Our previous study has suggested that paclitaxel-induced F-actin stabilization enhances the developmental competence of porcine in vitro fertilization (IVF) embryos; however, its role in bovine embryos remains unclear. Herein, we analyzed the embryo cleavage stages 48 h post-IVF to assess their correlation with blastocyst development and investigated the impact of paclitaxel-induced cytoskeletal stabilization. After IVF, bovine embryos were classified into two groups based on cleavage stage: LCG and ECG (late cleaved group; LCG, < 8-cell and early cleaved group; ECG, ≥ 8-cell). Bovine embryos cultured in the ECG developed to the blastocyst stage at a significantly higher rate (35.9 ± 9.5

Background: Heat stress (HS), driven by global climate change, adversely affects reproductive performance in livestock and mammals by impairing gamete development and reducing fertility. Exposure of oocytes to elevated temperatures during in vitro maturation (IVM) can induce persistent cellular stress that compromises subsequent embryonic development. However, the involvement of ATF5-mediated UPRmt signaling and GRP75-associated ER-mitochondria communication during early bovine embryogenesis under HS remains unclear. Methods: In this study, bovine cumulus–oocyte complexes (COCs) were matured for 22 h at either 38.5℃ (control; Con) or 41.5℃ (HS), followed by in vitro fertilization and embryo culture under standard conditions. Results: HS exposure during IVM significantly reduced cleavage and blastocyst formation rates (p < 0.01). Embryos derived from HS-exposed oocytes exhibited decreased mitochondrial membrane potential (ΔΨm), increased reactive oxygen species (ROS) production, and mitochondrial calcium (Ca2+) accumulation, suggesting disruption of GRP75 (known as HSPA9)-mediated ER-mitochondria communication. These embryos also showed increased expression of HSPA9 and HSPD1 at the cleavage stage, indicating persistence of stress signals beyond fertilization. Immunofluorescence analysis revealed enhanced nuclear localization of ATF5 in blastocysts, while qPCR analysis demonstrated increased expressions of ATF5-mediated UPRmt-related genes, including LONP1, HSPD1, and HSPA9. Conclusions: Collectively, these findings demonstrate that heat stress during oocyte maturation induces persistent mitochondrial dysfunction associated with GRP75-mediated mitochondrial Ca2+ dysregulation and ATF5-mediated UPRmt activation, which may represent an adaptive compensatory response in surviving bovine embryos.

Zearalenone (ZEN), a mycotoxin commonly found in animal feed, impairs the female reproductive function owing to its estrogen-mimicking ability. ZEN exposure induces endoplasmic reticulum (ER) stress, oxidative stress, and autophagy activation. However, the involvement of inositol-requiring enzyme 1 (IRE1)-mediated ER-phagy in ZEN-induced ER stress and embryonic competence during in vitro culture (IVC) remains unclear. Porcine embryos were cultured with 2.5, 5, and 10 μM ZEN for two days after in vitro fertilization (IVF) to assess blastocyst development. ZEN exposure caused a significant dose-dependent decrease in blastocyst formation, expanded blastocysts, and total cell number (p < 0.05). ZEN-treated embryos exhibited increased DNA fragmentation along with elevated mitochondrial and intracellular Ca²⁺ levels (p < 0.001), as confirmed using terminal deoxynucleotidyl transferase dUTP nick-end labeling assay and Rhod-2 and Fluo-4 AM staining. ER-tracker analysis revealed decreased ER fluorescence intensity and increased uneven cytoplasmic ER distribution in blastocysts exposed to 5 and 10 μM ZEN (p < 0.001). Western blotting showed activation of the unfolded protein response (UPR) with increased expression of glucose-regulated protein 78 (GRP78), p-IRE1, IRE1, p-c-Jun N-terminal kinase (JNK), JNK, C/EBP homologous protein (CHOP), and the autophagy marker microtubule-associated protein 1 light chain 3 beta (LC3B) (p < 0.05), whereas the ER-phagy receptor family with sequence similarity 134, member B was downregulated. Tauroursodeoxycholic acid (TUDCA) alleviated ZEN-induced ER stress, restoring ER distribution and calcium homeostasis, reducing DNA damage, and improving blastocyst development. These findings suggest that ZEN disrupts IRE1 signaling and suppresses ER-phagy during early porcine embryo development, whereas TUDCA alleviates ER stress and improves embryonic competence during IVC.

