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Biomedical subjects

C L Richer

Publications and source records attributed to C L Richer.

At least 55 records · Page 3Linked to original sources
PubMed1988

High-resolution R-banding at the 1250-band level. 1. Technical considerations on cell synchronization and R-banding (RHG and RBG).

A comparative analysis of variables for cell synchronization was made and led to the description of optimal conditions capable of yielding, in over 95% of cases, a large number of excellent quality cells in prometaphase and late prophase. Thymidine presents advantages over amethopterin as the synchronizing agent. The block was released with either thymidine or 5-bromo-2'-deoxyuridine (BrdU) for which various concentrations were tested. The presence of colcemid was also evaluated. Without colcemid, the optimal length of the release period was precisely determined so that the wave of synchronized cells could be harvested while going through the early stages of mitoses. Subsequently, GTG, RHG and RBG banding were produced on these elongated chromosomes. A comprehensive approach to RHG banding ensured an easier and well reproducible banding technique. Nine interrelated factors which influence banding quality were studied. The analysis of their effects on this banding pattern revealed new data for the understanding of its mechanism. A high-resolution R-banding technique after BrdU incorporation and Giemsa staining is presented; it is simple, reliable and reproducible. Different conditions for the FPG (Fluorochrome-Photolysis-Giemsa) technique were studied in order to obtain sharper borders and higher contrast between positive and negative bands. Optimal conditions for the incorporation of BrdU and the FPG method produced excellent band separation and band contrast even in very elongated prophase chromosomes. They did not decrease the mitotic index, did not increase chromosomal damage significantly, and did not greatly vary from subject to subject nor with the age of the slide (between 1 day and 36 months). Homologue discordance correlated to band range was compared for GTG, RHG and RBG banding.

Adult↗
PubMed1987

Two sisters with a distal deletion at the Xq26/Xq27 interface: DNA studies indicate that the gene locus for factor IX is present.

Two sisters with premature menopause and a small deletion of the long arm of one of their X chromosomes [del (X)(pter----q26.3:)] were investigated with polymorphic DNA probes near the breakpoint. The deleted chromosome retained the factor IX (F9) locus and the loci DXS51 (52A) and DXS100 (pX45h), which are proximal to F9. However, the factor VIII (F8) locus was not present, nor were two loci tightly linked to this locus, DXS52 (St14) and DXS15 (DX13). This deletion refines the location of the F9 locus to Xq26 or to the interface Xq26/Xq27, thus placing it more proximally than has been previously reported. The DNA obtained from these patients should be valuable in the mapping of future probes derived from this region of the X chromosome.

Adult↗
PubMed1987

Combined myeloid cell synchronization and chromosome G-banding by bromodeoxyuridine.

A combined high-resolution G-banding technique and cell synchronization of bone marrow cultures by BrdU is presented and compared with direct technique, standard 24-hour culture, and synchronization both with low temperature and methotrexate. Mitotic count and quality of spreading are improved in comparison with unsynchronized techniques. Precision of banding is increased significantly compared with the four other techniques used. Our procedure, which presents the advantage of combined synchronization and banding, is simple, time-saving, and provides more information about the chromosomes. Our results suggest that the GB-FPG technique should be a method of choice for the synchronization of bone marrow cells.

Bone Marrow↗
PubMed1987

Analysis of low concentrations of 5-bromo-2-deoxyuridine on sister chromatid exchanges in human lymphocytes.

To determine a concentration of 5-bromo-2-deoxyuridine (BrdU) sufficient for sister chromatid differentiation (SCD), and yet having a minimal effect on the number of sister chromatid exchanges (SCEs), we assessed the effect produced on the number of SCEs by low concentrations (1, 3, and 10 micrograms/mL) of BrdU. SCD was not obtained in 19% of the 31 subjects with 1 microgram/mL of BrdU, while the differentiation was adequate for all samples treated with 3 and 10 micrograms/mL. We statistically analysed the effects of these three different doses and found no significant difference in the number of SCEs obtained with the doses of 1 and 3 micrograms/mL, but a significant difference was observed between these two concentrations and 10 micrograms/mL. We therefore suggest that the dose of 3 micrograms/mL, while sufficient to produce reliable differential staining, still permits an adequate evaluation of the base line of SCEs and appears to enhance the sensitivity of the test to evaluate between-individual variations. Our experiments also underline that SCE counts should include the centromere exchanges.

Adolescent↗
PubMed1987

Idiograms of horse chromosomes at prometaphase, early metaphase, and midmetaphase after R-banding by BrdU incorporation followed by the fluorochrome-photolysis-Giemsa technique.

