Longitudinal Wobbling Motion in

The rare phenomenon of nuclear wobbling motion has been investigated in the nucleus ^{187}Au. A longitudinal wobbling-bands pair has been identified and clearly distinguished from the associated signature-partner band on the basis of angular distribution measurements. Theoretical calculations in the framework of the particle rotor model are found to agree well with the experimental observations. This is the first experimental evidence for longitudinal wobbling bands where the expected signature partner band has also been identified, and establishes this exotic collective mode as a general phenomenon over the nuclear chart.

Mouse offspring after microinjection of heated spermatozoa.

The thermostability of the mammalian sperm genome was previously reported, but no live offspring after conception with heated spermatozoa had yet been obtained. In the present study, mouse spermatozoa were heated at 56 degrees C for 30 min and microinjected into mouse oocytes. Fertilization did not occur unless activation was induced by incubation in a calcium-free medium containing strontium. Under these conditions fertilization and cleavage rates were comparable to those obtained after microinjection of control spermatozoa, but the developmental rate to the blastocyst stage was lower. When transferred to foster mothers, embryos derived from heated sperm developed into phenotypically normal offspring, which grew and reproduced normally. In the mouse, heated spermatozoa can therefore support full embryonic development after microinjection into oocytes.

Characterization of INTA 51-3, a new atypical strain of Bacillus thuringiensis from Argentina.

Several isolates of Bacillus thuringiensis native to Argentina obtained in a nationwide screening program showed atypical crystal morphology. One of these strains, INTA 51-3, was further characterized in order to determine other features like protein composition of its parasporal crystal, plasmid pattern, identification of cry genes and toxicological properties. B. thuringiensis INTA 51-3 (serovar tohokuensis) had an amorphous inclusion containing a major protein component of ca. 130 kDa. After trypsin digestion of solubilized crystals, SDS-PAGE resolved a unique protease-resistant peptide of ca. 90 kDa. The plasmid pattern from INTA 51-3 resembled that of the standard strain HD-1. However, Southern analysis showed no hybridization to fragments of cry1Aa, cry2Aa, cry3A, and cry11A genes. Degenerate primers were used for identification of the cry1 genes by PCR. Nevertheless, the presence of cry1 type gene(s) in B. thuringiensis INTA 51-3 was confirmed. Highly concentrated crystal suspensions showed to be weakly toxic only to lepidopteran species.

A trisomic germ cell line and precocious chromatid segregation leads to recurrent trisomy 21 conception.

A chromosomally normal 37-year-old woman was referred for preimplantation genetic diagnosis after having several conceptuses with trisomy 21. Segregation of chromosome 21 was assessed in unfertilised meiosis II oocytes and preimplantation embryos from PGD cycles using fluorescent in situ hybridisation (FISH). Of 7 preimplantation embryos, 5 were chromosomally abnormal with 4 having trisomy 21 and one being tetraploid. Of 4 oocytes, 3 had an abnormal chromosomal constitution with either an extra chromosome 21 or an extra chromatid 21. In one oocyte an extra chromatid 21 was detected in both the metaphase II complement and the first polar body providing the first direct evidence of a maternal trisomic germ cell line. Moreover, this result shows that the extra chromosome 21 can precociously divide into its two chromatids at the first meiotic division.

Preimplantation genetic diagnosis for couples at high risk of Down syndrome pregnancy owing to parental translocation or mosaicism.

The population risk for trisomy 21 is 1 in 700 births but some couples are at a much higher risk owing to parental translocation or mosaicism. We report on the first attempt to carry out preimplantation genetic diagnosis for two such couples using cleavage stage embryo biopsy and dual colour FISH analysis. Each couple underwent two treatment cycles. Couple 1 (suspected gonadal mosaicism for trisomy 21) had two embryos normal for chromosome 21 transferred, but no pregnancy resulted; 64% (7/11) unfertilised oocytes/embryos showed chromosome 21 aneuploidy. Couple 2 (46,XX,t(6;21)(q13;q22.3)) had a single embryo transferred resulting in a biochemical pregnancy; 91% (10/11) oocytes/embryos showed chromosome 21 imbalance, most resulting from 3:1 segregation of this translocation at gametogenesis. The opportunity to test embryos before implantation enables the outcome of female meiosis to be studied for the first time and the recurrence risk for a Down syndrome pregnancy to be assessed.

Meiotic behaviour of sex chromosomes investigated by three-colour FISH on 35,142 sperm nuclei from two 47,XYY males.