In this study, we investigated whether MIT-001, a small-molecule reactive oxygen species (ROS) scavenger, improves the re-expansion and viability of bovine blastocysts in response to vitrification-induced mitochondrial dysfunction and stress. Therefore, this study aimed to analyze the protective effects of MIT-001 on the mitochondrial function of bovine blastocysts following vitrification warming. In this experiment, MIT-001 (0.1 μM) was allocated to three culture conditions based on treatment period: (I) warming only (WARM); (II) vitrification only (VITR); and (III) both vitrification and warming (VITR-WARM), compared with the control (Non-treated). Survival analysis of cryopreserved bovine blastocysts revealed that MIT-001 supplementation during the warming period (WARM group) significantly improved (p < 0.01; Non-treated: 57.3 ± 2.3% vs WARM: 74.2 ± 7.3%) post-warm survival rates. It is noteworthy that surviving blastocysts in the WARM groups demonstrated significantly (p < 0.05) lower TUNEL positive cells (%) and a higher ratio of expanded blastocyst development compared to the other groups. Intracellular ROS, as well as mitochondrial and nuclear superoxide levels, were significantly reduced (p < 0.001) in the MIT-001-treated WARM group, accompanied by enhanced mitochondrial activation (MitoTracker Orange staining). Simultaneously, mitochondrial membrane potential, assessed using JC-1 staining, was elevated, whereas a reduction in mitochondrial fission marker dynamin-related protein 1 (DRP1) expression was observed in surviving blastocysts from the MIT-001-supplemented WARM group (p < 0.01). In addition, MIT-001 improved cytoskeletal stability by decreasing the aggregation thickness of filamentous actin (F-actin, p < 0.001; Non-treated: 10.73 μm vs. WARM: 6.03 μm) in bovine blastocyst of the WARM group. Finally, the enhanced developmental potential of vitrified-warmed blastocysts was linked to increased phospho-p38 mitogen-activated protein kinase (MAPK) expression exclusively in the WARM group compared to the other groups. Consequently, MIT-001 mitigates cryopreservation-induced cellular stress by improving mitochondrial function and regulates F-actin stabilization to enhance the viability and developmental potential of vitrified-warmed bovine blastocysts. These findings highlight the potential of MIT-001 to support cellular recovery and developmental capacity during cryopreservation, suggesting that it may play an effective protective role in bovine blastocyst cryopreservation.

Objective:To investigate the dual effects of anti-Müllerian hormone (AMH) on ovarian response, fertilization outcomes, and clinical pregnancy in assisted reproductive technology (ART) cycles. Methods:This retrospective cohort study screened 6,312 in vitro fertilization or intracytoplasmic sperm injection cycles performed between 2018 and 2023. After excluding cycles with missing AMH or body mass index (BMI) values, non-ART cycles, and extreme BMI values (<18 or ≥45 kg/m2), 4,399 cycles were included. AMH level, age, and BMI were evaluated as independent variables. Ovarian response (retrieved oocyte count) and normally fertilized embryos (two pronuclei) were analyzed using multivariable linear regression, and clinical pregnancy was evaluated using logistic regression. Results:AMH level showed a strong positive association with ovarian response (β=2.348; P<0.001), whereas age showed a significant negative association. After adjusting for age, BMI, fertilization method, and retrieved oocytes, AMH level showed a small but significant negative association with fertilized embryo count (β=-0.044; P=0.027). In the subgroup analysis, cycles with AMH ≥10 ng/mL showed higher oocyte yield but lower fertilization (0.561 vs. 0.694; P<0.001) and clinical pregnancy rates (5.7% vs. 23.1%; P<0.001) than those with AMH <10 ng/mL. Increasing age and AMH levels were independently associated with reduced odds of clinical pregnancy. Conclusion:AMH may reflect ovarian reserve quantity rather than optimal reproductive potential, showing a positive association with oocyte yield, but a negative association with fertilization and clinical pregnancy outcomes in ART cycles.

