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Mutation Research. Genetic Toxicology... 2024Cytogenetic studies have shown that human chromosomes 1, 9, and 16, with a large heterochromatic region of highly methylated classical satellite DNA, are prone to...
Cytogenetic studies have shown that human chromosomes 1, 9, and 16, with a large heterochromatic region of highly methylated classical satellite DNA, are prone to induction of chromatid breaks and interchanges by mitomycin C (MMC). A couple of studies have indicated that material from chromosome 9, and possibly also from chromosomes 1 and 16, are preferentially micronucleated by MMC. Here, we further examined the chromosome-specific induction of micronuclei (MN; with and without cytochalasin B) and chromosomal aberrations (CAs) by MMC. Cultures of isolated human lymphocytes from two male donors were treated (at 48 h of culture, for 24 h) with MMC (500 ng/ml), and the induced MN were examined by a pancentromeric DNA probe and paint probe for chromosome 9, and by paint probes for chromosomes 1 and 16. MMC increased the total frequency of MN by 6-8-fold but the frequency of chromosome 9 -positive (9) MN by 29-30-fold and the frequency of chromosome 1 -positive (1) MN and chromosome 16 -positive (16) MN by 12-16-fold and 10-17-fold, respectively. After treatment with MMC, 34-47 % of all MN were 9, 17-20 % 1, and 3-4 % 16. The majority (94-96 %) of the 9 MN contained no centromere and thus harboured acentric fragments. When MMC-induced CAs aberrations were characterized by using the pancentromeric DNA probe and probes for the classical satellite region and long- and short- arm telomeres of chromosome 9, a high proportion of chromosomal breaks (31 %) and interchanges (41 %) concerned chromosome 9. In 83 % of cases, the breakpoint in chromosome 9 was just below the region (9cen-q12) labelled by the classical satellite probe. Our results indicate that MMC specifically induces MN harbouring fragments of chromosome 9, 1, and 16. CAs of chromosome 9 are highly overrepresented in metaphases of MMC-treated lymphocytes. The preferential breakpoint is below the region 9q12.
Topics: Humans; Mitomycin; Male; Chromosome Aberrations; Micronuclei, Chromosome-Defective; Chromosomes, Human, Pair 9; Chromosomes, Human, Pair 1; Chromosomes, Human, Pair 16; Lymphocytes; Adult; Micronucleus Tests; Cells, Cultured; Cytochalasin B; In Situ Hybridization, Fluorescence
PubMed: 38821666
DOI: 10.1016/j.mrgentox.2024.503753 -
Cureus Apr 2024This case report demonstrates the management of primary infertility in a couple: the male was 37 years old and the female was 32 years old. The female had a submucosal...
This case report demonstrates the management of primary infertility in a couple: the male was 37 years old and the female was 32 years old. The female had a submucosal uterine fibroid. Later, the female underwent a myomectomy to remove submucosal fibroids in the uterus after two failed intrauterine insemination (IUI) cycles. After six months of her recovery period, she underwent ovum pickup for an in vitro fertilization (IVF) cycle. During the process of ovum pickup (OPU), four oocytes were retrieved: three in the metaphase one (M1) stage and one in the metaphase two (M2) stage. Subsequently, the couple underwent in vitro maturation (IVM) of oocytes, where the M1 stage oocytes were cultured for six hours. The M1 stage oocytes progressed to the M2 stage. These oocytes were then injected with sperm, which resulted in the formation of two blastocysts. These blastocysts were then cryopreserved for three months, and after three months, these frozen embryos were then transferred, leading to the successful conception. The case study evaluates a couple who suffered from infertility. This study includes a treatment of myomectomy and in vitro maturation.
PubMed: 38813276
DOI: 10.7759/cureus.59257 -
Journal of Biomedical Optics Jun 2024Preparation of a recipient cytoplast by oocyte enucleation is an essential task for animal cloning and assisted reproductive technologies in humans. The femtosecond...
SIGNIFICANCE
Preparation of a recipient cytoplast by oocyte enucleation is an essential task for animal cloning and assisted reproductive technologies in humans. The femtosecond laser is a precise and low-invasive tool for oocyte enucleation, and it should be an appropriate alternative to traditional enucleation by a microneedle aspiration. However, until recently, the laser enucleation was performed only with applying a fluorescent dye.
AIM
This work is aimed to (1) achieve femtosecond laser oocyte enucleation without applying a fluorescent dye and (2) to study the effect of laser destruction of chromosomes on the structure and dynamics of the spindle.
APPROACH
We applied polarized light microscopy for spindle visualization and performed stain-free mouse and human oocyte enucleation with a 1033 nm femtosecond laser. Also, we studied transformation of a spindle after metaphase plate elimination by a confocal microscopy.
