Spindle Positioning Research Articles

Subcortical Maternal Complex in Female Infertility: A Transition from Animal Models to Human Studies.

Female infertility is a significant healthcare burden that is frequently encountered among couples globally. While environmental factors, comorbidities, and lifestyle determine reproductive health, certain genetic variants in key reproductive genes can potentially cause unsuccessful pregnancies. Such crucial proteins have been identified within the subcortical maternal complex (SCMC) and play an integral role in the early stages of embryogenesis before embryo implantation. SCMC proteins are associated with crucial pathways during embryogenesis, causing changes that are necessary for the transition of an oocyte to an embryo. These vital processes include the formation of cytoplasmic spindles and lattices, accurate positioning of meiotic spindles, regulatory roles in various gene translations, organelle redistribution, and zygotic genome reprogramming. While these genes are well studied in animal models, often mice, translation to clinical studies is comparatively less. The present study elucidates the transition in genetic studies from animal to human models of SCMC proteins. The present literature review shows that the expression of various SCMC proteins impairs embryo development at different stages. The clinical translation of SCMC occurs via various pathways. Therefore, females experiencing multiple unsuccessful pregnancies after natural or assisted conception techniques are candidates for underlying SCMC mutations. Although the phenotype of affected individuals has been identified, the molecular mechanisms that lead to impaired pathways still require investigation. Therefore, the present study paves the way for future research leading to the early diagnosis of lethal variants and possible subsequent management.

Dynamics of spindle assembly and position checkpoints: Integrating molecular mechanisms with computational models

Dynamics of spindle assembly and position checkpoints: Integrating molecular mechanisms with computational models

Manipulation of Spindle Position Using Magnetic Tweezers in Sea Urchin Embryos.

The regulation of mitotic spindle position and orientation, and consequent division plane specification, is critical for early embryo development, tissue architecture and stem cells. Accordingly, defects in spindle positioning have been associated with the emergence of tissue disorders and mis-specification of cell fates, causing the formation of tumors, for instance, in stem cell populations. In such context, methods to physically manipulate mitotic spindle position or orientation in living cells bear the promise of dissecting the causal impact of spindle mis-positioning on cell or tissue behavior. Here, we describe a method to directly modulate mitotic spindle position and orientation with in vivo magnetic tweezers in developing sea urchin embryos. This method allows formonitoring the impact of spindle orientation on embryo development and quantifying forces and torques needed to move or rotate spindles. It may be transposable to many other cell, embryo or tissue types.

Numerical investigation of spindle position effects on steam ejector performance with a nonequilibrium condensation model

AbstractThis research studied the performance of a spindle‐controlled steam ejector using models such as ideal gas and wet steam under various operating conditions based on computational fluid dynamics (CFD). A wet steam model incorporating nonequilibrium condensation was employed to simulate the complex flow phenomena within the ejector. The structure of the flow, entrainment ratio (Er), and shock wave characteristics of the steam ejector were examined in two different models. Results indicate that the spindle position has a substantial impact on steam ejector performance. For the ideal gas and wet steam models, the optimal spindle position (SP‐5) at a .1 MPa motive pressure achieves the highest entrainment ratios (Er) of 1.01 and 1.042, respectively. However, an ejector with a fixed geometry achieves Er values of only .517 and .549 for the ideal gas and wet steam models, under identical working conditions. This represents a substantial improvement of 89.8% over the fixed‐geometry ejector. The wet steam model consistently predicts 2%–4% higher Er values compared with the ideal gas model across all spindle positions. The study also reveals that increasing the motive pressure from .1 to .3 MPa reduces Er by up to 45.8% at the optimal spindle position, with the shock train length extending to 35% of the mixing chamber at .3 MPa. These findings offer insights for improving the design and optimization of variable‐geometry steam ejectors, potentially increasing efficiency in industrial applications.

Spatiotemporal regulation of organelle transport by spindle position checkpoint kinase Kin4.