Post-ovulatory aging (POA) is associated with reduced fertilization rates and poor embryo quality both in vivo and in vitro. However, the relationship between nicotinamide adenine dinucleotide (NAD+) and the filamentous actin (F-actin) cytoskeleton in POA-induced oocytes remains unknown. Here, we investigated the mechanisms by which the NAD+ salvage pathways function in poor oocyte maturation upon POA through the F-actin cytoskeleton. Porcine oocytes were aged by extending in vitro maturation (IVM) for an additional 24 h to create a POA model. F-actin and adducin 1 (ADD1)-related spindle assembly were analyzed using immunofluorescence, western blotting, and RNA sequencing to identify key gene categories in the POA and IVM groups. To assess NAD+ function in restoring oocyte maturation, nicotinamide mononucleotide (NMN) was added and the maturation efficiency was evaluated. Expression of spindle assembly factors, F-actin cytoskeleton factors, aging markers, and NAD+-related genes was analyzed via quantitative polymerase chain reaction, immunofluorescence, and western blotting. We revealed unique interactions between the F-actin/ADD1-related cytoskeleton and aging factors (clusterin (CLU) and FAM111 trypsin-like peptidase A (FAM111A)) in poor-quality oocytes. POA oocytes were established with an extension of 24 h based on 44 h of IVM. They exhibited actin collapses and abnormal cortical F-actin, ADD1, and acetyl(Ac)-α-tubulin protein levels, which resulted in defective spindle assembly. RNA sequencing analysis was performed to identify differentially expressed genes involved in the oocyte viability response to aging, the cytoskeleton, and NAD metabolic processes using IVM and/or POA oocytes. This showed that NAD-binding genes were differentially expressed after POA induction, eight of which were downregulated compared with IVM oocytes. Importantly, activation of NAD+ pathways upon addition of NMN to the medium at 24 h after IVM rescued the maturation capability of POA oocytes with perturbations of spindle assembly and cortical F-actin. F-actin polymerization through NAD+ generated from NMN is an essential factor in determining oocyte quality. This effect is mediated by microtubules related to spindle assembly in POA oocytes.

Background: In mammals, DRP1 is a key regulator of mitochondrial fission during mitochondrial dynamics, whereas ATF5 promotes the mitochondrial unfolded protein response (UPRmt). Both pathways are essential for maintaining cellular homeostasis and protecting oocytes and embryos from external stressors. However, the relationship between ATF5 expression and DRP1 under heat stress conditions during porcine oocyte maturation remains unclear. Methods: In this study, we investigated the mitochondrial dynamics and ATF5 expression in porcine oocytes exposed to heat stress during in vitro maturation (IVM). Protein and gene expression levels were assessed using immunofluorescence staining, Western blotting, and quantitative PCR. Results: During IVM, both DRP1 and ATF5 expression were increased (p < 0.01) significantly. In contrast, heat stress markedly impaired (p < 0.05) meiotic progression and cumulus cell expansion. Mitochondrial dynamics were disrupted (p < 0.05), as fission and fusion markers displayed reciprocal changes relative to those in controls. Concomitantly, the expression of ATF proteins was significantly reduced (p < 0.01) under heat stress. Heat-stressed oocytes also exhibited decreased (p < 0.05) expression of genes involved in antioxidant defense and NAD metabolism, whereas autophagy- and apoptosis-related transcripts were significantly upregulated (p < 0.05). At the blastocyst stage, embryos derived from heat-stressed oocytes exhibited nuclear localization of the UPR-associated transcription factors ATF4, CHOP, and ATF5. Conclusions: Collectively, our findings suggest that heat stress disrupts mitochondrial dynamics and ATF5 expression during porcine oocyte maturation while the UPRmt pathway remains active during early embryonic development to mitigate heat-induced cellular damage.

Introduction Severe combined immunodeficiency (SCID) mini-pigs are a highly versatile model for human disease research and regenerative medicine. Objectives This study aims to generate a novel JAK3-deficient mini-pig model with a human-like immune system and to elucidate how JAK3 plays an important role in immune system. Methods JAK3 and RAG2 knockout (KO) mini-pigs were generated using CRISPR/Cas9 and somatic cell nuclear transfer. These mini-pigs were transferred to a sterilized isolator within a specific pathogen-free facility. Phenotypic characteristics and clinical manifestations were analyzed through histological and hematological analysis of SCID mini-pigs to explore the unique role of JAK3 in immune functions. Results JAK3 KO was characterized by defects in T and NK cells, very low levels of B cells, and a complete absence of thymus and lymph nodes. Notably, JAK3 KO mini-pigs had significantly reduced numbers of monocytes in peripheral blood, macrophages in tissue, and inflammatory cytokines, suggesting that JAK3 KO can induce a broad immunodeficiency that extends to the myeloid system as well as the lymphoid. Moreover, JAK3 KO mini-pigs had intestinal abnormalities similar to those of patients. Conclusion These results suggest that JAK3 KO mini-pigs can be used as an effective model for the development of therapies for SCID patients, as well as for regenerative medicine applications such as the development of patient-specific artificial organs.

