Uridine-5-H3 radioautography of the human sex chromatin body.

To obtain an estimate of the rate of RNA synthesis by the heterochromatic sex chromatin body, human female fibroblasts were labeled with uridine-5-H(3) and radioautographed. The number of grains over the sex chromatin body was compared with the number of grains over a comparable area of euchromatin. The ratio was 0.37. When corrected for the higher content of DNA per unit area in heterochromatin, the maximum rate of RNA synthesis by the DNA of the sex chromatin body was approximately 18% of the rate of RNA synthesis by a comparable amount of euchromatin DNA. The rate of RNA synthesis by the sex chromatin body did not increase significantly with partial despiralization of this chromatin at prophase.

The fine structure and chemical composition of nuclei during spermiogenesis in the house cricket. I. Initial stages of differentiation and the loss of nonhistone protein.

The early stages of nuclear differentiation in spermatids of the house cricket are described with regard to the fine structural elements and chemical components which occur. Particular attention is given to the loss of nonhistone protein from the nucleus and its relation to chromatin structure. Granular elements about 25 to 80 mmicro in diameter, and fibers about 8 mmicro in diameter occur in the earliest spermatid nucleus. The fibers are found in diffuse and condensed chromatin while granules are found only in diffuse material. DNA and histone parallel the chromatin fibers in distribution, while nonhistone protein and RNA parallel the granules in distribution. The granules and most of the nonhistone protein are lost, simultaneously, after the early spermatid stage. The protein loss occurs without detectable change in the structure of chromatin fibers. Chromatin fibers first show a structural change in mid spermiogenesis, when they become thicker and very contorted. Unusually thin fibers (about 5 mmicro) also appear in mid spermatid nuclei; they are apparently composed of nonhistone protein and free of DNA and histone.

Satellite DNA in constitutive heterochromatin of the guinea pig.

Total DNA and DNA from the heterochromatin and euchromatin fractions of male guinea pig liver nuclei were analyzed by cesium sulfate-silver density-gradient centrifugation. Total DNA is composed of three components: a heavy satellite DNA, a main DNA of intermediate density, and a light satellite DNA. Heterochromatin DNA shows a fourfold enrichment in the satellite components while euchromatin DNA is relatively devoid of them. The strands of both satellite DNA's are separable by centrifugation in alkaline cesium chloride. Base analyses on the separate strands demonstrate that the two satellite DNA's represent different species.

Cytogenetic evidence for premeiotic transformation of human testicular cancers.

To determine the point at which transformation of the germ cell occurs during meiosis in nonseminomatous testicular cancer, the sex chromosome compositions of 15 cell lines derived from primary tumors or metastases of 12 patients with testicular cancer were analyzed by trypsin G-banding analysis and Y-body staining. The simultaneous existence of both X- and Y-chromosomes in a single cell has been confirmed in 14 cell lines. This suggests that transformation of the cell occurs before the first meiotic division because it is known that segregation of X- and Y-chromosomes occurs during the first meiotic division. An incidental finding was the presence of Barr bodies in some cell lines containing more than one X-chromosome, which is consistent with the known primitive nature of testicular cancer and its ability to differentiate independently from the male host.

X chromatin and chromosome examination in aged women.

X chromatin frequencies were compared between aged, middle-aged and young women and statistically significant decrease in frequency was found in the aged as compared to the middle-aged and the young women. For a subsample of aged women results of chromosome examinations were also available which allowed comparison of the frequencies of X chromatin and monosomy C; no significant correlation emerged. This latter finding indicates that karyotypic analysis is necessary for the determination of monosomy C; and that X chromatin counts from buccal mucosal cells cannot be substituted for the examination of karyotypes.

A DNA probe from Schistosoma mansoni allows rapid determination of the sex of larval parasites.

A DNA clone representing a 0.4 kb degenerative repeat has been isolated. The DNA sequence is present only in the genome of female Schistosoma mansoni at different stages of the life cycle, at a frequency of approximately 75 copies per adult female genome. The sequence is not expressed and probably represents satellite DNA in the heterochromatin region of the W chromosome. It is demonstrated that the DNA clone may be used for the rapid determination of the sex of cercariae without the need for DNA isolation or Southern blotting.

