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PubMed1991

DNA ploidy pattern of flat adenomas of the large bowel.

In an attempt to clarify the nature of "flat adenomas," DNA content was measured by means of microspectrophotometry. Thirty-nine flat adenomas (FA), 13 with mild, 22 with moderate, and 4 with severe atypia, were collected for this study. In FA, diploidy (D), polyploidy (P), and aneuploidy (A) were found 100, 0, and 0 percent in mild atypia, 41.9, 4.5, and 54.5 percent in moderate atypia, and 0, 0, and 100 percent in severe atypia, respectively. It is assumed that FA have a much higher malignancy potential than previously expected since, histologically, benign-appearing FA with moderate atypia have already contained malignant DNA patterns. In particular, those more than 5 mm in diameter show aneuploidy in 80 percent. These data suggest that FA with moderate atypia play an important role in the pathogenesis of small colonic carcinomas.

Adenoma↗

PubMed1991

Highly repetitive DNA sequence in parthenogenetic Artemia.

The study of the structural organization of the eukaryotic genome is one of the most important tools for disclosing the evolutionary relationships between species. Artemia (Crustacea, Phyllopoda) offers a very interesting model for speciation studies. The genus, distributed all over the world, comprises both bisexual sibling species and parthenogenetic populations, exhibiting different chromosome numbers (diploidy, polyploidy, and heteroploidy). Digestion of genomic DNA of the parthenogenetic Artemia sp. from Tsing-Tao (China) with the restriction enzymes Eco RI and Alu I reveals that a highly repetitive sequence of 133 bp is present. The Eco RI fragment has been cloned and characterized by genomic organization. The distribution of the Eco RI family of repeats was also studied in several bisexual and parthenogenetic Artemia populations and compared with an Alu I repetitive fragment previously identified in Artemia franciscana.

Animals↗

PubMed1991

Parallel origins of duplications and the formation of pseudogenes in mitochondrial DNA from parthenogenetic lizards (Heteronotia binoei; Gekkonidae).

Analysis of mitochondrial DNAs (mtDNAs) from parthenogenetic lizards of the Heteronotia binoei complex with restriction enzymes revealed an approximately 5-kb addition present in all 77 individuals. Cleavage site mapping suggested the presence of a direct tandem duplication spanning the 16S and 12S rRNA genes, the control region and most, if not all, of the gene for the subunit 1 of NADH dehydrogenase (ND1). The location of the duplication was confirmed by Southern hybridization. A restriction enzyme survey provided evidence for modifications to each copy of the duplicated sequence, including four large deletions. Each gene affected by a deletion was complemented by an intact version in the other copy of the sequence, although for one gene the functional copy was heteroplasmic for another deletion. Sequencing of a fragment from one copy of the duplication which encompassed the tRNA(leu)(UUR) and parts of the 16S rRNA and ND1 genes, revealed mutations expected to disrupt function. Thus, evolution subsequent to the duplication event has resulted in mitochondrial pseudogenes. The presence of duplications in all of these parthenogens, but not among representatives of their maternal sexual ancestors, suggests that the duplications arose in the parthenogenetic form. This provides the second instance in H. binoei of mtDNA duplication associated with the transition from sexual to parthenogenetic reproduction. The increased incidence of duplications in parthenogenetic lizards may be caused by errors in mtDNA replication due to either polyploidy or hybridity of their nuclear genomes.

Amino Acid Sequence↗

PubMed1994

Intracytoplasmic single sperm injection: preclinical training and first clinical results.