We present three idiograms of equine chromosomes, R-banded after BrdU incorporation and stained by the fluorochrome-photolysis-Giemsa technique. The haploid set of prometaphasic chromosomes shows 591 bands (range 7-38 per chromosome), the early metaphasic set 404 (range 5-26), and the midmetaphasic set 272 (range 3-18). Following cell synchronization with thymidine, more than twice as many R-bands were revealed on the resulting prometaphasic chromosomes, making possible the establishment of a very accurate and characteristic representation of this banding pattern in the domestic horse.

Animals↗
PubMed1987

Protein supplements, mitogens and antibiotics in lymphoblast transformation and cytogenetic analysis.

The in vitro effects of four protein supplements, foetal calf serum (FCS), gammaglobulin-depleted FCS (FCS-A), human serum albumin (HSA) and autologous serum (AS), and of four antibiotics on lymphocyte blast transformation and on lymphocyte proliferation for cytogenetic studies, were evaluated. 3H-thymidine incorporation was measured in the lymphocytes obtained from sixteen healthy adults and stimulated with phytohaemagglutinin (PHA), concanavalin A (Con A), pokeweed mitogen (PWM), PWM + PHA and PWM + Con A. The most efficient protein supplement was AS, although great variability was observed between subjects. HSA came second, while FCS and FCS-A were the least effective. The presence of immunoglobulins did not modify mitogen-induced lymphocyte responses. In unsupplemented media, peak responses to PHA and Con A occurred at lower mitogen concentrations. Furthermore, the addition of the antibiotics tested did not impede blast transformation whether or not a protein supplement was present. This work also showed that for chromosome analysis, protein supplements are necessary for optimum whole-blood lymphocyte cultures and that HSA can replace FCS.

Adult↗
PubMed1987

Human serum albumin as a substitute for foetal calf serum in blast transformation assays and cytogenetic analyses.

Diverse factors in serum produce complex effects on lymphocytes. Indeed, when used as a protein supplement for lymphocyte culture, serum has many disadvantages. A need exists for media in which serum is replaced by purified active serum components. The in vitro effects of human serum albumin (HSA) were evaluated and compared with the widely used foetal calf serum (FCS). Unstimulated and mitogen-activated lymphocyte proliferation was measured by 3H-thymidine incorporation. Phytohaemagglutinin (PHA), concanavalin A (Con A) and pokeweed mitogen (PWM) were used to transform the lymphocytes obtained from 36 healthy adults. Six concentrations of HSA, 0.5, 1, 2.5, 5, 10 and 20%, were studied. The last two concentrations produced an inhibition of lymphocyte proliferation. HSA, which was not itself mitogenic permitted a 4- to 5-fold higher transformation than 10% FCS. Maximum transformation occurred at HSA concentrations between 2.5 and 5% for PHA and PWM, and between 1 and 2.5% for Con A. In whole-blood lymphocyte cultures, the addition of 1% HSA gave a number of mitoses comparable to that obtained with the 10% FCS supplement. HSA is an interesting substitute for FCS: it is chemically well understood and standardized, species specific, readily available, relatively inexpensive, and has a long shelf life. HSA appears to be an excellent protein supplement for support of human lymphocyte proliferation in vitro, and can be recommended for routine use in lymphocyte cultures.

Adult↗
PubMed1987

Analysis of X-chromosome inactivation in horse embryos.

To define the time of X-chromosome inactivation in the horse, 122 conceptuses were collected transcervically between Days 6 and 28 (ovulation = Day 0) and subjected to cytogenetic analysis: 59 of the embryos were divided and in 41 of these separate cytogenetic analysis of the embryonic disc and remaining tissues was possible. Conceptuses were measured and photographed before capsule removal, culture in the presence of 5-bromodeoxyuridine and subsequent fixation for cytogenetic analysis. On average, 15 slides were prepared per conceptus. C-banding was used to determine the sex of each conceptus and, in the females, the effects of 5-bromodeoxyuridine incorporation were revealed by u.v. irradiation which produced R-bands and allowed identification of the inactive X-chromosome. Inactivation of the X chromosome began gradually in the trophoblastic (+/- attached endoderm and mesoderm) cells around Day 7.5 and in the embryonic disc around Day 11.5. Cells with an inactive X-chromosome predominated in the trophoblast by Day 10.5 and in the disc by Day 12.5.

Animals↗
PubMed1987

Light and electron microscopy of Ag-NORs in domestic horse chromosomes identified after R-banding.

Silver staining shows the presence in the domestic horse of six NORs located on chromosomes 1, 26 and 31 as identified after R-banding. Following electron microscopy, the argyrophilic material was observed outside the terminal secondary constrictions (satellite stalks) on the terminal portion of the short arm of chromosome 1, outside the secondary constrictions on the proximal region of the long arms of chromosome 31, and beside the proximal region of the long arms of chromosome 26. Satellite staining applied to these chromosomes appears to reveal only the active NORs.