RESULTS
We demonstrated a fundamental possibility of inactivating the metaphase plate in mouse and human oocytes by 1033 nm femtosecond laser radiation without applying a fluorescent dye. Irradiation of the spindle area, visualized by polarized light microscopy, resulted in partly or complete metaphase plate destruction but avoided the microtubules impairment. After the metaphase plate elimination, the spindle reorganized, however, it was not a complete depolymerization.
CONCLUSIONS
This method of recipient cytoplast preparation is expected to be useful for animal cloning and assisted reproductive technologies.
Topics: Animals; Mice; Oocytes; Humans; Female; Lasers; Spindle Apparatus; Microscopy, Confocal; Metaphase; Microscopy, Polarization
PubMed: 38812963
DOI: 10.1117/1.JBO.29.6.065002 -
Cell Biochemistry and Function Jun 2024Cancer is the second leading cause of mortality worldwide. The development of anticancer therapy plays a crucial role in mitigating tumour progression and metastasis....
Cancer is the second leading cause of mortality worldwide. The development of anticancer therapy plays a crucial role in mitigating tumour progression and metastasis. Epithelioid hemangioendothelioma is a very rare cancer, however, with a high systemic involvement. Kynurenine metabolites which include l-kynurenine, 3-hydroxykynurenine, 3-hydroxyanthranilic acid and quinolinic acid have been shown to inhibit T-cell proliferation resulting in a decrease in cell growth of natural killer cells and T cells. Furthermore, metabolites such as l-kynurenine have been shown to inhibit proliferation of melanoma cells in vitro. Considering these metabolite properties, the present study aimed to explore the in vitro effects of l-kynurenine, quinolinic acid and kynurenic acid on endothelioma sEnd-2 cells and on endothelial (EA. hy926 cells) (control cell line). The in vitro effect at 24, 48, and 72 h exposure to a range of 1-4 mM of the respective kynurenine metabolites on the two cell lines in terms of cell morphology, cell cycle progression and induction of apoptosis was assessed. The half inhibitory concentration (IC), as determined using nonlinear regression, for l-kynurenine, quinolinic acid and kynurenic acid was 9.17, 15.56, and 535.40 mM, respectively. Optical transmitted light differential interference contrast and hematoxylin and eosin staining revealed cells blocked in metaphase, formation of apoptotic bodies and compromised cell density in l-kynurenine-treated cells. A statistically significant increase in the number of cells present in the sub-G phase was observed in l-kynurenine-treated sample. To our knowledge, this was the first in vitro study conducted to investigate the mechanism of action of kynurenine metabolites on endothelioma sEnd-2 cells. It can be concluded that l-kynurenine exerts an antiproliferative effect on the endothelioma sEnd-2 cell line by decreasing cell growth and proliferation as well as a metaphase block. These hallmarks suggest cell death via apoptosis. Further research will be conducted on l-kynurenine to assess the effect on cell adhesion in vitro and in vivo as cell-cell adhesion has been shown to increase metastasis to distant organs therefore, the inhibition of adhesion may lead to a decrease in metastasis.
Topics: Kynurenine; Humans; Apoptosis; Cell Proliferation; Quinolinic Acid; Kynurenic Acid; Cell Cycle; Cell Line, Tumor; Antineoplastic Agents; Endothelial Cells; Dose-Response Relationship, Drug
PubMed: 38807444
DOI: 10.1002/cbf.4065 -
Open Biology May 2024The precise spatial and temporal control of histone phosphorylations is important for the ordered progression through the different phases of mitosis. The...
The precise spatial and temporal control of histone phosphorylations is important for the ordered progression through the different phases of mitosis. The phosphorylation of H2B at S6 (H2B S6ph), which is crucial for chromosome segregation, reaches its maximum level during metaphase and is limited to the inner centromere. We discovered that the temporal and spatial regulation of this modification, as well as its intensity, are governed by the scaffold protein RepoMan and its associated catalytically active phosphatases, PP1α and PP1γ. Phosphatase activity is inhibited at the area of maximal H2B S6 phosphorylation at the inner centromere by site-specific Aurora B-mediated inactivation of the PP1/RepoMan complex. The motor protein Mklp2 contributes to the relocalization of Aurora B from chromatin to the mitotic spindle during anaphase, thus alleviating Aurora B-dependent repression of the PP1/RepoMan complex and enabling dephosphorylation of H2B S6. Accordingly, dysregulation of Mklp2 levels, as commonly observed in tumour cells, leads to the lack of H2B S6 dephosphorylation during early anaphase, which might contribute to chromosomal instability.
