High risk for unbalanced segregation of some reciprocal translocations: a large pedigree containing distal 4q trisomy from t(4;7)(q28;p22).

We report on a familial t(4;7)(q28;p22) with 2:2 adjacent-1 unbalanced segregation producing duplication of 4q28-->qter in multiple offspring. Within the large four-generation pedigree, a carrier had a reproductive outcome that was approximately equal for 1) the balanced translocation, 2) normal chromosomes, and 3) viable 4q trisomy or pregnancy loss. The three individuals with chromosomal confirmation of trisomy 4q28-->qter (comprising approximately 1.8% of the haploid autosomal length) had similar mental and developmental retardation, hypotonia, restricted speech, seizures, and facial anomalies but no cardiac, renal, or skeletal anomalies. It is suggested that these latter severe malformations, associated with the classic 4q2 to 3 group of anomalies, were from an imbalance outside 4q28-->qter and were not necessarily related to the relatively large size of the trisomic segment. Multiple different chromosomes are reported to be rearranged with 4q in the production of distal 4q trisomy. The incidence of 4q rearrangement remains unexplained, but once it is present in a family, viability of a large trisomy in 4q seems to explain the number of affected individuals reported.

Founder's Award, Society for Biomaterials. Sixth World Biomaterials Congress 2000, Kamuela, HI,May 15-20, 2000. Really smart bioconjugates of smart polymers and receptor proteins.

Over the past 18 years we have been deeply involved with the synthesis and applications of stimuli-responsive polymer systems, especially polymer-biomolecule conjugates. This article summarizes our work with one of these conjugate systems, specifically polymer-protein conjugates. We include conjugates prepared by random polymer conjugation to lysine amino groups, and also those prepared by site-specific conjugation of the polymer to specific amino acid sites that are genetically engineered into the known amino acid sequence of the protein. We describe the preparation and properties of thermally sensitive random conjugates to enzymes and several affinity recognition proteins. We have also prepared site-specific conjugates to streptavidin with temperature-sensitive polymers, pH-sensitive polymers, and light-sensitive polymers. The preparation of these conjugates and their many fascinating applications are reviewed in this article.

Maternal age specific risk rate estimates for Down syndrome among live births in whites and other races from Ohio and metropolitan Atlanta, 1970-1989.

Our primary objective was to estimate, by one year and five year intervals, maternal age specific risk rates for Down syndrome among whites and among other races from two different populations, metropolitan Atlanta and south west Ohio, using live birth and prenatally diagnosed cases ascertained during 1970-1989. The five year estimates were also calculated separately for each of the five four year periods during these 20 years. Additionally, we compared two different methods of estimating these risk rates by using a third population of whites, and compared two different statistical methods of smoothing the risk rates. The results indicate good agreement between the metropolitan Atlanta and south west Ohio estimates within races, but show a statistically significant difference between the two race categories. Because 86% of live births in the "other races" category in the combined population are to blacks, these data may be seen as the first estimates of maternal age specific risk rates for Down syndrome among blacks calculated by one year intervals. We found excellent agreement in the risk rate estimates among the five four year time periods, between the estimates obtained by using the two different methods of estimation, and between the estimates obtained using the two different methods of statistical smoothing. Our estimated risk rates for white women in their 20s strongly reinforce those from previous studies currently being used for genetic counselling purposes. While we did find somewhat higher rates for women under 20, and increasingly higher rates for those over 30 years of age, these differences are not substantial. Thus, this study in general supports the risk rates estimated from data collected mostly during the 1960s and 1970s.

Acceptance of amniocentesis by women in the state of Montana (USA) who are screen positive for Down's syndrome.

