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Contraception and Reproductive Medicine Feb 2024In the last decade, luteal-phase ovarian stimulation (LPOS) has been suggested as an alternative controlled ovarian stimulation (COS) protocol for in vitro...
BACKGROUND
In the last decade, luteal-phase ovarian stimulation (LPOS) has been suggested as an alternative controlled ovarian stimulation (COS) protocol for in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) cycles mainly in women with a history of poor ovarian response (POR). The present randomized controlled trial study aimed to compare the outcomes of follicular phase ovarian stimulation (FPOS) and LPOS protocols in POR cases undergoing ICSI cycles.
METHODS
Seventy-eight POR patients who met the Bologna criteria and underwent an ICSI cycle were included. In this study, 39 POR cases were allocated to the FPOS group, and 39 POR cases were allocated to the LPOS group. The primary outcome was the number of metaphase II (MII) oocytes. In addition, the total number of oocytes, number of top-quality day 3 embryo, day 3 embryo development rate, chemical pregnancy and clinical pregnancy rates were defined as secondary outcomes.
RESULTS
The obtained results demonstrated that the number of MII oocytes significantly increased in the LPOS group compared to the FPOS group (P = 0.007). However, there was no significant difference between the two groups regarding the number of GV and MI oocytes, number of top-quality day 3 embryos and day 3 embryo development rate among both categories of patients. Also, the number of total and MII oocytes was significantly higher in the LPOS group (P = 0.016).
CONCLUSION
These results suggest that LPOS protocol effectively increases the number of mature oocytes in women with a history of POR.
TRIAL REGISTRATION
IRCT20210405050852N1 (Registered at Iranian registry of clinical trials; available at https://en.irct.ir/trial/55402 ).
PubMed: 38368372
DOI: 10.1186/s40834-024-00265-z -
Heat stress at the bicellular stage inhibits sperm cell development and transport into pollen tubes.Plant Physiology Jun 2024For successful double fertilization in flowering plants (angiosperms), pollen tubes deliver 2 nonmotile sperm cells toward female gametes (egg and central cell,...
For successful double fertilization in flowering plants (angiosperms), pollen tubes deliver 2 nonmotile sperm cells toward female gametes (egg and central cell, respectively). Heatwaves, especially during the reproduction period, threaten male gametophyte (pollen) development, resulting in severe yield losses. Using maize (Zea mays) as a crop and grass model system, we found strong seed set reduction when moderate heat stress was applied for 2 d during the uni- and bicellular stages of pollen development. We show that heat stress accelerates pollen development and impairs pollen germination capabilities when applied at the unicellular stage. Heat stress at the bicellular stage impairs sperm cell development and transport into pollen tubes. To understand the course of the latter defects, we used marker lines and analyzed the transcriptomes of isolated sperm cells. Heat stress affected the expression of genes associated with transcription, RNA processing and translation, DNA replication, and the cell cycle. This included the genes encoding centromeric histone 3 (CENH3) and α-tubulin. Most genes that were misregulated encode proteins involved in the transition from metaphase to anaphase during pollen mitosis II. Heat stress also activated spindle assembly check point and meta- to anaphase transition genes in sperm cells. In summary, misregulation of the identified genes during heat stress at the bicellular stage results in sperm cell development and transport defects ultimately leading to sterility.
Topics: Pollen Tube; Heat-Shock Response; Zea mays; Gene Expression Regulation, Plant; Pollen; Germination; Hot Temperature; Plant Proteins
PubMed: 38366643
DOI: 10.1093/plphys/kiae087 -
Current Biology : CB Mar 2024The outer corona plays an essential role at the onset of mitosis by expanding to maximize microtubule attachment to kinetochores. The low-density structure of the corona...