Asymmetric cell division in Saccharomyces cerevisiae involves class V myosin-dependent transport of organelles along the polarised actin cytoskeleton to the emerging bud. Vac17 is the vacuole/lysosome-specific myosin receptor. Its timely breakdown terminates transport and results in the proper positioning of vacuoles in the bud. Vac17 breakdown is controlled by the bud-concentrated p21-activated kinase Cla4, and the E3-ubiquitin ligase Dma1. We found that the spindle position checkpoint kinase Kin4 and, to a lesser extent, its paralog Frk1 contribute to successful vacuole transport by preventing the premature breakdown of Vac17 by Cla4 and Dma1. Furthermore, Kin4 and Cla4 contribute to the regulation of peroxisome transport. We conclude that Kin4 antagonises the Cla4/Dma1 pathway to coordinate spatiotemporal regulation of organelle transport.

A noncanonical GTPase signaling mechanism controls exit from mitosis in budding yeast

In the budding yeast Saccharomyces cerevisiae, exit from mitosis is coupled to spindle position to ensure successful genome partitioning between mother and daughter cells. This coupling occurs through a GTPase signaling cascade known as the mitotic exit network (MEN). The MEN senses spindle position via a Ras-like GTPase Tem1 which localizes to the spindle pole bodies (SPBs, yeast equivalent of centrosomes) during anaphase and signals to its effector protein kinase Cdc15. How Tem1 couples the status of spindle position to MEN activation is not fully understood. Here, we show that Cdc15 has a relatively weak preference for Tem1GTP and Tem1's nucleotide state does not change upon MEN activation. Instead, we find that Tem1's nucleotide cycle establishes a localization-based concentration difference in the cell where only Tem1GTP is recruited to the SPB, and spindle position regulates the MEN by controlling Tem1 localization to the SPB. SPB localization of Tem1 primarily functions to promote Tem1-Cdc15 interaction for MEN activation by increasing the effective concentration of Tem1. Consistent with this model, we demonstrate that artificially tethering Tem1 to the SPB or concentrating Tem1 in the cytoplasm with genetically encoded multimeric nanoparticles could bypass the requirement of Tem1GTP and correct spindle position for MEN activation. This localization/concentration-based GTPase signaling mechanism for Tem1 differs from the canonical Ras-like GTPase signaling paradigm and is likely relevant to other localization-based signaling scenarios.

Experimental analysis of the R290 variable geometry ejector with a spindle

The aim of this work was to experimentally analyze the variable geometry gas ejector with a spindle designed for operation with the natural working fluid propane (R290). The performance of the ejector was evaluated based on the mass entrainment ratio and critical temperature, being the most crucial parameters for optimizing the cooling capacity in ejector refrigeration systems, as well as the ejector efficiency using common literature notation. Additionally, local pressure drop measurements allowed for the determination of pressure profiles inside for different spindle positions used for capacity regulation. The experimentally defined ejector efficiency curves demonstrated the ability to control the ejector capacity by means of the spindle, irrespective of the unfavorable operating conditions. By decreasing the effective throat area using spindle, the ejector mass entertainment ratio increased by 35%. Spindle movement allowed for the decrease of the motive nozzle flow rate by up to 65%, while still maintaining the suction of the secondary flow of the ejector. Moreover, the behavior of a slight increase in suction nozzle flow with a decrease in motive nozzle flow has been observed for the initial movement of the spindle followed by a huge drop for further reduction of the effective throat area, confirming the previous conclusions shown in the literature.