OBJECTIVE: Our purpose was to reduce oocyte damage before clinical application of intracytoplasmic single sperm injection by training on aged unfertilized oocytes. DESIGN: Intracytoplasmic single sperm injection (ICSI) was accomplished by micromanipulation of sperm and oocytes. PATIENTS: Thirty-four patients consented to donate unfertilized aged oocytes to train for ICSI. Forty-four patients suffering from severe male infertility were treated with ICSI. INTERVENTION: Oocytes were inseminated by intracytoplasmic single sperm injection. MAIN OUTCOME MEASURES: Oocyte damage and fertilization and pregnancy rates were the outcome measures. RESULTS: One hundred fifty-one aged unfertilized oocytes were gathered for training of which 121 were injected with a single sperm and 30 without a spermatozoon as a control group for activation. Oocyte damage, initially as high as 40%, was reduced to 15% after 60 oocytes. Normal fertilization (2PN) occurred in 18%, and polyploidy in 4.4%. The cleavage rate was 69%; none of these embryos were transferred. In the control group, seven oocytes were damaged, seven (30%) showed one pronucleus, and one showed two pronuclei. No cleavage was observed in the control group. In the clinical trial, 44 patients (61 cycles) were clinically treated with the same ICSI procedure, including 575 of 721 collected oocytes. Damage was 13%, activation was 11%, normal fertilization was 30%, and 5 (1%) polypoid zygotes were observed. The fertilization rate ranged from 5 to 100%, with a mean of 39.5 +/- 4% (SE). Nine patients had no fertilization (15%). Ninety-six percent of the zygotes cleaved and 47% were at the four-cell stage 45 hr after injection. One hundred twelve embryos were replaced in 48 transfers (2.3 embryos/ET). One live birth, one miscarriage, and eight ongoing pregnancies were obtained (22%/ET). CONCLUSION: Preclinical practice on aged unfertilized oocytes seems useful before starting clinical ICSI, as the high initial oocyte damage can be reduced and subsequent clinical treatment successfully applied.

Cell Survival↗

PubMed1990

The effects of hyperthermia on the cell cycle of Ehrlich ascites cancer cells in vivo.

The effects of hyperthermia on the cell cycle of Ehrlich ascites cancer cells were studied, and these effects simultaneously evaluated in terms of prolonging the survival of test mice inoculated with tumor cells from heat-treated mice. DDY mice bearing Ehrlich ascites cancer cells were placed in a water bath at 37 degrees C, 39 degrees C, 41 degrees C, 42 degrees C. The heating of mice at 41 degrees C, 42 degrees C and 43 degrees C induced the accumulation of cancer cells at the G2M phase of the cell cycle with many cells exhibiting polyploidy (16 C). The extent of accumulation increased as the temperature of incubation was raised, however the interrupted cell cycle resumed 120 hours after heating. The retransplantation of cells from the heat-treated mice revealed that the mice which were inoculated with Ehrlich ascites cancer cells from mice heated at 43 degrees C survived longer, while the mice which were inoculated with Ehrlich ascites cancer cells from mice heated at 39 degrees C survived for only a slightly shorter time than those which were inoculated with cells from mice heated at 37 degrees C.

Animals↗

PubMed1989

Cytophotometric investigations on a chronic tubular nephropathy simulating preneoplastic condition.

Cytophotometric investigations were performed on the atypical renal tubular epithelium of a young man who died of renal failure, possibly due to chronic lead exposure. The aim of this analysis was to determine whether nuclear atypia indicates incipient malignancy or is a type of so-called "benign atypias". DNA histogram, showing euploid polyploidy supports the latter view and the dysplastic epithelial lesion is not even considered to be a preneoplastic condition.

Adult↗

PubMed1983

Toxicity of a male contraceptive, gossypol, in mammalian cell cultures.

Gossypol, a substance extracted from cotton plants, is a potential male contraceptive. Although human users reported only mild or no side effects, our study found gossypol to be cytotoxic to cultured mammalian cells. A dose dependent decrease in the number of viable cells and mitotic index was demonstrated with both Chinese hamster ovary (CHO) cells and human lymphocytes after the cultures received treatment of gossypol for various durations. Cellular DNA, RNA, and protein synthetic activities were reduced in the presence of gossypol. However, gossypol did not seem to induce chromosome breakage or polyploidy.

Animals↗

PubMed1991

Isolation of two morphologically distinct cell lines from rat arterial smooth muscle expressing high tumorigenic potentials.

Smooth muscle cell proliferation is an important feature of atherogenesis. Some works have hypothesized that a transformation of smooth muscle cells could arise during this pathological process. The present paper describes two spontaneously transformed cell lines of arterial smooth muscle cells (SMC) established from aortic media of adult rat. The cell lines have been designated V6 and V8; some of their morphologic, growth, and metabolic characteristics are described and compared to their parent cells. The two cell lines appeared distinct by their morphology and by their degree of transformation. V6 cells appeared as elongated spindle-shaped cells whereas V8 cells were spread cells with a cobblestone pattern. Karyotypes of both cell lines showed a high polyploidy level. V6 and V8 cell lines were immortalized and showed growth characteristics of transformed cells: low requirement of serum to grow, ability to form colonies in soft agar and tumorigenicity in nude mice; V8 cells presented a higher malignancy than V6 cells. Both V6 and V8 cells exhibited characteristics of cultured arterial SMC: ultrastructure, alpha actin expression at the protein and mRNA level, prostacyclin production. The remarkably different morphologies of the V6 and V8 lines and their transformed phenotype suggest that these cell lines could be useful models to study SMC differentiation and proliferation with respect to atherosclerotic or hypertensive vascular diseases.