Topics: Aurora Kinase B; Phosphorylation; Humans; Histones; Mitosis; Protein Phosphatase 1; Cell Cycle Proteins; HeLa Cells; Spindle Apparatus; Centromere; Nuclear Proteins
PubMed: 38806145
DOI: 10.1098/rsob.230460 -
Journal of Ovarian Research May 2024The key to enhancing the efficacy of antagonistic regimens in pregnancy is to better synchronize follicular growth during cycles of controlled ovarian stimulation (COS),...
BACKGROUND
The key to enhancing the efficacy of antagonistic regimens in pregnancy is to better synchronize follicular growth during cycles of controlled ovarian stimulation (COS), especially in patients with diminished ovarian reserve (DOR). During in vitro fertilization-embryo transfer (IVF-ET) treatment, luteal phase estrogen pretreatment may enhance follicular development synchronization and yield of mature oocytes. However, the effect of estrogen pretreatment in DOR patients with elevated basal follicle-stimulating hormone (FSH) levels has not been well studied.
METHODS
We retrospectively analyzed the clinical data of patients with elevated basal FSH levels and DOR (401 cycles) who underwent IVF/intracytoplasmic monosperm injection (ICSI)-assisted conception. Both groups were treated with a flexible gonadotropin-releasing hormone (GnRH) antagonist regimen and were further divided into two groups according to whether they received luteal estrogen pretreatment. There were 79 patients in the estrogen pretreatment group and 322 patients in the control group. On the second day of the menstrual cycle, gonadotropin (Gn) stimulation of the ovaries was initiated. The general characteristics, clinical, biological parameters and outcomes of the two groups were compared.
RESULTS
The basic profiles of the two groups were similar (P > 0.05). More patients in the pretreatment group showed FSH rebound after gonadotropin (Gn) initiation, resulting in a significantly higher number of Gn days and total Gn than those in the control group (P < 0.05). There was no statistically significant difference in the number of days of antagonist use, follicle output rate (FORT), number of metaphase II(MII)eggs obtained, number of Two pronuclei (2PN) fertilized, number of D quality embryos, blastocyst formation rate, fresh embryo clinical pregnancy rate, cumulative pregnancy rate, and non-transferable embryo rate between the two groups (P > 0.05).
CONCLUSIONS
The use of luteal phase estrogen pretreatment in patients with elevated basal FSH combined with DOR resulted in high FSH levels after the release of negative feedback, which was detrimental to early follicular growth, did not increase the follicular output rate, may have increased the use and duration of controlled ovarian stimulation drugs, and did not increase the number of eggs gained or improve clinical outcomes.
Topics: Humans; Female; Retrospective Studies; Adult; Follicle Stimulating Hormone; Ovulation Induction; Ovarian Reserve; Estrogens; Fertilization in Vitro; Pregnancy; Gonadotropin-Releasing Hormone; Pregnancy Rate; Embryo Transfer
PubMed: 38802887
DOI: 10.1186/s13048-024-01415-2 -
JBRA Assisted Reproduction May 2024Following the advancement of medically assisted reproduction (MAR) technology, and the rationale to extend the culture to the blastocyst stage, performing elective... (Review)
Review
Following the advancement of medically assisted reproduction (MAR) technology, and the rationale to extend the culture to the blastocyst stage, performing elective single embryo transfer (eSET), gamete quality and assessment have acquired large relevance in ART. Embryo quality is strictly correlated with gametes quality and culture conditions. Oocyte maturity assessment is therefore imperative for fertilization and embryo evolution. Mature oocytes at the metaphase II stage result in a higher fertilization rate compared to immature oocytes. Indeed, oocyte morphology evaluation represents an important and challenging task that may serve as a valuable prognostic tool for future embryo development and implantation potential. Different grading systems have been reported to assess human embryos, however, in many cases, it is still a major challenge to select the single embryo to transfer with the highest implantation potential. Further, eSET has conferred a challenge to embryologists, who must try to enhance embryo culture and selection to provide an adequate success rate, whilst reducing the overall number of embryos transferred. Above the standard morphological assessment, there are several invasive or non-invasive approaches for embryo selection such as preimplantation genetic testing, time-lapse technology, proteomics and metabolomics, as well as oxygen utilization and analysis of oxidative stress in culture medium. This short review is not designed to be a comprehensive review of all possible features that may influence oocyte quality. It does give, however, a brief overview and describes the prognostic value of the morphological characteristics of human oocytes on their developmental capacity following ART treatments.
PubMed: 38801314
DOI: 10.5935/1518-0557.20240034 -
JBRA Assisted Reproduction May 2024One of the techniques that has gained much attention is the in vitro maturation of oocytes for patients who use assisted reproduction techniques. However, its results...