Hydrodynamic Mechanism for Stable Spindle Positioning in Meiosis II Oocytes

Cytoplasmic streaming, the persistent flow of fluid inside a cell, induces intracellular transport, which plays a key role in fundamental biological processes. In meiosis II mouse oocytes (developing egg cells) awaiting fertilization, the spindle, which is the protein structure responsible for dividing genetic material in a cell, must maintain its position near the cell cortex (the thin actin network bound to the cell membrane) for many hours. However, the cytoplasmic streaming that accompanies this stable positioning would intuitively appear to destabilize the spindle position. Here, through a combination of numerical and analytical modeling, we reveal a hydrodynamic mechanism for stable spindle positioning beneath the cortical cap. We show that this stability depends critically on the spindle size and the active driving from the cortex and demonstrate that stable spindle positioning can result purely from a hydrodynamic suction force exerted on the spindle by the cytoplasmic flow. Our findings show that local fluid dynamic forces can be sufficient to stabilize the spindle, explaining robustness against perturbations not only perpendicular but also parallel to the cortex. Our results shed light on the importance of cytoplasmic streaming in mammalian meiosis. Published by the American Physical Society 2024

Mitotic spindle positioning protein serves as prognostic biomarker in patients with colorectal cancer

Background Colorectal cancer (CRC) ranks among the most aggressive types of cancer globally. Currently, clinical tumor prognostic biomarkers still lack accuracy. Mitotic spindle positioning (MISP) protein connects microtubules to the actin cytoskeleton and adhesive plaques, playing a critical role in spindle positioning, orientation, and the process of cell division. MISP can regulate the malignant biological functions of pancreatic cancer and intrahepatic cholangiocarcinoma and it acts as biomarker for prognosis, but its role in CRC remains unclear. Methods This study has collected 37 CRC tissue samples and 37 corresponding adjacent nontumor tissue samples, and 57 additional CRC tissues samples. Clinical data were obtained from the patients with CRC. MISP mRNA and protein expression levels were analyzed in normal control and CRC tissues using the GEPIA and Human Protein Atlas website. MISP protein levels in the collected tissues were analyzed using immunohistochemistry. Results MISP mRNA and protein expression levels were significantly increased in CRC tissues compared to adjacent nontumor tissues. Higher MISP protein levels were associated with distant metastasis, recurrence, and lower survival rates. Kaplan–Meier analysis showed that high expression levels of MISP protein were associated with recurrence and death in CRC patients. In addition, a high expression level of MISP protein, lymph node metastasis, and distance metastasis were risk factors for recurrence and a poor prognosis in patients with CRC. Conclusion Elevated MISP protein correlated with tumor metastasis, recurrence, and lower survival rates in patients with CRC, and thus, MISP has the potential to become a prognostic marker for CRC.

First person – Md Hashim Reza

ABSTRACT First Person is a series of interviews with the first authors of a selection of papers published in Journal of Cell Science, helping researchers promote themselves alongside their papers. Md Hashim Reza is first author on ‘ Expansion microscopy reveals characteristic ultrastructural features of pathogenic budding yeast species’, published in JCS. Md Hashim is a postdoc in the lab of Kaustuv Sanyal at JNCASR, Jakkur, India, investigating the evolution of factors regulating spindle positioning and spindle pole body (SPB) inheritance in non-model yeasts.

A noncanonical GTPase signaling mechanism controls exit from mitosis in budding yeast.

In the budding yeast Saccharomyces cerevisiae, exit from mitosis is coupled to spindle position to ensure successful genome partitioning between mother and daughter cell. This coupling occurs through a GTPase signaling cascade known as the mitotic exit network (MEN). The MEN senses spindle position via a Ras-like GTPase Tem1 which localizes to the spindle pole bodies (SPBs, yeast equivalent of centrosomes) during anaphase and signals to its effector protein kinase Cdc15. How Tem1 couples the status of spindle position to MEN activation is not fully understood. Here, we show that Cdc15 has a relatively weak preference for and Tem1's nucleotide state does not change upon MEN activation. Instead, we find that Tem1's nucleotide cycle establishes a localization-based concentration difference in the cell where only is recruited to the SPB, and spindle position regulates the MEN by controlling Tem1 localization. SPB localization of Tem1 primarily functions to promote Tem1-Cdc15 interaction for MEN activation by increasing the effective concentration of Tem1. Consistent with this model, we demonstrate that artificially tethering Tem1 to the SPB or concentrating Tem1 in the cytoplasm with genetically encoded multimeric nanoparticles could bypass the requirement of and correct spindle position for MEN activation. This localization/concentration-based GTPase signaling mechanism for Tem1 differs from the canonical Ras-like GTPase signaling paradigm and is likely relevant to other localization-based signaling scenarios.