OBJECTIVE: To assess factors influencing uptake of amniocentesis after a positive Down's syndrome screening result. METHODS: Interviews of 53 Montana women with screening risks > or = 1 in 300 after delivery. RESULTS: Thirty had accepted amniocentesis ("yes" group) and 23 had declined ("no" group) (57% uptake). Age at delivery was significantly higher (p = 0.02) for the "no" than the "yes" group (mean 35.3 nu 31.7 years). The mean risk of Down's syndrome ascertained by screening was 1 in 190 for the "no" group and 1 in 115 for the "yes" group (p = 0.05). Statistically significant differences (p < or = 0.05) between opinions in the two groups included: (a) desire to know if the fetus had Down's syndrome; (b) perception of the burden of care for an affected child; (c) support of doctor, spouse, and relatives for choice about amniocentesis; (d) attitudes toward abortion; (e) importance of religion; and (f) concerns about the amniocentesis procedure. The most important factor for those choosing amniocentesis was knowing if the fetus had Down's syndrome, and for those not choosing amniocentesis, attitude about abortion. CONCLUSION: Our results show the need for prescreening education to enable pregnant women to make informed decisions about screening for Down's syndrome and diagnostic testing.

Incidence and significance of chromosome mosaicism involving an autosomal structural abnormality diagnosed prenatally through amniocentesis: a collaborative study.

Among 179,663 prenatal diagnosis cases collected from ten institutions and two publications, 555 (0.3 per cent) were diagnosed as having chromosome mosaicism. Of these, 57 (10.3 per cent) were mosaic for an autosomal structural abnormality, 28 (5 per cent) for a sex chromosome structural abnormality, and 85 (15.3 per cent) were mosaic for a marker chromosome. Ninety-five cases of prenatally diagnosed mosaicism with a structural abnormality in an autosome and a normal cell line, and with a known phenotypic outcome, were collected for karyotype-phenotype correlations through our collaboration (40 cases), a prior survey (26 cases), and published reports (29 cases). They included 13 balanced reciprocal translocations, one unbalanced reciprocal translocation, four balanced Robertsonian translocations, four unbalanced Robertsonian translocations, four inversions, 17 deletions, three ring chromosomes, 19 i(20q), seven +i(12p), six other isochromosomes, and 17 partial trisomies resulting from a duplication or other rearrangement. All cases mosaic for a balanced structural rearrangement resulted in a normal phenotype. All cases of 46/46,i(20q) resulted in normal liveborns. Five of seven cases with 46/47,+i(12p) had an abnormal phenotype compatible with Killian-Pallister syndrome. The overall risk for an abnormal outcome for a mosaic case with an unbalanced structural abnormality, excluding 46/46,i(20q) and 46/47,+i(12p), is 40.4 per cent. In the same category, the study also suggested a correlation between the percentage of abnormal cells and an abnormal phenotype. For mosaicism involving a terminal deletion, the possibility of a familial fragile site should be considered.

Tissue specificity and stability of mosaicism in Pallister-Killian +i(12p) syndrome: relevance for prenatal diagnosis.

We ascertained +i(12p) mosaicism during third trimester in a case of polyhydramnios and diaphragmatic hernia. Primary cultures of amniocytes had colonies with +i(12p), colonies without +i(12p), and mixed colonies with 46/47,+i(12p). The likely explanation was instability and loss of i(12p) during somatic divisions of amniocytes. Fetal blood in third trimester retained +i(12p) in 13% of cells. A review of mosaicism in published cases indicates that factors influencing the presence of +i(12p) include tissue type and in vitro and in vivo age. In blood, amniocyte, and probably bone marrow cultures, +i(12p) is less stable than in fibroblast-like cultures derived from skin and other tissues. Young cultures at early passage are more likely to have +i(12p) than old cultures. Cultures from young (especially fetal) donors are more likely to retain +i(12p) than cultures from adult donors. These rules will be important in determining appropriate tissues for diagnosis and interpretation of mosaicism in this disorder.

Pre- and postnatal diagnosis of trisomy 4 mosaicism.