Background: Post-ovulatory aging (POA) of oocytes is related to a decrease in the quality and quantity of oocytes caused by aging. Previous studies on the characteristics of POA have investigated injury to early embryonic developmental ability, but no information is available on its effects on mitochondrial fission and mitophagy-related responses. In this study, we aimed to elucidate the molecular mechanisms underlying mitochondrial fission and mitophagy in in vitro maturation (IVM) oocytes and a POA model based on RNA sequencing analysis. Methods: The POA model was obtained through an additional 24 h culture following the IVM of matured oocytes. NMN treatment was administered at a concentration of 25 μM during the oocyte culture process. We conducted MitoTracker staining and Western blot experiments to confirm changes in mitochondrial function between the IVM and POA groups. Additionally, comparative transcriptome analysis was performed to identify differentially expressed genes and associated changes in mitochondrial dynamics between porcine IVM and POA model oocytes. Results: In total, 32 common genes of apoptosis and 42 mitochondrial fission and function uniquely expressed genes were detected (≥ 1.5-fold change) in POA and porcine metaphase II oocytes, respectively. Functional analyses of mitochondrial fission, oxidative stress, mitophagy, autophagy, and cellular apoptosis were observed as the major changes in regulated biological processes for oocyte quality and maturation ability compared with the POA model. Additionally, we revealed that the activation of NAD+ by nicotinamide mononucleotide not only partly improved oocyte quality but also mitochondrial fission and mitophagy activation in the POA porcine model. Conclusions: In summary, our data indicate that mitochondrial fission and function play roles in controlling oxidative stress, mitophagy, and apoptosis during maturation in POA porcine oocytes. Additionally, we found that NAD+ biosynthesis is an important pathway that mediates the effects of DRP1-derived mitochondrial morphology, dynamic balance, and mitophagy in the POA model.

The cytotoxic mycotoxin deoxynivalenol (DON) reportedly has adverse effects on oocyte maturation and embryonic development in pigs. Recently, the interplay between cell apoptosis and endoplasmic reticulum (ER) stress has garnered increasing attention in embryogenesis. However, the involvement of the inositol-requiring enzyme 1 (IRE1)/c-jun N -terminal kinase (JNK)/C/EBP homologous protein (CHOP) pathways of unfolded protein response (UPR) signaling in DON -induced apoptosis in porcine embryos remains unknown. In this study, we revealed that exposure to DON (0.25 mu M) substantially decreased cell viability until the blastocyst stage in porcine embryos, concomitant with initiation of cell apoptosis through the IRE1/JNK/CHOP pathways in response to ER stress. Quantitative PCR confirmed that UPR signaling -related transcription factors were upregulated in DON -treated porcine blastocysts. Western blot analysis showed that IRE1/JNK/CHOP signaling was activated in DON -exposed porcine embryos, indicating that ER stress -associated apoptosis was instigated. The ER stress inhibitor tauroursodeoxycholic acid protected against DON -induced ER stress in porcine embryos, indicating that the toxic effects of DON on early developmental competence of porcine embryos can be prevented. In conclusion, DON exposure impairs the developmental ability of porcine embryos by inducing ER stress -mediated apoptosis via IRE1/JNK/CHOP signaling.

Malignant melanoma represents a form of skin cancer characterized by a bleak prognosis and heightened resistance to traditional therapies. Quercetin has demonstrated notable anti-carcinogenic, anti-inflammatory, anti-oxidant, and pharmacological effects across various cancer types. However, the intricate relationship between quercetin’s anti-cancer properties and ganglioside expression in melanoma remains incompletely understood. In this study, quercetin manifests specific anti-proliferative, anti-migratory, and cell-cycle arrest effects, inducing mitochondrial dysfunction and apoptosis in two melanoma cancer cell lines. This positions quercetin as a promising candidate for treating malignant melanoma. Moreover, our investigation indicates that quercetin significantly reduces the expression levels of ganglioside GD3 and its synthetic enzyme. Notably, this reduction is achieved through the inhibition of the FAK/paxillin/Akt signaling pathway, which plays a crucial role in cancer development. Taken together, our findings suggest that quercetin may be a potent anti-cancer drug candidate for the treatment of malignant melanoma.