The outer corona plays an essential role at the onset of mitosis by expanding to maximize microtubule attachment to kinetochores. The low-density structure of the corona forms through the expansion of unattached kinetochores. It comprises the RZZ complex, the dynein adaptor Spindly, the plus-end directed microtubule motor centromere protein E (CENP-E), and the Mad1/Mad2 spindle-assembly checkpoint proteins. CENP-E specifically associates with unattached kinetochores to facilitate chromosome congression, interacting with BubR1 at the kinetochore through its C-terminal region (2091-2358). We recently showed that CENP-E recruitment to BubR1 at the kinetochores is both rapid and essential for correct chromosome alignment. However, CENP-E is also recruited to the outer corona by a second, slower pathway that is currently undefined. Here, we show that BubR1-independent localization of CENP-E is mediated by a conserved loop that is essential for outer-corona targeting. We provide a structural model of the entire CENP-E kinetochore-targeting domain combining X-ray crystallography and Alphafold2. We reveal that maximal recruitment of CENP-E to unattached kinetochores critically depends on BubR1 and the outer corona, including dynein. Ectopic expression of the CENP-E C-terminal domain recruits the RZZ complex, Mad1, and Spindly, and prevents kinetochore biorientation in cells. We propose that BubR1-recruited CENP-E, in addition to its essential role in chromosome alignment to the metaphase plate, contributes to the recruitment of outer corona proteins through interactions with the CENP-E corona-targeting domain to facilitate the rapid capture of microtubules for efficient chromosome alignment and mitotic progression.
Topics: Humans; Cell Cycle Proteins; Chromosomal Proteins, Non-Histone; Kinetochores; Microtubules; Mad2 Proteins; Mitosis; Dyneins; Spindle Apparatus; HeLa Cells
PubMed: 38354735
DOI: 10.1016/j.cub.2024.01.042 -
Health Science Reports Feb 2024Endometriosis is a common reason for infertility and poor outcomes of assisted reproductive technology (ART). Inflammation is involved in the pathogenesis of this...
Association between follicular fluid bacteria with inflammatory markers of the complete blood count and the outcomes of assisted reproductive technology in women with endometriosis: A case-control study.
BACKGROUND AND AIMS
Endometriosis is a common reason for infertility and poor outcomes of assisted reproductive technology (ART). Inflammation is involved in the pathogenesis of this disease. The presence of microorganisms in women with endometriosis may increase levels of inflammatory markers. The purpose of this study is to determine the relationship between the presence of bacteria in the follicular fluid with the inflammatory markers of the complete blood count (CBC) and the outcomes of in vitro fertilization (IVF) in women with endometriosis.
METHODS
This case-control study was conducted on 74 patients undergoing IVF, referred to Al-Zahra Hospital in Rasht (Iran) in 2021. The patients were divided into two case groups including 37 women with endometrioma and the control group, including 37 infertile women with a male factor and normal ultrasound. In total, 74 follicular fluids were collected from the case and control groups and were cultured in the laboratory. The relationship between culture results with IVF outcomes and the levels of CBC inflammatory markers including the number of white blood cells (WBCs), lymphocytes, neutrophils, neutrophil-to-lymphocyte ratio (NLR), lymphocyte-to-monocyte ratio (LMR), and platelet-to-lymphocyte ratio (PLR), erythrocyte sedimentation rate (ESR), and c-reactive protein (CRP) was analyzed.
RESULTS
There was no significant statistical difference between the frequency of bacteria present in the follicular fluid ( = 0.861), the mean rate of fertilization ( = 0.363), the frequency of CRP ( = 0.999), and the mean WBCs, lymphocytes, neutrophils, NLR, LMR, and PLR in the two groups. There was a significant statistical difference between the mean number of oocytes of metaphase II ( = 0.034) and the mean ESR ( = 0.018) in the two groups.
CONCLUSIONS
It seems necessary to evaluate follicular fluid as a biological substance that is considered an optimal factor for predicting oocyte quality, fertilization rate, embryo quality, and the success rate of ART.
PubMed: 38343663
DOI: 10.1002/hsr2.1874 -
BMC Cancer Feb 2024Histiocytoses are rare disorders manifested by increased proliferation of pathogenic myeloid cells sharing histological features with macrophages or dendritic cells and...
BACKGROUND
Histiocytoses are rare disorders manifested by increased proliferation of pathogenic myeloid cells sharing histological features with macrophages or dendritic cells and accumulating in various organs, i.a., bone and skin. Pre-clinical in vitro models that could be used to determine molecular pathways of the disease are limited, hence research on histiocytoses is challenging. The current study compares cytophysiological features of progenitor, stromal-like cells derived from histiocytic lesions (sl-pHCs) of three pediatric patients with different histiocytoses types and outcomes. The characterized cells may find potential applications in drug testing.
