97th percentile) and occipital frontal circumference (OFC) was 56 cm (50th percentile). The head was brachycephalic with a flat occiput and midface. There was bilateral ptosis and normal irides. In addition to generalized hirsutism, there was a low posterior hairline and slight synophrys. The palate was somewhat high-arched; the neck was webbed. Cardiac examination was normal. The chest was barrel shaped. There was no clinodactyly. Dermal ridge patterns showed nine ulnar loops and one whorl; palmar creases were normal. Repeated peripheral blood chromosome studies, including R-banding, were within normal limits (table 1). However, a skin biopsy at age 16 showed substantial mosaicism for a small accessory ring chromosome which could not be further identified (fig. 1). There was no increased chromosome "breakage" in these cultures. The euploid cells prevailed at later passages (table 1). Since the patient's phenotype suggested Down syndrome, although atypical, the accessory ring chromosome might represent chromosome 21 material, albeit without complete trisomy for band 21q22, which is considered specific for the Down phenotype [1]. No numerical or structural chromosome change was found in 25 lymphocyte metaphases examined in both parents. Patient 2 Our second case was the 2,890 gm, 51 cm product of an uncomplicated term pregnancy of a 30-year-old primagravida and 28-year-old unrelated father. When evaluated at 9 months for unusual facies, hypotonia, failure to thrive and developmental delay, she was felt to have clinical features typical of the 18q- syndrome [2]. At 5 years of age she was functioning at an 8-9 month performance level. Family history is negative for mental retardation and congenital malforma- tions. The patient's height, weight and OFC were below the third percentile. She had frontal bossing and narrow biparietal diameter with mid-face hypoplasia, flat nasal bridge and epicanthal folds. Esotropia and retinal coloboma of the left eye were not associated with nystagmus. The philtrum was short with a turned-down mouth and the palate was high-arched. The ears had prominent antihelices and antitragi. The labia minora were absent, and the clitoris was small. The fingers were tapered with clinodactyly of the second, fourth, and fifth digits bilaterally. Dermal ridges were hypoplastic with two whorls and eight ulnar loops. The patient's first cytogenetic evaluation on cultured lymphocytes at 9 months (table 1) revealed only normal metaphases; however, a repeat blood culture at age 5 showed minor mosaicism for 18q-. All of the metaphases from skin fibroblast cultures exhibited deletion of a portion of the long arm of chromosome 18 (fig. 1) for which R-banding suggested the following designation: 46,XX,del( 1 8)(pter-*q22:). Patient 3 Our third case was an 11 -year-old female referred for possible Turner syndrome. She was a 1,950 gm product of a 36 week gestation, born to a 37-year-old G8 P8 mother and a 48-year-old unrelated father. She was smaller and slower in development than her siblings; her IQ was 65. At age 91/2 years bilateral streak gonads, absence of the uterus and the left fallopian tube, 56 PAGON ET AL. C~~~ 't00 S~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~\0 3- x V0) X~~~~c* E X C-4X+ + 3 HX mN N ( c___s,,r,~. 3N XX X m E _NZ ._ 0 ) to-55 e to 0x--X E ~~~~~0 ~ ~&'.'~~~TB._ s , o00 FIBROBLAST VERSUS LYMPHOCYTE CHROMOSOMES 57 FIG. 1. -Partial karyotypes of R-banded preparations from fibroblast cultures. Shown are the G-group chromosomes from patients 1, 3, and 4, and the E-group from patient 2. malrotation of the intestine, and ectopic right kidney were incidental findings at laparotomy for appendicitis. She had mixed conductive and sensorineural hearing loss. Family history is negative for mental retardation and congenital anomalies. The patient's height was 116 cm and weight was 20 kg (both below the third percentile). The face was asymmetric with telecanthus, bilateral ptosis, slight synophrys, and downward slanting palpebral fissures. Preauricular pits were present bilaterally. The neck was short with slight webbing and a low posterior hairline. The labia majora were small and the clitoris slightly enlarged. There was clinodactyly of the right fifth finger and camptodactyly of the left fifth finger with partial syndactyly of the second, third, and fourth toes bilaterally. The nails were spoon shaped. Dermal ridge patterns showed eight low arches and two ulnar loops. Lymphocyte cultures at different ages (table 1) yielded only euploid and structurally normal metaphases, including 100 spreads evaluated from the second culture. Fibroblast karyotypes consistently revealed a supernumerary G-group-like chromosome with R-banding characteristics reminiscent of chromosome 22 (fig. 1 and table 1). A phenotypically similar patient with repeatedly normal blood cytogenetics and presumptive trisomy 22 mosaicism in skin fibroblast cultures has been previously described [3]. Patient 4 Our final case, who had typical Down syndrome features, but normal developmental milestones, was referred for evaluation of Down syndrome at 7 months. She was the 3,040 gm, 53 cm product of an uncomplicated term pregnancy of a 24-year-old mother. At 7 months, she was rolling over, sitting alone, and transferring objects from hand to hand. At 13 months, she had normal muscle tone, spoke several words, and was walking with support. Chromosome mosaicism was considered because of her normal psychomotor development. A 2-year-old female sib was normal; there was no family history of mental retardation or malformations. At 5 months, height was 64 cm (25th percentile), and weight, 6.2 kg (fifth percentile). She had the typical facies of Down syndrome. Brushfield spots were present, as well as wide space between the first and second toes, and clinodactyly of the fifth fingers. Muscle tone and developmental milestones were normal. Dermal ridge patterns showed eight ulnar loops and a fused proximal transverse crease. Analysis of 30 metaphases from lymphocyte cultures at 8 months revealed only euploid and 58 PAGON ET AL. structurally normal chromosomes (table 1); however, seven of 56 R-banded metaphases from skin fibroblast cultures included an additional chromosome 21 (fig. 1). METHODS PHA stimulated samples of peripheral blood were cultivated in Dulbecco-Vogt media and harvested in the usual manner. Skin fibroblast cultures, established from explants of small biopsies from the forearm, were maintained in serial passage [4]. Chromosome analyses of the fibroblasts were carried out on specimens prepared in situ on 1 x 3 in slides. R-banding was accomplished by the method of Sehested [5]. DISCUSSION Chromosome disorders are suspected in patients with features