A liveborn girl with 46,XX/47,XX+4 mosaicism is reported for the first time. The diagnosis of true mosaicism was established initially in the assay of cultured amniotic fluid cells, although fetal blood obtained by percutaneous umbilical blood sampling showed a 46,XX chromosome constitution. The liveborn infant had manifestations previously reported in dup(4p) and dup(4q) syndromes. Cells in cord and arterial blood samples also were 46,XX, but cultures of placenta and bilateral forearm skin biopsies showed 46,XX/47,XX,+4 mosaicism. This case illustrates the disadvantage of chromosome analysis from blood alone when tissue-specific mosaicism is present.

Cell kinetic disturbances induced by treatment of human diploid fibroblasts with 5-azacytidine indicate a major role for DNA methylation in the regulation of the chromosome cycle.

BrdU-Hoechst flow cytometry was used to investigate the effects of DNA hypomethylation, induced by treatment with 5-azacytidine (5AC), on cell proliferation. When human fibroblast-like cells derived from skin and amniotic fluid were exposed to 5AC during three successive cell cycles, their clone-forming ability was diminished after removal of the drug. Treated cells were rendered quiescent by culture with low serum in the absence of the drug. Upon serum stimulation, they showed a diminished fraction of proliferating cells, which exhibited a prolonged transit through the S and G2 phase of the cell cycle, and a permanent arrest within the G2 compartment. This pattern of disturbed cell proliferation may in part explain the changes in replication banding pattern reported in the literature. Cytogenetic analysis of 5AC-treated cells revealed numerous endomitoses and tetraploid metaphases indicating a disturbed chromosome cycle in association with these cell kinetic perturbations.

Prenatal ascertainment of triploidy by maternal serum alpha-fetoprotein screening.

We report our experience in ascertaining fetal triploidy during routine maternal serum alpha-fetoprotein (MSAFP) screening. Three cases were identified after elevated MSAFP tests. Two of the three had normal amniotic fluid alpha-fetoprotein (AFAFP). The third had amniocentesis too late for AFAFP interpretation. Three additional cases were detected by amniocentesis without prior MSAFP screening and none had an elevated AFAFP. A literature review revealed eight triploid fetuses detected as a result of an elevated MSAFP. Of the five with AFAFP quantitation, only one had an abnormal value and the elevation was minimal. In these 14 cases from our own and other reports, ultrasound findings of placental and fetal abnormalities were often noted, but a pattern diagnostic of triploidy was not present. We conclude that, for optimal prenatal detection of triploidy, fetal karyotyping should be included when an amniocentesis is performed for elevated MSAFP.

Differentiation in human chorionic villus cultures: hCG and HLA expression.

The present report describes methods to separate, culture, and study syncytio-cytotrophoblast and mesenchymal core of the first-trimester human chorionic villus. The cultured outer layer cells (syncytio-cytotrophoblast) are multinucleated, pleomorphic, and active in the formation of human chorionic gonadotrophin (hCG). The mesenchymal core cells are more fibroblast-like in appearance, do not show multinucleation, and have less hCG in their culture media. Both cultured cell types express HLA (ABC) Class I histocompatibility antigens but not HLA (DR) Class II antigens. These and previous studies from this laboratory postulate different embryonic origins: (1) Syncytio-cytotrophoblast cultures of chorionic villus derive from differentiated trophoblast and preserve multinucleation as well as hCG hormone function. (2) Cells cultured from the chorionic villus core originate from extraembryonic mesenchyme. (3) Amniocytes (AF cells) cultured from amniotic fluid resemble the multipotential and early-stage trophoblast, retaining pleomorphism, multinucleation, and lacunae formation as well as production of hCG, progesterone, oestrogen, basement membrane glycoprotein, and Type IV collagen. These cell types cultured from the chorionic villus and amniotic fluid provide a means for in vitro study of specific embryonic cell lineages.

Treatment of a mandibular fracture in a neonate.