Lymphoid chimerism after allogeneic bone marrow transplantation. Y-chromatin staining of peripheral T and B lymphocytes and allotyping of serum immunoglobulins.

Lymphoid cell engraftment was monitored for several years after bone marrow transplantation by Y-chromatin staining of T and B lymphocytes in the peripheral blood and/or by immunoglobulin allotyping in the serum of 20 of 52 pediatric patients grafted successively between October 1973 and October 1983. Data on 2 patients with severe combined immunodeficiency, grafted earlier in December 1968 and April 1971, are also included. These children received an allogeneic bone marrow graft for leukemia (n = 7), severe aplastic anemia (n = 11), or severe combined immunodeficiency (n = 4) and were informative for this study, because they differed from their donor by sex (n = 16) and/or by immunoglobulin phenotype (n = 13). Of 16 pairs in which the donor was of the opposite sex, 11 patients ultimately showed circulating T and B lymphocytes of donor origin after bone marrow transplantation; in the remaining 5, there was an incomplete chimerism of the circulating lymphoid cells. Of 13 pairs with a difference in immunoglobulin phenotype between donor and recipient, 8 patients exhibited donor allotypes 3 months or later after transplantation, in 3 of them together with recipient allotypes. In the remaining 5 patients, recipient allotypes were detected after transplantation, but the simultaneous presence of donor-type immunoglobulin production could not be excluded in 4. The persistence of either a split (T lineage of donor origin and B lineage of recipient origin) or mixed (T and/or B lineage of donor and recipient origin) chimerism was related to the type of disease. In 3 children circulating B cells of donor-origin did not fit with the recipient origin of the sessile immunoglobulin-secreting plasma cells. This implies that different immune compartments--e.g., bone marrow and peripheral lymphoid tissues--should be investigated following allogeneic bone marrow transplantation. A prolonged presence of recipient-type lymphoid cells increased the risk of leukemic relapse in the patients investigated.

DNA replication sequence in a dicentric (functionally monocentric) X chromosome formed by the joining of two X chromosomes at region p22.

In this study we used densitometry to evaluate DNA replication kinetics in a rearranged chromosome formed by the joining of two X chromosomes at region p22. No 45X mosaicism is present in peripheral blood or fibroblast cultures. The patient has primary amenorrhea, short stature, and gonadal dysgenesis. The sequence of replication in the majority of cells is p11, q11, q13, q22-24, q12, p22, q26, q28, q27, q25, and p21, q21. Thus p11 is the earliest region to replicate, and q21 is the last. In 66% of 127 cells analyzed, the replication pattern is asymmetric, and bands q12, q26, and q28 are most likely to be out of phase on the two sides of the breakpoint. We find that band p22 has a delay of replication compared to an abnormal X derived from two X chromosomes joined at the q23 region previously reported by us. Structural rearrangement may therefore delay replication in the region of the break.

Origin of the retrocorneal membrane in the rabbit.

Electron microscopic studies have offered presumptive evidence that the origin of both cellular and extracellular components of the retrocorneal membrane (RCM) derive from corneal endothelial cells. In an attempt to demonstrate conclusively the origin of the cell in the RCM, we performed 8-mm exchange, penetrating keratoplasties between male and female rabbits. After allowing for complete healing and using only clear corneas, we produced central RCM by freezing the central cornea after inducing intraocular inflammation. Care was taken to ensure that the membrane was surrounded by normal donor endothelium and Descemet's membrane was intact. Sex chromatin counts of the cells in the RCM of a female host with a male graft showed an average of 2% Barr bodies. Cells in the membrane of a male host with a female graft showed an average of 40% Barr bodies. The experimental and control sex chromatin counts were almost identical. This study provides conclusive evidence that central retrocorneal, fibroblast-like cells and membranes in rabbits are derived from corneal endothelial cells.

Turner phenotype in mother and daughter.