Rat animal models are widely used owing to their relatively superior cognitive abilities and higher similarity compared with mouse models to human physiological characteristics. However, their use is limited because of difficulties in establishing embryonic stem cells and performing genetic modifications, and insufficient embryological research. In this study, we established optimal superovulation and fertilized–egg transfer conditions, including optimal hormone injection concentration (≥150 IU/kg of PMSG and hCG) and culture medium (mR1ECM), to obtain high-quality zygotes and establish in vitro fertilization conditions for rats. Next, sgRNA with optimal targeting activity was selected by performing PCR analysis and the T7E1 assay, and the CRISPR/Cas9 system was used to construct a rat model for muscular dystrophy by inducing a deficiency in the fukutin gene without any off-target effect detected. The production of fukutin knockout rats was phenotypically confirmed by observing a drop-in body weight to one-third of that of the control group. In summary, we succeeded in constructing the first muscular dystrophy disease rat model using the CRISPR/CAS9 system for increasing future prospects of producing various animal disease models and encouraging disease research using rats.

Cadmium (Cd) is toxic metal that can induce various diseases, such as cardiovascular, nervous, and reproductive systems. This study investigated the effect of Cd exposure on porcine oocyte maturation and the underlying mechanism. Porcine cumulus-oocyte complexes were exposed various Cd concentration and tauroursodeox-ycholic acid (TUDCA), an inhibitor of endoplasmic reticulum (ER) stress during in vitro maturation (IVM). After IVM, we evaluated meiotic maturation, ER stress, and oocyte quality by Cd exposure. Cd exposure inhibited cumulus cell expansion and meiotic maturation, increased oocyte degeneration, and induced ER stress. The levels of spliced XBP1 and ER stress-associated transcripts, markers of ER stress, were elevated in Cd-treated cumu-lus-oocyte complexes and denuded oocytes during IVM. Moreover, Cd-induced ER stress impaired oocyte quality by disrupting mitochondrial function and elevating intracellular reactive oxygen species levels while decreasing ER function. Interestingly, TUDCA supplementation significantly decreased the expression of ER stress-related genes and increased the quantity of ER compared with the Cd treatment. Additionally, TUDCA was also able to rescue excessive levels of ROS and restore normal mitochondrial function. Moreover, the addition of TUDCA under Cd exposure greatly ameliorated Cd-mediated detrimental effects on meiotic maturation and oocyte quality, including cumulus cell expansion and MII rate. These findings suggest that Cd exposure during IVM impairs the meiotic maturation of oocytes by inducing of ER stress.

AIMS:Mdivi-1 (Md-1) is a well-known inhibitor of mitochondrial fission and mitophagy. The mitochondrial superoxide scavenger Mito-TEMPO (MT) exerts positive effects on the developmental competence of pig embryos. This study aimed to explore the adverse effects of Md-1 on developmental capacity in porcine embryos and the protective effects of MT against Md-1-induced injury. MAIN METHODS:We exposed porcine embryos to Md-1 (10 and 50 μM) for 2 days after in vitro fertilization (IVF). MT (0.1 μM) treatment was applied for 4 days after exposing embryos to Md-1. We assessed blastocyst development, DNA damage, mitochondrial superoxide production, and mitochondrial distribution using TUNEL assay, Mito-SOX, and Mito-tracker, respectively. Subsequently, the expression of PINK1, DRP1, and p-DRP1Ser616 was evaluated via immunofluorescence staining and Western blot analysis. KEY FINDINGS:Md-1 compromised the developmental competence of blastocysts. Apoptosis and mitochondrial superoxide production were significantly upregulated in 50 μM Md-1-treated embryos, accompanied by a downregulation of p-DRP1Ser616, PINK1, and LC3B levels and lower mitophagy activity at the blastocyst stage. We confirmed the protective effects of MT against the detrimental effect of Md-1 on blastocyst developmental competence, mitochondrial fission, and DRP1/PINK1-mediated mitophagy activation. Eventually, MT recovered DRP1/PINK1-mediated mitophagy and mitochondrial fission by inhibiting superoxide production in Md-1-treated embryos. SIGNIFICANCE:MT protects against detrimental effects of Md-1 on porcine embryos by suppressing superoxide production. These findings expand available scientific knowledge on improving outcomes of IVF.