MISP Is Overexpressed in Intestinal Metaplasia and Gastric Cancer.

Gastric cancer is the fifth most common cancer and the fourth cause of global cancer mortality. The identification of new biomarkers and drug targets is crucial to allow the better prognosis and treatment of patients. The mitotic spindle positioning (MISP) protein has the function of correcting mitotic spindle positioning and centrosome clustering and has been implicated in the cytokinesis and migration of cancer cells. The goal of this work was to evaluate the expression and clinical relevance of MISP in gastric cancer. MISP expression was evaluated by immunohistochemistry in a single hospital series (n = 286) of gastric adenocarcinomas and compared with normal gastric mucosa and intestinal metaplasia, a preneoplastic lesion. MISP was detected on the membrane in 83% of the cases, being overexpressed in gastric cancer compared to normal gastric mucosa (n = 10). Its expression was negatively associated with diffuse and poorly cohesive types. On the other hand, it was strongly expressed in intestinal metaplasia where it was associated with MUC2 and CDX2 expression. Furthermore, when we silenced MISP in vitro, a significant decrease in the viability of gastric carcinoma cells was observed. In conclusion, MISP is overexpressed in gastric cancer, being associated with an intestinal phenotype in gastric carcinogenesis and having a role in cellular proliferation.

PLD1 promotes spindle assembly and migration through regulating autophagy in mouse oocyte meiosis

ABSTRACT PLD1 has been implicated in cytoskeletal reorganization and vesicle trafficking in somatic cells; however, its function remains unclear in oocyte meiosis. Herein, we found PLD1 stably expresses in mouse oocytes meiosis, with direct interaction with spindle, RAB11A+ vesicles and macroautophagic/autophagic vacuoles. The genetic or chemical inhibition of PLD1 disturbed MTOC clustering, spindle assembly and its cortical migration, also decreased PtdIns(4,5)P2, phosphorylated CFL1 (p-CFL1 [Ser3]) and ACTR2, and their local distribution on MTOC, spindle and vesicles. Furthermore in PLD1-suppressed oocytes, vesicle size was significantly reduced while F-actin density was dramatically increased in the cytoplasm, the asymmetric distribution of autophagic vacuoles was broken and the whole autophagic process was substantially enhanced, as illustrated with characteristic changes in autophagosomes, autolysosome formation and levels of ATG5, BECN1, LC3-II, SQSTM1 and UB. Exogenous administration of PtdIns(4,5)P2 or overexpression of CFL1 hyperphosphorylation mutant (CFL1S3E) could significantly improve polar MTOC focusing and spindle structure in PLD1-depleted oocytes, whereas overexpression of ACTR2 could rescue not only MTOC clustering, and spindle assembly but also its asymmetric positioning. Interestingly, autophagy activation induced similar defects in spindle structure and positioning; instead, its inhibition alleviated the alterations in PLD1-depleted oocytes, and this was highly attributed to the restored levels of PtdIns(4,5)P2, ACTR2 and p-CFL1 (Ser3). Together, PLD1 promotes spindle assembly and migration in oocyte meiosis, by maintaining rational levels of ACTR2, PtdIns(4,5)P2 and p-CFL1 (Ser3) in a manner of modulating autophagy flux. This study for the first time introduces a unique perspective on autophagic activity and function in oocyte meiotic development. Abbreviations: ACTR2/ARP2: actin related protein 2; ACTR3/ARP3: actin related protein 3; ATG5: autophagy related 5; Baf-A1: bafilomycin A1; BFA: brefeldin A; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GOLGA2/GM130: golgin A2; GV: germinal vesicle; GVBD: germinal vesicle breakdown; IVM: in vitro maturation; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MI: metaphase of meiosis I; MII: metaphase of meiosis II; MO: morpholino; MTOC: microtubule-organizing center; MTOR: mechanistic target of rapamycin kinase; PB1: first polar body; PLA: proximity ligation assay; PLD1: phospholipase D1; PtdIns(4,5)P2/PIP2: phosphatidylinositol 4,5-bisphosphate; RAB11A: RAB11A, member RAS oncogene family; RPS6KB1/S6K1: ribosomal protein S6 kinase B1; SQSTM1/p62: sequestosome 1; TEM: transmission electron microscopy; TUBA/α-tubulin: tubulin alpha; TUBG/γ-tubulin: tubulin gamma; UB: ubiquitin; WASL/N-WASP: WASP like actin nucleation promoting factor