Actins↗

PubMed1992

Numerical chromosome anomalies after fertilization of freeze-thawed mouse oocytes.

The chromosome complement of first cleavage stage mouse embryos was analyzed to investigate the effect of slow freezing-fast thawing cryopreservation on chromosome numbers by comparing these numbers with those found fresh after fertilization of control oocytes. Fewer frozen-thawed (34.1%) than control oocytes (75.0%) cleaved to the 2-cell stage after in vitro fertilization. The incidence of hyperploidy was significantly increased by freezing (4.5% vs. 0% in controls). Polyploidy was not significantly affected (17.0% for freeze-thaw embryos vs. 26.2% for controls).

Aneuploidy↗

PubMed1991

A microspectrophotometric study of DNA ploidy patterns of colorectal adenomas.

Eighty-four adenomas (31 mild, 25 moderate, 28 severe atypia), 32 invasive carcinomas and 15 samples of normal mucosa were collected form polypectomy and surgically resected specimens, and their DNA ploidy patterns were examined by microspectrophotometry. The frequency of polyploidy and aneuploidy were 7% and 0% in normal mucosa, 13% and 0% in mild atypia, 20% and 44% in moderate atypia, 32% and 22% in severe atypia and 31% and 53% in invasive carcinomas, respectively. Furthermore, these abnormal ploidy patterns were found frequently in moderate atypia accompanying severe atypia and mild atypia accompanying moderate atypia. These findings suggest that adenomas with moderate atypia might be the early stage of histologically overt cancer. The thin-section method (4 microns or 7 microns) was useful for measuring regional DNA ploidy patterns of colorectal adenoma.

Adenoma↗

PubMed2005

Cloning of resistance gene analogs located on the alien chromosome in an addition line of wheat-Thinopyrum intermedium.

Homology-based gene/gene-analog cloning method has been extensively applied in isolation of RGAs (resistance gene analogs) in various plant species. However, serious interference of sequences on homoeologous chromosomes in polyploidy species usually occurred when cloning RGAs in a specific chromosome. In this research, the techniques of chromosome microdissection combined with homology-based cloning were used to clone RGAs from a specific chromosome of Wheat-Thinopyrum alien addition line TAi-27, which was derived from common wheat and Thinopyrum intermedium with a pair of chromosomes from Th. intermedium. The alien chromosomes carry genes for resistance to BYDV. The alien chromosome in TAi-27 was isolated by a glass needle and digested with proteinase K. The DNA of the alien chromosome was amplified by two rounds of Sau3A linker adaptor-mediated PCR. RGAs were amplified by PCR with the degenerated primers designed based on conserved domains of published resistance genes (R genes) by using the alien chromosome DNA, genomic DNA and cDNA of Th. intermedium, TAi-27 and 3B-2 (a parent of TAi-27) as templates. A total of seven RGAs were obtained and sequenced. Of which, a constitutively expressed single-copy NBS-LRR type RGA ACR 3 was amplified from the dissected alien chromosome of TAi-27, TcDR 2 and TcDR 3 were from cDNA of Th. intermedium, AcDR 3 was from cDNA of TAi-27, FcDR 2 was from cDNA of 3B-2, AR 2 was from genomic DNA of TAi-27 and TR 2 was from genomic DNA of Th. intermedium. Sequence homology analyses showed that the above RGAs were highly homologous with known resistance genes or resistance gene analogs and belonged to NBS-LRR type of R genes. ACR 3 was recovered by PCR from genomic DNA and cDNA of Th. intermedium and TAi-27, but not from 3B-2. Southern hybridization using the digested genomic DNA of Th. intermedium, TAi-27 and 3B-2 as the template and ACR 3 as the probe showed that there is only one copy of ACR 3 in the genome of Th. intermedium and TAi-27, but it is absent in 3B-2. The ACR 3 could be used as a specific probe of the R gene on the alien chromosome of TAi-27. Results of Northern hybridization suggested that ACR 3 was constitutively expressed in Th. intermedium and TAi-27, but not 3B-2, and expressed higher in leaves than in roots. This research demonstrated a new way to clone RGAs located on a specific chromosome. The information reported here should be useful to understand the resistance mechanism of, and to clone resistant genes from, the alien chromosome in TAi-27.