Meiotic segregation of sex chromosomes from two fertile 47,XYY men was analysed by a three-colour fluorescence in situ hybridisation procedure. This method allows the identification of hyperhaploidies (spermatozoa with 24 chromosomes) and diploidies (spermatozoa with 46 chromosomes), and their meiotic origin (meiosis I or II). Alpha-satellite probes specific for chromosomes X, Y and 1 were observed simultaneously in 35,142 sperm nuclei. For both 47,XYY men (24,315 sperm nuclei analysed from one male and 10,827 from the other one) the sex ratio differs from the expected 1:1 ratio (P < 0.001). The rates of disomic Y, diploid YY and diploid XY spermatozoa were increased for both 47,XYY men compared with control sperm (142,050 sperm nuclei analysed from five control men), whereas the rates of hyperhaploidy XY, disomy X and disomy 1 were not significantly different from those of control sperm. These results support the hypothesis that the extra Y chromosome is lost before meiosis with a proliferative advantage of the resulting 46,XY germ cells. Our observations also suggest that a few primary spermatocytes with two Y chromosomes are able to progress through meiosis and to produce Y-bearing sperm cells. A theoretical pairing of the three gonosomes in primary spermatocytes with an extra sex chromosome, compatible with active spermatogenesis, is proposed.

Increased aneuploid frequency in spermatozoa from a Hodgkin's disease patient after chemotherapy and radiotherapy.

The frequency of sperm aneuploidy was investigated by fluorescence in situ hybridization (FISH) in a Hodgkin's disease patient shortly after he had received chemotherapy and radiotherapy. Sperm karyotyping of the same patient had previously shown multiple structural abnormalities in most spermatozoa immediately after radiotherapy (day 0), whereas most spermatozoa collected 5 wk later (day 38) exhibited normal metaphase divisions (Rousseaux et al., 1993). Variations in the frequency of aneuploidy could not be detected by sperm karyotyping. Multicolor FISH on interphase spermatozoa revealed an increase in the rate of disomy for chromosomes 1, 6, 11, X, and Y at day 0 as well as at day 38. The high frequency of 24,XY (nondisjunction at meiosis I) and 24,XX (nondisjunction at meiosis II) spermatozoa (8.46% and 1.64% at day 0, respectively) from the Hodgkin's disease patient suggests that both meiosis I and II are affected and that the X chromosome is frequently involved in such malsegregation events. The rate of 46,XY diploidy was also increased in the patient's sperm, up to 0.62% at day 0. While radiotherapy probably affected the postmeiotic cells (spermatids), the patient's cancer and/or chemotherapy are the two major factors that could have affected the dividing spermatogonia and/or spermatocytes, resulting in high aneuploidy rates.

[Intracytoplasmic sperm injection and Klinefelter syndrome]. / ICSI et syndrome de Klinefelter.

Sperm cytogenetic study in two patients with Klinefelter's syndrome have demonstrated that there existed a risk low but highly significant, to transmit a sex chromosome abnormality to the offsprings. This result argues for a systematic karyotype before ICSI, and if such mosaïcs can be treated by ICSI, they must be firstly associated to a genetic counselling.

Increased incidence of hyperhaploid 24,XY spermatozoa detected by three-colour FISH in a 46,XY/47,XXY male.

Meiotic segregation of gonosomes from a 46,XY/47,XXY male was analysed by a three-colour fluorescence in situ hybridisation (FISH) procedure. This method allows the identification of hyperhaploid spermatozoa (with 24 chromosomes), diploid spermatozoa (with 46 chromosomes) and their meiotic origin (meiosis I or II). Alpha satellite DNA probes specific for chromosomes X, Y and 1 were observed on 27,097 sperm nuclei. The proportions of X- and Y-bearing sperm were estimated to 52.78% and 43.88%, respectively. Disomy (24,XX, 24,YY, 24,X or Y,+1) and diploidy (46,XX, 46,YY, 46,XY) frequencies were close to those obtained from control sperm, whereas the frequency of hyperhaploid 24,XY spermatozoa (2.09%) was significantly increased compared with controls (0.36%). These results support the hypothesis that a few 47,XXY germ cells would be able to complete meiosis and to produce mature spermatozoa.

Sperm nuclei analysis of a Robertsonian t(14q21q) carrier, by FISH, using three plasmids and two YAC probes.

The meiotic segregation of chromosomes 14 and 21 was analysed in 1116 spermatozoa from an oligoasthenospermic carrier of a Robertsonian translocation t(14q21q), and in 16,392 spermatozoa from a control donor, using two-colour fluorescence in situ hybridisation (FISH). Two YAC probes (cloned in yeast artificial chromosomes) specific for regions on the long arms of these chromosomes were co-hybridised. Of the spermatozoa, 12% were unbalanced, resulting from adjacent segregations. Chromosomes X, Y and 1 were also simultaneously detected in 1335 spermatozoa from the same carrier. Whereas gonosomal disomy rates were not significantly different from those of the control donors, disomy 1 were slightly but significantly increased to 0.7%. The diploidy rate was also slightly increased to approximately 1% in the translocation carrier.

Human sperm chromosome analysis after microinjection into hamster oocytes.