METHODS
Molecular phenotype of the cells, i.e. expression of CD1a and CD207 (langerin), was determined using flow cytometry. Cytogenetic analysis included GTG-banded metaphases and microarray (aCGH) evaluation. Furthermore, the morphology and ultrastructure of cells were evaluated using a confocal and scanning electron microscope. The microphotographs from the confocal imaging were used to reconstruct the mitochondrial network and its morphology. Basic cytophysiological parameters, such as viability, mitochondrial activity, and proliferation, were analyzed using multiple cellular assays, including Annexin V/7-AAD staining, mitopotential analysis, BrdU test, clonogenicity analysis, and distribution of cells within the cell cycle. Biomarkers potentially associated with histiocytoses progression were determined using RT-qPCR at mRNA, miRNA and lncRNA levels. Intracellular accumulation of histiocytosis-specific proteins was detected with Western blot. Cytotoxicyty and IC50 of vemurafenib and trametinib were determined with MTS assay.
RESULTS
Obtained cellular models, i.e. RAB-1, HAN-1, and CHR-1, are heterogenic in terms of molecular phenotype and morphology. The cells express CD1a/CD207 markers characteristic for dendritic cells, but also show intracellular accumulation of markers characteristic for cells of mesenchymal origin, i.e. vimentin (VIM) and osteopontin (OPN). In subsequent cultures, cells remain viable and metabolically active, and the mitochondrial network is well developed, with some distinctive morphotypes noted in each cell line. Cell-specific transcriptome profile was noted, providing information on potential new biomarkers (non-coding RNAs) with diagnostic and prognostic features. The cells showed different sensitivity to vemurafenib and trametinib.
CONCLUSION
Obtained and characterized cellular models of stromal-like cells derived from histiocytic lesions can be used for studies on histiocytosis biology and drug testing.
Topics: Humans; Child; Histiocytosis, Langerhans-Cell; Vemurafenib; Macrophages; Biomarkers; Phenotype; Antigens, CD; Lectins, C-Type; Mannose-Binding Lectins
PubMed: 38342891
DOI: 10.1186/s12885-023-11807-0 -
Veterinary Research Communications Jun 2024The Mitochondrial distribution pattern or MDP in mammalian oocytes serves as an indicator of their cytoplasmic maturity, with a heterogeneous pattern associated with...
The Mitochondrial distribution pattern or MDP in mammalian oocytes serves as an indicator of their cytoplasmic maturity, with a heterogeneous pattern associated with mature cytoplasm. Currently, MDP assessment involves fluorescent labelling of mitochondria followed by visual evaluation, as no quantitative method exists. Our objective was to develop a quantitative approach to assess MDP in mature equine oocytes. Equine oocytes, obtained by ovum pick up (OPU) were matured in vitro, and only metaphase II oocytes were used in the study (n = 56). Following denudation, oocytes were fixed, stained with MitoTracker™ Red CMXRos (50 nM in TCM-199 with Hank´s salts and 10% FBS) for 15 min at 38 °C, and then incubated with 2.5 µg/ml Hoechst 33342 for 10 min at 38 °C. Confocal microscope images were acquired, and the oocyte's MDP was visually classified as either homogeneous (HoD; n = 17) or heterogeneous (HeD; n = 39). For quantitative analysis, Fiji-ImageJ software was employed. Background subtraction was performed, and a 1-pixel line along the diameter was drawn to calculate the intensity profile. Fluorescence intensities were normalized, and ratios of peripheral to central fluorescence intensity were determined. Student´s t-test was used for comparations; MDP ratio was (mean ± standard error of the mean): 0.8 ± 0.02 for HoD and 0.3 ± 0.02 for HeD (p < 0.001). These results demonstrate concordance between quantitative and qualitative MDP assessment in mature equine oocytes. Our study describes a new approach to quantify mitochondrial distribution pattern and cytoplasmic maturation in mature equine oocytes.
Topics: Animals; Oocytes; Horses; Mitochondria; Female; Microscopy, Confocal
PubMed: 38340267
DOI: 10.1007/s11259-024-10325-z -
International Journal of Molecular... Jan 2024Cryopreservation is an essential step for utilizing various cell types for biological research and medical purposes. At the same time, there is a lack of data on the...