Animals↗
PubMed1986

Deletion mapping of the testis determining locus with DNA probes in 46,XX males and in 46,XY and 46,X,dic(Y) females.

Eleven Y-specific DNA probes hybridizing with DNA from one or more 46,XX males were isolated from a recombinant phage DNA library constructed from flow sorted human Y chromosomes. Two probes hybridized with DNA from nine out of eleven, i.e. greater than 80% of these 46,XX males. The relative frequency of hybridization of the probes in the 46,XX males and in a 46,X,dic(Y) female, together with in situ hybridization data, allowed mapping of the probes on Yp in relation to a putative testis determining locus. Several of those probes were also absent in a 46,XY female, further refining a model for ordering the probes on Yp. The DNA of one XX male hybridized both with probes from Yp and probes from proximal Yq (excluding the pericentral region). This suggests that complex translocations may occur into the DNA of 46,XX males that involve not only parts of Yp but also parts of Yq.

Chromosome Aberrations↗
PubMed1986

Analysis of sister-chromatid exchanges (SCEs) in familial and sporadic multiple sclerosis.

The levels of sister-chromatid exchanges (SCEs) were calculated in 17 multiple sclerosis (MS) patients (14 familial and 3 sporadic cases) and 16 healthy controls matched for sex and age. The SCEs were significantly increased for MS patients (p = 0.0002), while there was no statistically significant difference between men and women and between younger and older subjects in both groups. Such factors as familial occurrence and severity of MS, smoking habits, and distribution of lymphocyte subpopulations were discussed. Although there was a significant difference between the MS patients taking medication and the patients taking none (p = 0.038), the latter were still significantly different from the controls (p = 0.035), supporting the fact that the disease itself increases SCEs. Our study, done with 2 doses of BrdU, also shows that the increased SCEs in MS patients are not due to a hypersensitivity to this substance known to be an inducer of SCEs. Thus we suggest that the increased SCEs found are probably disease-related.

Adolescent↗
PubMed1986

Effect of cannabinoids on spermatogenesis in vivo: a cytological study.

The cytogenetic effects of delta 9-tetrahydrocannabinol (THC) and cannabinol (CBN) (10 mg/kg) were investigated in hybrid mice of genotype (C57BL x C3H)F1. Mice were treated for 5 consecutive days with the specific cannabinoid; 16 days after the last treatment the meiotic cells were evaluated. Analysis of the spermatocyte bivalents at the first meiotic metaphase failed to reveal any numerical or structural abnormality. Contrary to previous reports we failed to find any major meiotic abnormalities associated with THC and CBN treatments. There was no evidence of ring or chain figures. The centromeric banding procedure (C banding) was used to support observations of Giemsa stained cells. It has previously been reported that cannabinoids suppress the incorporation of 3H-uridine into pachytene spermatocytes and round spermatids; the absence of chromosome aberrations in the present study suggests that an adverse effect appears to be present, but may be at a level other than cytogenetic.

Animals↗
PubMed1985

Deletion of the long arm of the Y chromosome and review of Y chromosome abnormalities.

We report on a patient whose karyotype is 45,X/46,X,del(Y) (pter----q11.212). We also present a review of literature on the Y chromosome in which evidence is presented that there are genes on the Y chromosome that prevent Ullrich-Turner syndrome manifestations; aid in testes maturation and spermatogenesis; and affect height, tooth size, and bone maturation.

Adult↗
PubMed1985

Analysis of high-resolution R-bands, obtained by heat-denaturation and Giemsa staining, on human prophase chromosomes.

RHG-bands (heat-denatured Giemsa R-bands) of human prophase chromosomes were analyzed at high resolution, and the banding patterns at prophase and metaphase are presented. The bands were compared with those of the International Standard Cytogenetic Nomenclature idiograms and of the G-band idiograms proposed by J. J. Yunis. The number, size, and position of the RHG-bands correspond rather well with their equivalent G-negative bands, but some differences were noted in the zones of preferential stretching, the juxtacentromeric regions, and the telomeres. Variations in the centromere index and the banding pattern in heterochromatin were also discussed.

Adult↗
PubMed1985

A direct technique for the preparation of chromosomes from early equine embryos.

A technique is described for the preparation of banded chromosomes from early equine embryos cultured for less than 10 h in a medium containing bromodeoxyuridine. In addition to standard Giemsa staining and C-banding, chromosomes thus prepared can also be R-banded by either the RBA or the RB-FPG methods. This technique is rapid, repeatable, and limits cell loss, making it ideal for the preparation of early embryos.

Animals↗