The treatment and long-term follow-up of mandibular fracture in a neonate have been presented. Early treatment can prevent the need for later extensive and expensive reconstructive surgery.

Molecular evidence for true isochromosome 21q.

In one family a duplicated 21q was shown to be a true isochromosome, which segregates from mosaic mother to non-mosaic child with full Down syndrome phenotype. Densitometric analysis of Southern blots, using probe pPW228C for the distal long arm of chromosome 21, indicated that the 21q duplication contains two copies of the allele detected by the probe. Maternal mosaic karyotype of 45,XX,-21/46,XX/46, XX,-21,+21i(21q) also suggested transverse mitotic centromere division as the origin of the 21q isochromosomes. Morphologic analysis of chromosome heteromorphisms strengthened this interpretation because the free 21 missing in the cell line with 45 chromosomes was also missing in cells with the isochromosome. In a second family the cytogenetic data also suggested transmission of an i(21q) from mosaic mother to non-mosaic Down syndrome child but molecular evidence did not prove identity of alleles in the duplicated chromosome 21.

Chromosome and growth factor abnormalities in melanoma.

Cultures from metastatic melanomas of 15 patients had detailed melanoma growth stimulatory activity (MGSA) and cytogenetic analysis. The presence of melanoma cells was confirmed by microscopic identification of melanin, tyrosinase activity, and electron microscopy characterization of melanosomes. The MGSA is found in cytoplasmic granules after immunocytochemical stain. Three of the cultures did not produce MGSA and showed no distinctive cytogenetic differences. Breakpoints in derivative chromosomes were concentrated in region 1p1, and among all cultures chromosome 1 was the most frequently rearranged. It also has a low copy number of normal homologs. Chromosomes 18, X, and Y were never derivative, and chromosomes 2 and 4 were rarely so. Thus the cytogenetic data indicate that 4q13-21, the hybridization site for MGSA cDNA, is spared from gross change, although it could be under the influence of another site on chromosome 1 that is lost or rearranged. The ratio of abnormal to normal autosomes (mean per cell) in no culture exceeded 0.5, and for no autosome exceeded 0.8, suggesting a limit to the rearrangement tolerated for cell survival. If the Y is retained, the X:Y ratio varies around a normal figure of 1. The ratio of autosomes to sex chromosomes varies around a normal figure of 22. These data suggest stability of the X chromosome in cells undergoing multiple rearrangements of the autosomes.

2D NMR studies of aminoglycoside antibiotics. Use of relayed coherence transfer for 1H resonance assignment and in situ structure elucidation of amikacin derivatives in reaction mixtures.

Phase-sensitive 2D 1H/1H COSY spectra can be used to identify the structures of individual pure specimens of the aminoglycoside antibiotic amikacin and its N-hemisuccinyl derivatives. However, even at 500 MHz the 2D chemical shift dispersion does not allow for unambiguous assignment of all cross-peaks. By use of 2D relayed coherence transfer experiments (RELAY) optimized to detect two-step 1H/1H scalar interactions in which one of the J-values is small, sufficient additional correlations can be obtained from the frequency-isolated resonances to allow facile tracing of all scalar connectivities. Complete assignments of the 1H NMR spectra of amikacin, its 6'-N-hemisuccinamide, and a novel bis(acylate) [gamma-N-(p-vinylbenzoyl)amikacin 6'-N-hemisuccinamide] were obtained for aqueous media. The NMR spectrum of amikacin free base was also assigned in dimethyl sulfoxide solution. The RELAY experiment can be extended to the analysis of reaction mixtures, which allows for the identification and resonance assignment of regioisomeric amikacin haptens in the mixture state. All of the N-monohemisuccinyl isomers of amikacin have been identified in reaction mixtures through the RELAY experiment. The relative reactivities of the amino functions of amikacin toward acylating agents were found to be 6'-N greater than 3-N equal to or greater than 3"-N equal to or greater than gamma-N. However, this reactivity order is altered after the initial acylation event.