Cryopreservation is an essential step for utilizing various cell types for biological research and medical purposes. At the same time, there is a lack of data on the effect of cryopreservation, especially when prolonged, on the karyotype of cells. In the present work, we analyzed the genetic stability of cells subjected to a cryopreservation procedure. The objects were immortalized Chinese hamster lung fibroblasts (CHL V-79 RJK line) and human endometrial mesenchymal stem/stromal cells (eMSCs). We showed that short-term cryopreservation in liquid nitrogen for up to 6 months did not affect the karyotype stability of CHL V-79 RJK and eMSCs. On the contrary, karyotyping of G-banded metaphase chromosomes in cells underwent 10-year cryopreservation, which revealed genomic instability in both cell lines associated with the variability of chromosome number in cells, random chromosomal rearrangements, and condensation disorder in homologs. In addition, we found out that long-term cryopreservation of eMSCs does not affect the expression of their typical surface markers and morphology, but results in a significant reduction in proliferative potential and early manifestation of cellular senescence features upon eMSCs culturing. Thus, we concluded that the long-term cryopreservation of cells of different types and biological origin can lead to irreversible changes of their karyotype and acceleration of cellular senescence.
Topics: Cricetinae; Animals; Humans; Karyotyping; Cell Line; Karyotype; Cricetulus; Genomic Instability; Cryopreservation
PubMed: 38338745
DOI: 10.3390/ijms25031467 -
Communications Biology Feb 2024Accurate mitosis is coordinated by the spindle assembly checkpoint (SAC) through the mitotic checkpoint complex (MCC), which inhibits the anaphase-promoting complex or...
Accurate mitosis is coordinated by the spindle assembly checkpoint (SAC) through the mitotic checkpoint complex (MCC), which inhibits the anaphase-promoting complex or cyclosome (APC/C). As an essential regulator, Cdc20 promotes mitotic exit through activating APC/C and monitors kinetochore-microtubule attachment through activating SAC. Cdc20 requires multiple interactions with APC/C and MCC subunits to elicit these functions. Functionally assessing these interactions within cells requires efficient depletion of endogenous Cdc20, which is highly difficult to achieve by RNA interference (RNAi). Here we generated Cdc20 RNAi-sensitive cell lines which display a penetrant metaphase arrest by a single RNAi treatment. In this null background, we accurately measured the contribution of each known motif of Cdc20 on APC/C and SAC activation. The CRY box, a previously identified degron, was found critical for SAC by promoting MCC formation and its interaction with APC/C. These data reveal additional regulation within the SAC and establish a novel method to interrogate Cdc20.
Topics: Anaphase-Promoting Complex-Cyclosome; Cdc20 Proteins; Cell Cycle Proteins; M Phase Cell Cycle Checkpoints; Spindle Apparatus; Signal Transduction; Humans
PubMed: 38337031
DOI: 10.1038/s42003-024-05859-6 -
Human Reproduction Open 2024Does palmitic acid (PA), the most common saturated free fatty acid (FFA) in individuals with obesity, contribute to anovulation through upregulation of the...
STUDY QUESTION
Does palmitic acid (PA), the most common saturated free fatty acid (FFA) in individuals with obesity, contribute to anovulation through upregulation of the collagen-crosslinking enzyme lysyl oxidase (LOX) in the ovary?
SUMMARY ANSWER
Increased PA in individuals with obesity can cause LOX upregulation via the activation of hypoxia-inducible factor-1α (HIF-1α), resulting in abnormal collagen deposition in the ovary and anovulation, which can be ameliorated by metformin therapy.
WHAT IS KNOWN ALREADY
The underlying cause of anovulation in individuals with obesity is poorly defined, and accumulating evidence indicates that hormonal disturbance, insulin resistance, and inflammation may all play a role in the development of ovulation disorders in individuals with obesity. However, it remains to be determined whether PA plays a role in the regulation of LOX expression, thus disrupting ovarian extracellular matrix (ECM) remodelling in the ovary and resulting in impaired ovulation in individuals with obesity.
STUDY DESIGN SIZE DURATION
PA concentration and LOX protein abundance and activity in follicular fluid and ovarian tissue were compared between control (n = 21) subjects, patients with obesity with ovulation (n = 22), and patients with obesity with anovulation (n = 16). The effect of PA on LOX protein expression, and the underlying mechanism, was examined in primary human granulosa cells . The improvements in obesity conditions induced by LOX inhibition combined with metformin were investigated in a high-fat diet-induced obese rat model.