Mitotic Spindle Positioning (MISP) Facilitates Colorectal Cancer Progression by Forming a Complex with Opa Interacting Protein 5 (OIP5) and Activating the JAK2-STAT3 Signaling Pathway.

Patients with inflammatory bowel disease (IBD) who experience long-term chronic inflammation of the colon are at an increased risk of developing colorectal cancer (CRC). Mitotic spindle positioning (MISP), an actin-binding protein, plays a role in mitosis and spindle positioning. MISP is found on the apical membrane of the intestinal mucosa and helps stabilize and elongate microvilli, offering protection against colitis. This study explored the role of MISP in colorectal tumorigenesis using a database, human CRC cells, and a mouse model for colitis-induced colorectal tumors triggered by azoxymethane (AOM)/dextran sodium sulfate (DSS) treatment. We found that MISP was highly expressed in colon cancer patient tissues and that reduced MISP expression inhibited cell proliferation. Notably, MISP-deficient mice showed reduced colon tumor formation in the AOM/DSS-induced colitis model. Furthermore, MISP was found to form a complex with Opa interacting protein 5 (OIP5) in the cytoplasm, influencing the expression of OIP5 in a unidirectional manner. We also observed that MISP increased the levels of phosphorylated STAT3 in the JAK2-STAT3 signaling pathway, which is linked to tumorigenesis. These findings indicate that MISP could be a risk factor for CRC, and targeting MISP might provide insights into the mechanisms of colitis-induced colorectal tumorigenesis.

Measurement of ring spinning spun yarn spindle position based on binocular vision

Aiming at the problem of insufficient positioning accuracy of the ring spinning spun yarn automatic jointing machine using an automated guided vehicle platform after spun yarn breakage, a three-dimensional coordinate detection method of the ring spinning spun yarn spindle position based on binocular vision is proposed. The bottom column of the spindle was selected as the detection target, the contrast of the background of the bottom column of the spindle was physically enhanced, the picture of the bottom of the spindle was collected with a calibrated binocular camera, the two-dimensional coordinates of the midpoint of the bottom edge of the spindle bottom profile were obtained by the processing of the local picture, and the three-dimensional world coordinates of the midpoint of the bottom edge of the spindle bottom profile were calculated based on the method of single-pair feature-point matching. Comparing this paper’s single-pair feature-point matching method with the traditional multi-pair feature-point matching detection method (semi-global block matching), the experimental results showed that: the detection errors of both the single-pair feature-point matching method and the traditional semi-global block matching detection method were less than 2 mm; both methods satisfied the requirements of the positional detection accuracy of the spindle joints. The position detection method based on single-pair feature-point matching saved 80% of the program time compared with the traditional semi-global block matching method.

PAK1-Dependent Regulation of Microtubule Organization and Spindle Migration Is Essential for the Metaphase I-Metaphase II Transition in Porcine Oocytes.