Amino Acid Sequence↗

PubMed2025

Genetic basis and role of exotic accessions in cultivated cotton fiber quality improvement.

Exotic Gossypium accessions still harbor QTL‑validated alleles that, combined with CRISPR pyramiding and genomic selection, can break the entrenched fiber length-strength trade‑off. Cotton's four independent domestications twice in diploids and twice in allotetraploids offer a natural experiment in fiber improvement. Synthesizing three decades of data, we chart how polyploidy, selection and modern breeding have repeatedly reshaped the Gossypium genome. More than 15,000 quantitative trait locus (QTL) and genome wide association mapping studies (GWAS) hits converge on a handful of chromosomal "hotspots"; new MAGIC, NAM, NIL and long-read resources now narrow these peaks to < 200 kb, resolving causal genes such as GhHOX3, GhZF14 and GhMYB7. Multi-omics evidence links auxin, ethylene, gibberellin, brassinosteroid and strigolactone signaling to HDZIP IV, MYB, bHLH/HLH and ERF networks that drive fiber initiation, extreme cell elongation and cellulose deposition. Population genomics shows that ~ 40% of favorable fiber alleles are fixed in elite Gossypium hirsutum, yet wild diploids and landraces still harbor variants that could break the length strength trade-off. We propose a three-step roadmap genomic selection, CRISPR gene pyramiding and accelerated introgression to expand cotton's genetic base and deliver fibers suited to sustainable textile demands.

Gossypium↗

PubMed2004

Analysis of genomic and expressed major histocompatibility class Ia and class II genes in a hexaploid Lake Tana African 'large' barb individual (Barbus intermedius).

Expression of too many co-dominant major histocompatibility complex (MHC) alleles is thought to be detrimental to proper functioning of the immune system. Polyploidy of the genome will increase the number of expressed MHC genes unless they are prone to a silencing mechanism. In polyploid Xenopus species, the number of MHC class I and II genes has been physically reduced, as it does not increase with higher ploidy genomes. In the zebrafish some class II B loci have been silenced, as only two genomically bona fide loci, DAA/DAB and DEA/DEB, have been described. Earlier studies indicated a reduction in the number of genomic and expressed class II MHC genes in a hexaploid African 'large' barb. This prompted us to study the number of MHC genes present in the genome of an African 'large' barb individual (Barbus intermedius) in relation to those expressed, adopting the following strategy. Full-length cDNA sequences were generated from mRNA and compared with partial genomic class Ia and II sequences generated by PCR using the same primer set. In addition, we performed Southern hybridizations to obtain a verification of the number of class I and II B genes. Our study revealed three beta2-microglobulin, five class Ia, four class II A, and four class II B genes at the genomic level, which were shown to be expressed in the hexaploid barb individual. The class Ia and class II data indicate that the ploidy status does not correlate with the presence and expression of these MHC genes.

Amino Acid Sequence↗

PubMed2001

Requirement of thrombopoietin-induced activation of ERK for megakaryocyte differentiation and of p38 for erythroid differentiation.

Thrombopoietin (TPO) plays a critical role not only in proliferation and differentiation of megakaryocytes but also in erythroid differentiation. We have investigated whether the different pathway of mitogen-activated protein kinase (MAPK) after TPO stimulation may discriminate megakaryocyte and erythroid differentiation. In this study, we have used human CD34+ hematopoietic progenitor cells (HPCs) from cord blood (CB) in serum-free liquid culture supplemented with TPO, to compare the respective effects of specific inhibitors of MAPK kinase (MEK) (PD98059) and p38 MAP kinase (p38) (SB203580) on megakaryocyte and erythroid development. PD98059, but not SB203580, significantly suppressed TPO-induced megakaryocyte differentiation when examined by the expression of CD41 and polyploidy assay. In the presence of SB203580, CD34+/CD36+ erythroid progenitors clearly decreased, whereas they increased when cultured with PD98059. These results indicate that activation of extracellular-signal-regulated kinase (ERK) is required for TPO-induced megakaryocyte differentiation and that p38 is required for TPO-induced erythroid differentiation.

Antigens, CD34↗

PubMed2004

Quantitative determination of mosaic GFP gene expression in tobacco.