Rapamycin induces autophagosome formation and activity during oocyte maturation, improved fertilization ability of matured oocytes, and early embryonic developmental competence. However, potential changes in mitochondrial fission and mitophagy via regulation of autophagy in early porcine embryonic development have not been previously studied. Here, we investigated embryonic developmental ability and quality of porcine embryos 2 days after in vitro fertilization and following treatment with 1 and 10 nM rapamycin. As a results, 1 nM rapamycin exposure significantly improved (p < 0.05) blastocyst developmental competence compared to that in nontreated embryos (nontreated: 26.2 ± 5.7% vs. 1 nM rapamycin: 35.3 ± 5.1%). We observed autophagic (LC3B) and mitochondrial fission protein expression (dynamin-related protein-1 [DRP1] and pDRP1-Ser616) at the cleavage stage of 1 and 10 nM rapamycin-treated porcine embryos, using Western blot and immunofluorescence analyses. Interestingly, 1 nM rapamycin treatment significantly improved autophagy formation, mitochondrial activation, and mitochondrial fission protein levels (p < 0.05; p-DRP1 [Ser616]) at the cleavage stage of porcine embryos. Additionally, mitophagy was significantly increased in blastocysts treated with 1 nM rapamycin. In conclusion, our results suggest that rapamycin promotes blastocyst development ability in porcine embryos through mitochondrial fission, activation, and mitophagy in in vitro culture.

Ochratoxin A (OTA), a mycotoxin found in foods, has a deleterious effect on female reproduction owing to its endocrine-disrupting activity mediated through endoplasmic reticulum (ER) stress and reactive oxygen species (ROS) production. However, the mechanisms of OTA-induced ER stress in pig embryos during in vitro culture (IVC) are not yet fully understood. In the present study, porcine embryos were cultured for two days in an IVC medium supplemented with 0.5, 1.0, and 5.0 μM OTA, which led to an OTA-induced reduction in the developmental rate of blastocysts. The mRNA-seq transcriptome analysis revealed that the reduced blastocyst development ability of OTA-exposed porcine embryos was caused by ER stress, ultimately resulting in the accumulation of ROS and the occurrence of apoptosis. The expression levels of some UPR/PERK signaling-related genes (DDIT3, EIF2AK3, EIF2S1, NFE2L2, ATF4, EIF2A, and KEAP1) were found to differ in OTA-exposed pig embryos. OTA induces DNA damage by triggering an increase in RAD51/γ-H2AX levels and suppressing p-NRF2 activity. This effect is mediated through intracellular ROS and superoxide accumulation in the nuclei of porcine embryos. The cytotoxicity of OTA increased the activation of the PERK signal pathways (p-PERK, PERK, p-eIF2α, eIF2α, ATF4, and CHOP) in porcine embryos, with abnormal distribution of the ER observed around the nucleus. Collectively, our findings indicate that ER stress is a major cause of decline in the development of porcine embryos exposed to OTA. Therefore, OTA exposure induces ER stress and DNA damage via oxidative stress by disrupting PERK/NRF2 signaling activity in the developmental competence of porcine embryos during IVC.

Luteolin (Lut), a polyphenolic compound that belongs to the flavone subclass of flavonoids, possesses anti-inflammatory, cytoprotective, and antioxidant activities. However, little is known regarding its role in mammalian oocyte maturation. This study examined the effect of Lut supplementation during in vitro maturation (IVM) on oocyte maturation and subsequent developmental competence after somatic cell nuclear transfer (SCNT) in pigs. Lut supplementation significantly increased the proportions of complete cumulus cell expansion and metaphase II (MII) oocytes, compared with control oocytes. After parthenogenetic activation or SCNT, the developmental competence of Lut-supplemented MII oocytes was significantly enhanced, as indicated by higher rates of cleavage, blastocyst formation, expanded or hatching blastocysts, and cell survival, as well as increased cell numbers. Lut-supplemented MII oocytes exhibited significantly lower levels of reactive oxygen species and higher levels of glutathione than control MII oocytes. Lut supplementation also activated lipid metabolism, assessed according to the levels of lipid droplets, fatty acids, and ATP. The active mitochondria content and mitochondrial membrane potential were significantly increased, whereas cytochrome c and cleaved caspase-3 levels were significantly decreased, by Lut supplementation. These results suggest that Lut supplementation during IVM improves porcine oocyte maturation through the reduction of oxidative stress and mitochondria-mediated apoptosis.