OBJECTIVE
One of the techniques that has gained much attention is the in vitro maturation of oocytes for patients who use assisted reproduction techniques. However, its results are still inferior to controlled ovarian stimulation methodologies. Understanding the maturation mechanisms based on analyses can help improve this methodology's results. The work aims to identify the central genes differentially expressed in oocytes after in vitro maturation in the germinal vesicle and metaphase II stages.
METHODS
This work is a computational analysis. The entire search will be conducted using the Gene Expression Omnibus (GEO) database. To carry out and obtain the data present in the work, an advanced research search was carried out in the GEO database within the period from January 1, 2013, to January 1, 2023. A total of 27 genomic data were available in the GEO database, of which only two were used.
RESULTS
Two datasets were identified on the Gene Expression Omnibus database platform: registration data GSE158802 and GSE95477. From the analysis, we identified five downregulated and thirty-six upregulated genes; the central genes that correlated with the main gene proteins found were CLTA and PANK1.
CONCLUSIONS
There was a differential regulation of gene expression. The most central ones are related to energy capture.
PubMed: 38801311
DOI: 10.5935/1518-0557.20240030 -
BioRxiv : the Preprint Server For... May 2024Chromosome congression and alignment on the metaphase plate involves lateral and microtubule plus-end interactions with the kinetochore. Here we take advantage of our...
UNLABELLED
Chromosome congression and alignment on the metaphase plate involves lateral and microtubule plus-end interactions with the kinetochore. Here we take advantage of our ability to efficiently generate a GFP-marked acentric X chromosome fragment in neuroblasts to identify forces acting on chromosome arms that drive congression and alignment. We find acentrics efficiently align on the metaphase plate, often more rapidly than kinetochore-bearing chromosomes. Unlike intact chromosomes, the paired sister acentrics oscillate as they move to and reside on the metaphase plate in a plane distinct and significantly further from the main mass of intact chromosomes. Consequently, at anaphase onset acentrics are oriented either parallel or perpendicular to the spindle. Parallel-oriented sisters separate by sliding while those oriented perpendicularly separate via unzipping. This oscillation, together with the fact that in monopolar spindles acentrics are rapidly shunted away from the poles, indicates that distributed plus-end directed forces are primarily responsible for acentric migration. This conclusion is supported by the observation that reduction of EB1 preferentially disrupts acentric alignment. In addition, reduction of Klp3a activity, a gene required for the establishment of pole-to-pole microtubules, preferentially disrupts acentric alignment. Taken together these studies suggest that plus-end forces mediated by the outer pole-to-pole microtubules are primarily responsible for acentric metaphase alignment. Surprisingly, we find that a small fraction of sister acentrics are anti-parallel aligned indicating that the kinetochore is required to ensure parallel alignment of sister chromatids. Finally, we find induction of acentric chromosome fragments results in a global reorganization of the congressed chromosomes into a torus configuration.
ARTICLE SUMMARY
The kinetochore serves as a site for attaching microtubules and allows for successful alignment, separation, and segregation of replicated sister chromosomes during cell division. However, previous studies have revealed that sister chromosomes without kinetochores (acentrics) often align to the metaphase plate, undergo separation and segregation, and are properly transmitted to daughter cells. In this study, we discuss the forces acting on chromosomes, independent of the kinetochore, underlying their successful alignment in early mitosis.
PubMed: 38798431
DOI: 10.1101/2023.11.14.567057 -
Mathematical Biosciences May 2024This paper develops a theory for anaphase in cells. After a brief description of microtubules, the mitotic spindle and the centrosome, a mathematical model for anaphase...
This paper develops a theory for anaphase in cells. After a brief description of microtubules, the mitotic spindle and the centrosome, a mathematical model for anaphase is introduced and developed in the context of the cell cytoplasm and liquid crystalline structures. Prophase, prometaphase and metaphase are then briefly described in order to focus on anaphase, which is the main study of this paper. The entities involved are modelled in terms of liquid crystal defects and microtubules are represented as defect flux lines. The mathematical techniques employed make extensive use of energy considerations based on the work that was developed by Dafermos (1970) from the classical Frank-Oseen nematic liquid crystal energy (Frank, 1958; Oseen, 1933). With regard to liquid crystal theory we introduce the concept of regions of influence for defects which it is believed have important implications beyond the subject of this paper. The results of this paper align with observed biochemical phenomena and are explored in application to HeLa cells and Caenorhabditis elegans. This unified approach offers the possibility of gaining insight into various consequences of mitotic abnormalities which may result in Down syndrome, Hodgkin lymphoma, breast, prostate and various other types of cancer.
PubMed: 38795952
DOI: 10.1016/j.mbs.2024.109219