PARTICIPANTS/MATERIALS SETTING METHODS
The abundance of PA concentration and LOX activity was measured via a LOX activity assay and ELISA, respectively. The effect of PA on LOX protein expression was examined in the presence or absence of inhibitors of signalling molecules and siRNA-mediated knockdown of the putative transcription factor. Chromatin immunoprecipitation assays were subsequently conducted to further identify the responsible transcription factor. The role of metformin in the treatment of anovulation by LOX inhibition was investigated in a high-fat diet (HFD)-induced obese rat model. The numbers of retrieved total oocytes and metaphase II oocytes were recorded upon ovarian stimulation. Masson's trichrome staining was used to measure the total collagen content, and immunohistochemical staining and western blotting were used to measure LOX, HIF-1α, and collagen I and IV in the ovary.
MAIN RESULTS AND THE ROLE OF CHANCE
Significantly increased FFA, LOX, and collagen abundance were observed in the ovaries of obese women with anovulation, compared to healthy controls or obese women with ovulation. In a HFD-induced obese rat model, metformin corrected the distortion of ovarian morphology by decreasing LOX and collagen protein abundance in the ovary and improving oestrous cyclicity and ovulation. PA increased LOX expression via the activation of HIF-1α in human granulosa cells, which was attenuated by metformin.
LARGE SCALE DATA
N/A.
LIMITATIONS REASONS FOR CAUTION
Several other saturated and polyunsaturated FFAs, such as stearic acid and arachidonic acid, are also increased in the blood of individuals with obesity, and increased levels of other FFAs may also contribute to the development of anovulation in individuals with obesity, which needs to be further verified in the future.
WIDER IMPLICATIONS OF THE FINDINGS
Elevated PA in individuals with obesity can cause LOX dysregulation via activation of HIF-1α, resulting in abnormal collagen deposition in the ovary and anovulation. This dysregulation can be ameliorated by metformin therapy through its local effect on ECM remodelling in the ovary, which is independent of its systemic effect on insulin sensitivity and chronic inflammation.
STUDY FUNDING/COMPETING INTERESTS
This work was supported by the National Natural Science Foundation of China (grant numbers 82101730, 82130046, and 31900598) and Innovative Research Team of High-level local Universities in Shanghai (SHSMU-ZLCX20210201). All the authors declare no conflicts of interest in relation to this work.
PubMed: 38333108
DOI: 10.1093/hropen/hoae002 -
Pakistan Journal of Medical Sciences Jan 2024Chromosome-1 abnormalities (C1As) are common genetic aberrations in hematological malignancies. We sought to evaluate significance of these abnormalities with reference...
BACKGROUND
Chromosome-1 abnormalities (C1As) are common genetic aberrations in hematological malignancies. We sought to evaluate significance of these abnormalities with reference to clinical characteristics and survival outcome in a pediatric B-Lymphoblastic Leukemia (B-ALL) cohort.
METHODS
This is a retrospective study conducted in cytogenetic section of Indus Hospital and Health Network. Data was retrieved from October 2020 to July 2022 for childhood B-ALL cases exhibiting C1As. Chromosome analysis was performed on Cytovision MB8 using G-banded metaphases derived from unstimulated bone marrow culture. Results were recorded according to the International System for Human Cytogenetic Nomenclature (ISCN-2020). Data analyzed using SPSS, version 24.0.
RESULTS
C1As were observed in 60/450 (13.3%) cases of B-ALL. Among C1As, 29 (48%) cases had t(1;19). There were 13 (45%) balanced and 16 (55%) unbalanced translocations. The aberrations without t(1;19) were seen in 31 (52%) cases including 1q duplication with hyperdiploidy in 14 (45%) cases. The median age for C1As with and without t(1;19) was eight years and six years while the median leukocyte count was 32 x 10/L vs. 17 x 10/L. Event-free survival (EFS) for cases with and without t(1;19) was 69% and 74.2% respectively.
CONCLUSION
Despite the fact that the t(1;19) positive group had a higher median age, a higher white cell count and more CNS positives, the difference in EFS is statistically insignificant when compared to the t(1;19) negative cases. Furthermore, we found a survival difference between balanced and unbalanced t(1;19) groups, which is statistically insignificant but warrants large-scale prospective studies for further understanding.
PubMed: 38328656
DOI: 10.12669/pjms.40.2(ICON).8946