PURPOSE: Subzonal sperm insemination (SUZI) into hamster oocytes was performed to establish the karyotypes of the fertilizing spermatozoa. METHODS: Spermatozoa from two males with normal semen parameters were microinjected. Of 72 (52 + 20) analyzed sperm chromosome metaphases, only 1 (1.4%) was considered abnormal, showing a structural abnormality. RESULTS: No hyperhaploidy was observed. Rates of sperm chromosomal abnormalities after microinjection were not higher than those reported previously using zona-free egg insemination, suggesting that the SUZI procedure per se does not increase sperm chromosomal abnormalities. CONCLUSIONS: The use of subzonal insemination into hamster oocytes for the study of human sperm chromosomes in males with low sperm counts is discussed.

Meiotic segregation of the X and Y chromosomes and chromosome 1 analyzed by three-color FISH in human interphase spermatozoa.

Meiotic segregation of the X and Y chromosomes and chromosome 1 was analyzed by three-color fluorescence in situ hybridization (FISH) in 94,575 human interphase spermatozoa from four control subjects. More than 99% of the sperm cells were labeled. The proportions of X- and Y-bearing sperm were estimated to be 49.83% and 48.30%, respectively. The disomy rates were 0.04%, 0.009%, and 0.20% for the X and Y chromosomes and chromosome 1, respectively. Hyperhaploidy with an extra gonosome was found in 0.34% of spermatozoa, due to nondisjunction during meiosis I. The frequency of diploidy was 0.11% at meiosis I and 0.036% at meiosis II. Cohybridization of one autosomal and two gonosomal probes, in three-color FISH in interphase spermatozoa, seems to accurately discriminate diploidies from disomies, as well as the meiotic origin of gonosomal aneuploidies in sperm cells.

Meiotic segregation in males heterozygote for reciprocal translocations: analysis of sperm nuclei by two and three colour fluorescence in situ hybridization.

The meiotic segregation of chromosomes was analysed in three reciprocal translocation carriers, using FISH on interphase spermatozoa. The segregation pattern was first studied in 27,844 spermatozoa from two siblings carrying the reciprocal translocation t(6;11)(q14;p14). Three centromeric probes, specific for chromosomes 6, 11 and 1, were simultaneously hybridized so that all centric fragments as well as the ploidy of each cell could be determined by three colour FISH. For both subjects, the respective frequencies of alternate/adjacent 1, adjacent 2, 3:1 and 4:0 segregation modes were 88%, 9%, 3+ and < 1%. In another reciprocal translocation t(2;14)(p23.1;q31), a two colour FISH analysis was performed on 4,610 spermatozoa, using a chromosome 2 centromeric probe and a YAC probe located on the centric fragment of chromosome 14. Frequencies of alternate/adjacent 1, adjacent 2, and 3:1 segregations were 89%, 5.2%, and 5.8% respectively. The segregation of chromosomes X, Y and 1 were also analyzed with three colour FISH on the spermatozoa from all three translocation carriers, in order to detect an interchromosomal effect. Aneuploidy rates for the X and Y chromosomes were found to be in the same range in the three translocation carriers and control donors, but disomy 1 rates were slightly increased in the translocation carriers.

Male meiotic segregation of gonosomes analysed by two colour FISH in human interphase spermatozoa.

Human meiotic segregation of X and Y chromosomes was simultaneously analysed by dual fluorescence in situ hybridization (FISH) on 10,638 interphase spermatozoa from the same donor. A modified method for sperm decondensation ensured access of both X and Y probes to the sperm chromatin and a 99% hybridization efficiency. Expected sex ratios were obtained (49.30% haploidy X and 49.22% haploidy Y). The frequencies of meiotic II non-disjunctions for X and Y chromosomes (0.05%) were similar to those observed in sperm karyotypes after heterospecific fertilization of hamster eggs. In contrast, the frequency of XY bearing cells was significantly higher (0.42%). However, XY cells detected by FISH could either be diploid somatic cells, diploid germinal cells or hyperhaploid XY spermatozoa, the latter resulting from meiotic I non-disjunctions.

[Immune complex glomerulonephritis associated with pulmonary tuberculosis]. / Glomerulonefritis por inmunocomplejos asociada a tuberculosis pulmonar.