Two females are described, mother and daughter, who had the Turner phenotype and spontaneous sexual development. The mother is short and had ovulatory menstrual cycles, normal breast development, X-chromatin negative buccal smear, 45,X chromosomal pattern in her peripheral blood lymphocytes, and 45,X/46,X,r(X) mosaicism in her skin, with the majority of the cells (85%) showing X monosomy. She had a successful uncomplicated pregnancy at the age of 25 years. The daughter is short and had spontaneous sexual development, including menstruation at the age of 15 years. Her buccal smear was X-chromatin negative and karyotypes from peripheral blood lymphocytes and skin fibroblasts showed a 45,X chromosome constitution. Her menstrual cycles are irregular and, most probably, anovulatory. She has a horseshoe kidney. Six women with a 45, X chromosome complement are known to have delivered normal infants with no chromosomal abnormality. Five children with 45,X mosaicism have been born to mothers with 45,X mosaicism; all had a 46,XX cell line as well. This is the first report of a 45,X female born to a mother with mosaicism composed of 2 abnormal cell lines, 1 with X monosomy and 1 with a ring X chromosome.

Clinical and laboratory evaluation of patients with primary amenorrhea.

Sixty-two patients with primary amenorrhea were retrospectively categorized into 4 groups: 1) breast development absent and uterus present (29 patients), 2) breast development present and uterus absent (9 patients), 3) both breast development and uterus absent (2 patients), and 4) both breast development and uterus present (22 patients). Patients in category 1 were diagnosed as having hypogonadotropic hypogonadism (low or normal follicle-stimulating hormone [FSH]) or gonadal dysgenesis (elevated FSH). Patients in category 2 were diagnosed as having congenital absence of the uterus (female range testosterone [T] or testicular feminization [male range T]). In the 2 patients in category 3, a 46,XY karyotype occurred with an enzyme defect (17,20 desmolase) in 1 and the other had agonadism. In category 4, 5 patients with elevated prolactin and a pituitary adenoma were identified. The remaining 17 patients were divided into 2 groups based on progesterone withdrawal bleeding. Patients who had withdrawal bleeding and had elevated luteinizing hormone level were diagnosed as having polycystic ovaries and patients with normal gonadotropins as having hypothalamic dysfunction. Patients who did not bleed were diagnosed as having hypothalamic failure (normal or low FSH) or primary ovarian failure (elevated FSH). This study demonstrates that it is possible to classify patients with primary amenorrhea into 4 useful diagnostic categories based on physical examination and a minimal laboratory investigation.

Cytogenetic analysis in human bone marrow transplantation.

Chromosome and sex chromatin studies have contributed significantly to the evaluation of human bone marrow transplants. In addition to their use in documenting bone marrow engraftment following transplantation, they have been used to (1) evaluate whether recurrent leukemia occurs in donor or recipient cells and whether recovery in aplastic anemia results from growth of host or donor cells, (2) demonstrate the bone marrow origin of pulmonary macrophages and hepatic Kupffer cells, (3) show that bone marrow fibroblasts have an origin different from that of the hematopoietic bone marrow cells, (4) evaluate twin zygosity in preparation for transplantation, and (5) show that the defect in Fanconi's anemia is intracellular.

Barr bodies in testis with Klinefelter syndrome.

Barr bodies are described in the interstitial cells of testes of a chromatin-positive patient with Klinefelter syndrome. Ultrastructural studies confirm the origin of the Barr body from the nuclear chromatin. Ultrastructurally the interstitial cells showed diminished, smooth endoplasmic reticulum and lipid droplets probably indicating impaired function.

Study of the role of steroid receptors in human breast cancer.

Cytoplasmic estrogen receptors (ER) were studied in 200 female patients with primary breast cancer. Both ER and progesterone receptors (PgR) were studied in 38 patients. Of the 200 tumors, 55% were estrogen receptor-positive, of the 38 tumors, 34.2% were progesterone receptor-positive. The level of steroid receptors was compared with some parameters characterizing the host and the tumor. It was found that the level of ER correlated with the age factors and the menstrual status. The level of ER was higher in women with adrenal or involutive pathogenetic forms of breast cancer and in women with great body weight. There was not found any relation between the level of estrogen receptors and the stage of menstrual cycle as well as between the level of estrogen receptors and the content of sex chromatin in the tumors.