P21-activated kinase 1 (PAK1) is a critical downstream target that mediates the effect of small Rho GTPase on the regulation of cytoskeletal kinetics, cell proliferation, and cell migration. PAK1 has been identified as a crucial regulator of spindle assembly during the first meiotic division; however, its roles during the metaphase I (MI) to metaphase II (MII) transition in oocytes remain unclear. In the present study, the potential function of PAK1 in regulating microtubule organization and spindle positioning during the MI-MII transition was addressed in porcine oocytes. The results showed that activated PAK1 was co-localized with α-tubulin, and its expression was increased from the MI to MII stage (p < 0.001). However, inhibiting PAK1 activity with an inhibitor targeting PAK1 activation-3 (IPA-3) at the MI stage decreased the first polar body (PB1) extrusion rate (p < 0.05), with most oocytes arrested at the anaphase-telophase (ATI) stage. IPA-3-treated oocytes displayed a decrease in actin distribution in the plasma membrane (p < 0.001) and an increase in the rate of defects in MII spindle reassembly with abnormal spindle positioning (p < 0.001). Nevertheless, these adverse effects of IPA-3 on oocytes were reversed when the disulfide bond between PAK1 and IPA-3 was reduced by dithiothreitol (DTT). Co-immunoprecipitation revealed that PAK1 could recruit activated Aurora A and transform acidic coiled-coil 3 (TACC3) to regulate spindle assembly and interact with LIM kinase 1 (LIMK1) to facilitate actin filament-mediated spindle migration. Together, PAK1 is essential for microtubule organization and spindle migration during the MI-MII transition in porcine oocytes, which is associated with the activity of p-Aurora A, p-TACC3 and p-LIMK1.

Reconfigurable open architecture control system with integrated digital twin for 3-axis woodworking milling machine

ABSTRACT This study proposes a reconfigurable open architecture control system for the Computer Numerical Control (CNC) woodworking educational machine tool called “EMCO F1 CNC”. On this machine physical hardware reconfiguration (spindle position can be vertical and horizontal) is possible. The idea was to develop a control system that would follow the configuration changes of the machine tool. The educational character of the machine and its reconfigurability demand possibilities for machining programme simulations in order to determine optimal workpiece position in the workspace and programme verification. Those are the reasons for developing a virtual machine tool integrated with the control system as a digital twin. The study presents a novel methodology for the development of control systems, merging the concept of “virtual commissioning” with iterative error-driven processes. The objective was to develop a dynamic control system capable of promptly adjusting to kinematics changes in real time. In a wood machining experiment, a test workpiece for machine tools for both machine configurations verified the operation of the developed control system.

Dynein directs prophase centrosome migration to control the stem cell division axis in the developing Caenorhabditis elegans epidermis.

The microtubule motor dynein is critical for the assembly and positioning of mitotic spindles. In Caenorhabditis elegans, these dynein functions have been extensively studied in the early embryo but remain poorly explored in other developmental contexts. Here, we use a hypomorphic dynein mutant to investigate the motor's contribution to asymmetric stem cell-like divisions in the larval epidermis. Live imaging of seam cell divisions that precede formation of the seam syncytium shows that mutant cells properly assemble but frequently misorient their spindle. Misoriented divisions misplace daughter cells from the seam cell row, generate anucleate compartments due to aberrant cytokinesis, and disrupt asymmetric cell fate inheritance. Consequently, the seam becomes disorganized and populated with extra cells that have lost seam identity, leading to fatal epidermal rupture. We show that dynein orients the spindle through the cortical GOA-1Gα-LIN-5NuMA pathway by directing the migration of prophase centrosomes along the anterior-posterior axis. Spindle misorientation in the dynein mutant can be partially rescued by elongating cells, implying that dynein-dependent force generation and cell shape jointly promote correct asymmetric division of epithelial stem cells.