A 32 year old man was admitted for dyspnea, hemoptysis, macroscopic hematuria, hypertension (140/100), peripheral edema and hemodynamic decompensation. Lung Xrays revealed pulmonary edema and a cavity in the left apex. Laboratory determinations revealed an altered renal function with increased creatinine and urea levels and nephrotic syndrome. There was leucocyturia, hematuria and cylindruria. The sputum showed a large number of acid-fast bacilli. The patient began anti-tuberculosis treatment with three drugs (isoniacid, rifampicin, pirazinamide). On ultrasonography, both kidneys revealed ecogenic lesions with size, shape and cortico-medular relationship preserved. The patient persisted with altered renal function, steady levels of urea nitrogen, creatinine and potassium, preserved diuresis and hypertension. Bidimensional echocardiogram: LVDD 55 mm, hypoquinetic septum, pericardic effusion, thickened pericardium, pleural effusion, shortening fraction decreased. He received treatment for this congestive cardiac failure and hypertension with enalapril, nifedipine and fursemide. A percutaneous renal biopsy was performed with anatomopathologic diagnosis of diffuse encocapillar proliferative glomerulonephritis with crescents (15%) and total glomerular sclerosis (33%). Immunofluorescence: positive, immune-complexes with IgM and C3. The patient gradually recovered his normal renal function, improved his pleural effusions and normalized his cardiac function. He was discharged in good clinical condition on the 69th day of anti-tuberculosis treatment. An association between pulmonary tuberculosis and glomerulonephritis is discussed. It is proposed that renal lesions might be the consequence of the tuberculosis due to the sedimentation of circulating immune-complexes.

Achievement of meiosis in XXY germ cells: study of 543 sperm karyotypes from an XY/XXY mosaic patient.

Human sperm chromosomes from a 46,XY/47,XXY male were obtained using the technique of in vitro penetration of zona-free hamster eggs. The analysis of 543 sperm complements shows a significantly increased incidence (0.9%) of hyperhaploid gonosomal 24,XY sets, with a lack of the expected corresponding gonosomal hypohaploidies, and a normal rate of autosomal non-disjunctions. These results support the suggestion that 47,XXY cells are able to go through meiosis and to form spermatozoa. Only 24,XY sperm chromosomal constitutions were observed suggesting a preferential pairing of homologous sex chromosomes in 47,XXY spermatocytes.

Glomerulonefritis por inmunocomplejos asociada a tuberculosis pulmonar / Glomerulonephritis by immune complexes associated with pulmonary tuberculosis

Se presenta el caso de un varón de 32 años que ingresó por dispnea de esfuerzo, hemoptisis, hematuria macroscópica, hipertensión, edemas perifáricos y descompensación hemodinámica con radiología compatible con edema de pulmón y cavidad en vértice izquierdo. Los exámines de laboratorio mostraron función renal alterada con cifras elevadas de urea y creatinina y síndrome nefrótico. Se encontró leucocituria, hematuria y cilindruria intensas. Se hizo diagnóstico de tuberculosis pulmonar por baciloscopía positiva comenzando tratamiento tuberculostático. El paciente persistió con su función renal gravada con cifras estables de uremia, creatininemia y potasemia con ritmo diurético conservado e hipertenso. Se efectuó punción-biopsia renal con diagnóstico anatomopatológico de glomerulonefritis proliferativa difusa endocapilar con formación de semilunas (15 por ciento) y esclerosis glomerular total (33 por ciento). Por inmunofluorescencia se detectaron depósitos difusos de inmunocomplejos granulares localizados en mesangio, con positividad IgM ++ C3 +. El paciente se recuperó gradualmente hacia una función renal normal, disminuyó sus derrames pleurales y normalizó su función cardíaca con el tratamiento tuberculostático. Se comenta la asociación entre glomerulonefritis y tuberculosis pulmonar a la luz de la bibliografía disponible y se sugiere una relación causal entre ambas entidades

Immediate rearrangements of human sperm chromosomes following in-vivo irradiation.

The short-term effects of radiotherapy on sperm chromosomes was evaluated in two Hodgkin patients. Sperm cytogenetics were analysed by using human-hamster cross-fertilization. Immediately after irradiation, most sperm metaphases from the first patient exhibited multiple rearrangements, whereas in the second one, the incidence of such abnormalities was only slightly increased. Multi-fragmented chromosomes and structural aberrations of the chromatid-type were observed, suggesting a specific immediate response.

[Chromosome abnormalities of human gametes]. / Anomalies chromosomiques dans les gamètes humains.

Human newborns carrying chromosomal abnormalities are the survivors of a considerably larger cohort of affected conceptions. A direct cytogenetic study of these conceptuses would imply their destruction. Moreover, the ability to determine the parental origin of an additional or missing chromosome is limited from a methodological point of view. The cytogenetic study of human gametes provides information on these mechanisms. Several large-scale studies on sperm complements of normal men give an estimated abnormality rate of 10%. Our results concerning four carriers of reciprocal translocations show no evidence for a selection against abnormal sperm. Since about 30% of the recovered eggs fail to become fertilized in IVF programs, human oocytes have become available for large cytogenetic studies. Analysis of oocytes II provides information on the first meiotic non-disjunction rate. In our sample of 405 karyotypes, the rate of aneuploidy was 27%. No relationship was established between this frequency and the mode of stimulation of ovulation. No increase was observed with maternal aging.