Decrease of Y chromatin frequency with time after fixation of blood smear.

The average frequency of Y chromatin positive lymphocytes was 59.4% among males aged 13 - 19 years. The frequency decreased significantly with time after fixation of the blood smear.

Sex determination from plucked human hairs without epithelial root sheath. III. Fluorescent feulgen reaction using acriflavine.

In order to determine sex from plucked human hair roots devoid of epithelial root sheath, the frequency of the X-chromatin was examined in hair cortex nuclei of plucked hairs stained by the fluorescent Feulgen reaction using acriflavine. There was a distinct difference between male and female hairs, the frequency ranging from 27% to 70% (average 45.8%) in female samples and from 0% to 8% (average 2.9%) in male samples. The difference in the frequency between male and female was detected in samples kept in a dried condition for 32 weeks.

Sex determination from plucked human hairs without epithelial root sheath. II. Depigmentation of melanin in the hair cortex before Feulgen reaction.

In hair roots devoid of the epithelial root sheath, an attempt was made to decolorize the melanin granules without affecting the Feulgen reaction for the sex chromatin. The hair samples were treated with 0.25% potassium permanganate for 1 hour, 0.3% hydrogen peroxide for 1 minute and 5% potassium permanganate for 1 hour, 0.3% hydrogen peroxide for 1 minute and 5% oxalic acid for 5 minutes, and then stained with Feulgen. The frequency of sex chromatins ranged from 22% to 47% (average 32%) in female samples and from 0% to 8% (average 5%) in male samples. Thus, the frequency distributions of the male and female samples were completely independent of each other. The sex chromatins in dried female hairs were detectable at a frequency of 16 - 26% several weeks after plucking. The depigmentation procedure almost completely bleached the melanin granules in the hair cortex, and produced no harmful effect on the Feulgen reaction that followed.

Post-transfusion graft-versus-host disease following open heart surgery. Report of six cases.

Six cases of post-operative erythroderma after open heart surgery are described. About 10 days after seemingly uneventful recovery, all patients developed fever, erythroderma, liver enzyme elevation, pancytopenia, and an aplastic bone marrow. Their condition rapidly deteriorated, and they died within 20 days of the onset of symptoms. Skin biopsy specimens from two patients showed mild leukocytic infiltration in the epidermal basal layer and upper dermis. Immunostaining by the ABC method showed that most of these infiltrating cells were suppressor/cytotoxic T cells. HLA study of peripheral lymphocytes from two patients and their families revealed that the patients' HLA phenotypes were incompatible from their children's HLA findings. Y chromatin was present in the lymphocytes of the skin biopsy specimen of a female patient. Based on the clinical picture, skin biopsy, HLA study, and Y chromatin study, the authors strongly suspect post-transfusion GVHD as the etiology of postoperative erythroderma, although these patients lacked any known immunodeficiency.

An XO/XX mosaic sheep with associated gonadal dysgenesis.

A female sheep with an apparent XO/XX karyotype was discovered while screening barren ewes. The animal exhibited a normal phenotype. This is the first reported case of mosaicism associated with gonadal dysgenesis in the sheep.

Prognosis and incidence of sex chromatin in breast cancer. A preliminary report.

Results of two studies, based on a total of 126 patients (prospective--76 cases and retrospective--50 cases), on the incidence of the sex chromatin in the cancerous tissue are presented. For evaluation of prognosis in the retrospective study with a five year follow up, sex chromatin incidence was correlated with other prognositc factors such as lymph node metastasis, the disease free interval of distant metastasis and the size of the tumor. Whereas positive significant correlation between sex chromatin incidence and a five year survival time or disease free interval of distant metastasis was observed, no correlation was seen between sex chromatin counts and lymph node metastasis or the size of the tumor.