The relationship between controlled ovarian stimulation protocol, meiotic spindle visibility, position of the meiotic spindle relative to the polar body in the human oocyte, and clinical outcomes following ICSI

AbstractPurposeTo investigate the effects of different controlled ovarian stimulation (COS) protocols, including the progestin‐primed ovarian stimulation (PPOS), long, short, and the gonadotropin‐releasing hormone antagonist protocols, on meiotic spindle visibility and position within the oocyte and clinical outcomes following ICSI.MethodsBefore ICSI, spindle position (θ) just below the polar body (PB) was defined as 0° and categorized as follows: θ = 0°, 0° &lt; θ ≤ 30°, 30° &lt; θ ≤ 60°, 60° &lt; θ ≤ 90°, 90° &lt; θ ≤ 180°, between the PB and the oolemma, and nonvisible. The clinical outcomes after ICSI were retrospectively analyzed.ResultsThe normal fertilization rate was significantly higher in oocytes with visible spindles than in oocytes with nonvisible spindles after each COS protocol, but did not differ based on spindle positioning (0° ≤ θ ≤ 180°). The rates of pregnancy, live birth/ongoing pregnancy, and miscarriage did not differ based on spindle visibility or positioning. In multinominal logistic regression analysis, female age was associated with spindle position, and the odds of a spindle located at 30° &lt; θ ≤ 60°, at 60° &lt; θ ≤ 90°, or at 90° &lt; θ ≤ 180° were increased relative to θ = 0° in older women (odds ratio; 1.020, 1.030, and 1.060, respectively; p &lt; 0.05).ConclusionMeiotic spindle positioning in the oocyte does not affect normal fertilization, blastulation, pregnancy, live birth/ongoing pregnancy, and miscarriage after ICSI, independent of the COS protocol used.

#267 : The Spindle Localization ICSI (SL-ICSI) Improves the Fertilization Rate Decreased by Aging

Background and Aims: The position of meiotic spindle in metaphase II oocyte can be different from the underneath of the first polar body. In order to prevent the meiotic spindle damage during ICSI, confirming the position of the spindle is effective. In this study, we analyzed the meiotic spindle deviation and outcomes of ICSI with the patient age. Method: From 2017 to 2022, 5620 oocytes with the first polar body underwent SL-ICSI were analyzed. SL-ICSI was performed 4-5 hours after oocyte retrieval. Furthermore, the meiotic spindle deviation from the polar body was classified according to the angle; Group A: 0-5∘, Group B: 6-45∘, Group C: 46-90∘, Group D: 90∘&lt;. The spindle invisible oocytes were regarded as group E. The average age, the fertilization rate and good blastocyst rate in each group were analyzed. Statistical analysis was performed by one-way ANOVA and Tukey’s post-hoc test. Results: The proportions of spindle-visible oocyteswere 98.7%; 41.2%, 36.1%, 14.6%, 6.8%, and 1.3% respectively, in group A to E. The mean agesin each group were 39.0, 39.6, 40.1, 40.6, and 40.7, respectively. In addition, the correlation between the age and the location of spindle was revealed (A-BCDE: [Formula: see text] &lt; 0.001). The fertilization rates were 84.8%, 84.9%, 84.7%, 83.5%, 50.7%, respectively, and significantly decreased in group E ([Formula: see text] &lt; 0.001). Abnormal fertilization rates were 4.3%, 5.1%, 6.4%, 7.3%, 20.5%, respectively, and significantly increased in group E ([Formula: see text] &lt; 0.001). Good blastocyst rates were 49.8%, 52.4%, 47.6%, 50.5%, 3.3%, respectively, and significantly decreased in group E ([Formula: see text] &lt; 0.001). Conclusion: The introduction of SL-ICSI can be effective for elderly patients. The meiotic spindle of oocytes is highly likely to deviate from the first polar body by aging. Furthermore, if the spindle-invisible oocyte is observed, ICSI should be carefully carried out.