A specific form of gene silencing that was observed visually as a mosaic distribution of fluorescent and non-fluorescent cells apparently dispersed at random within tissues was found in a few green fluorescent protein (GFP)-transformed tobacco lines. To characterize this event quantitatively, we studied flow cytometric measurements in GFP-expressing and -silenced cells in T1 and T2 progeny of four selected plants. The proportion of silenced cells varied considerably among the T1 lines but with notable genotype differences. Mosaic expression was inherited into the T2 generation in which the majority of progenies tested exhibited a level of silencing similar to that of their T1 parental plants. However, in some T2 progenies segregation, evident as a decrease or increase in the proportion of fluorescent cells, was observed. We discuss several factors, such as copy number, promoter activity or polyploidy, that may be the possible causes of the gene silencing, but none sufficiently explain the appearance of the mosaic distribution.

Base Sequence↗

PubMed2003

Investigation of c-myc and p53 gene alterations in the tumor and surgical borderline tissues of NSCLC and effects on clinicopathologic behavior: by the FISH technique.

Genetic alterations on the primary tumoral tissues and surgical borderline tissues of 51 patients with NSCLC on which radiotherapy and chemotherapy had not been performed were analyzed by using the FISH method with locus-specific probes for p53 tumor suppressor gene and c-myc oncogene and centromere-specific probes for chromosome 17 and chromosome 8 on which these genes are located. P53 deletions were detected in 7 patients (13.7%), c-myc amplification in 4 patients (7.8%), monosomy 17 in 2 patients (3.9%) and trisomy 8 in 3 patients (5.8%), and a high level of polyploidy in tumoral tissues of 6 patients (11.7%). P53 deletion and c-myc amplification were found at surgical borderlines of 2 patients and 1 patient, respectively. Although both p53 deletion and c-myc amplification have low frequency at surgical border tissues, not only their detection is important for the follow-up of recurrency and metastasis, but it is also important for genetical and pathological staging. The results of this study show that c-myc amplification in NSCLC is related to the shortening of survival (p < 0.01). C-myc amplification and p53 deletion are also effective for the occurrence of metastasis (p < 0.05) and the effect of c-myc amplification in this matter is much higher than p53 deletion. The gain or loss of copy number of chromosome 8 and monosomy 17 show parallel effects with c-myc amplification and p53 deletion, respectively, on the clinicopathological behavior of tumors.

Adult↗

PubMed2004

Spontaneous chromosome doubling results from nuclear fusion during in vitro maize induced microspore embryogenesis.

A multidisciplinary study was carried out to analyse the chromosome doubling process during the early stages of in vitro maize microspore embryogenesis. The main stages (microspore derivatives) that were formed in the course of the culture were analysed. Chromosome number was determined from squashed cells, and DNA content was measured by cytometry. In parallel, an ultrastructural analysis of the microspore derivatives demonstrated the occurrence of a nuclear fusion process. It seems likely that nuclear fusion ensures chromosome doubling at early stages of induced microspore embryogenesis. It occurs precisely at the 5/7 day stage in the embryonic domain and probably leads to polyploidy in the endosperm domain of the microspore derivatives. As a conclusion a scheme summarises the results and proposes an interpretation of the sequence of chromosome doubling events during early maize microspore embryogenesis. Understanding of this process will be important for future efforts to increase the percentage of homozygous plants for crop improvement.

Cell Fusion↗

PubMed1998

Estimation of the mutagenic potential of the trematode Opisthorchis felineus in experimentally infected guinea pigs.

We studied the mutagenic potential of the trematode Opisthorchis felineus in experimentally infected guinea pigs by chromosome analysis. The investigation was conducted on 30 treated (experimentally infected) and 20 normal (uninfected) guinea pigs (Cavia porcellus). A sample of living metacercariae of O. felineus was obtained from the muscle of naturally infected crucian carp captured in the Tom River, and 1 ml suspension containing approximately 50 metacercariae in 0.9% NaCl solution was individually given p.o. to guinea pigs that were used as treated specimens, whereas normal guinea pigs from the same stock were given 1 ml 0.9% NaCl solution per individual to serve as controls. The treated and control animals were killed by CO2 asphyxiation and decapitation at 7, 15, 30, 45, and 120 days after contamination. The smears of bone marrow cells for chromosome analysis were prepared by a standard method. Statistical analysis of the data was done using Student's t-test. We found that experimental infection of guinea pigs with O. felineus induced significantly high frequencies of hypoploid cells and, at the 120th day p.i., led to a considerable incidence of chromatid breaks and polyploidy in bone marrow cells.

Animals↗