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Calcium ions (Ca2+) regulate cell proliferation and differentiation and participate in various physiological activities of cells. The calcium transfer protein inositol 1,4,5-triphosphate receptor (IP3R), located between the endoplasmic reticulum (ER) and mitochondria, plays an important role in regulating Ca2+ levels. However, the mechanism by which IP3R1 affects porcine meiotic progression and embryonic development remains unclear. We established a model in porcine oocytes using siRNA-mediated knockdown of IP3R1 to investigate the effects of IP3R1 on porcine oocyte meiotic progression and embryonic development. The results indicated that a decrease in IP3R1 expression significantly enhanced the interaction between the ER and mitochondria. Additionally, the interaction between the ER and the mitochondrial Ca2+ ([Ca2+]m) transport network protein IP3R1-GRP75-VDAC1 was disrupted. The results of the Duolink II in situ proximity ligation assay (PLA) revealed a weakened pairwise interaction between IP3R1-GRP75 and VDAC1 and a significantly increased interaction between GRP75 and VDAC1 after IP3R1 interference, resulting in the accumulation of large amounts of [Ca2+]m. These changes led to mitochondrial oxidative stress, increased the levels of reactive oxygen species (ROS) and reduced ATP production, which hindered the maturation and late development of porcine oocytes and induced apoptosis. Nevertheless, after treat with [Ca2+]m chelating agent ruthenium red (RR) or ROS scavenger N-acetylcysteine (NAC), the oocytes developmental abnormalities, oxidative stress and apoptosis caused by Ca2+ overload were improved. In conclusion, our results indicated IP3R1 is required for meiotic progression and embryonic development by regulating mitochondrial calcium and oxidative damage.
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Hexabromocyclododecane (HBCD) has been detected in animals and humans blood. As an environment contamination, HBCD damages tissues and organs in animals and humans and produces cytotoxicity. In current study, we explored the effect of HBCD on premature testicular aging in vivo and in vitro. In vivo, C57 mice (8-week-old) were used as model to estimate the effect of HBCD on premature testicular aging. The results showed that testes were premature aging through measuring several aging-related markers (such as p16INK4a, hereafter p16; p21CIP, hereafter p21) in response to HBCD exposure for 20 weeks. In addition, HBCD exposure can cause oxidative stress and inflammation. Further, mouse spermatogonial cells (GC-1spg cells) were premature senescence after HBCD exposure by the evaluation of cellular senescence marker molecules. Hence, GC-1spg cell line was applied for cell model to investigate the molecule mechanism by which HBCD cause premature testicular aging., Through eliminating Fe2+ in senescent GC-1spg cells, cellular senescence was greatly alleviated. Thus, Fe2+ was identified as the key driver molecule in HBCD-induced premature cellular senescence. Next, we found that elevated iron levels in HBCD-triggered senescent GC-1spg cells were due to Nuclear receptor coactivator 4 (NCOA4)-mediated ferritinophagy. Furthermore, our results revealed that HBCD-induced senescence was caused by Fe2+ mediated oxidative stress. In summary, HBCD-induced premature testicular aging is dependent on NCOA4/Fe2+/ROS signaling molecule. The current study lays the foundation for further exploration of the effects of HBCD on reproductive toxicology.
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Senescência Celular , Hidrocarbonetos Bromados , Camundongos Endogâmicos C57BL , Coativadores de Receptor Nuclear , Espécies Reativas de Oxigênio , Testículo , Masculino , Animais , Testículo/efeitos dos fármacos , Testículo/patologia , Testículo/metabolismo , Hidrocarbonetos Bromados/toxicidade , Camundongos , Coativadores de Receptor Nuclear/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Senescência Celular/efeitos dos fármacos , Linhagem Celular , Ferro/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Senilidade Prematura/induzido quimicamente , HumanosRESUMO
Ochratoxin A (OTA) is a widespread environmental toxin that poses a serious threat to human and animal health. OTA has been shown to cause cellular and tissue damage and is a global public health problem. However, the effects of OTA on gastrointestinal aging have not been reported. The aim of this study was to investigate the effects of OTA on intestinal aging in vitro and in vivo. In vitro experiments showed that OTA induced cellular inflammation through calcium overload and oxidative stress, significantly up-regulated the expression of P16, P21, and P53 proteins, markedly increased senescence-associated ß-galactosidase activity (SA-ß-gal) positive cells, and obviously decreased the expression of proliferating cell nuclear antigen (PCNA) proteins, which led to intestinal cell senescence. Meanwhile, we found that treatment with ß-carotene ameliorated OTA-induced intestinal cell senescence. Consistent with the results of the in vitro experiments, in vivo studies showed that the intestinal aging of mice fed OTA was significantly higher than that of the control group. In conclusion, OTA may induce intestinal aging through calcium overload, oxidative stress and inflammation. This study lays a foundation for further research on the toxicological effects of OTA.
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Cálcio , Proteína 3 que Contém Domínio de Pirina da Família NLR , Ocratoxinas , Estresse Oxidativo , Transdução de Sinais , Ocratoxinas/toxicidade , Estresse Oxidativo/efeitos dos fármacos , Camundongos , Animais , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Cálcio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Intestinos/efeitos dos fármacos , Senescência Celular/efeitos dos fármacosRESUMO
Prolactin (PRL) and its receptor, PRLR, are closely related to the occurrence and development of breast cancer. hPRL-G129R, an hPRLR antagonist, has been found to induce apoptosis in breast cancer cells via mechanisms currently unknown. Recent studies have indicated that PRLR exhibits dual functions based on its membrane/nucleus localization. In that context, we speculated whether hPRL-G129R is a dual-function antagonist. We studied the internalization of the hPRLR-G129R/PRLR complex using indirect immunofluorescence and Western blot assays. We found that hPRL-G129R not only inhibited PRLR-mediated intracellular signaling at the plasma membrane, but also blocked nuclear localization of the receptor in T-47D and MCF-7 cells in a time-dependent manner. Clone formation and transwell migration assays showed that hPRL-G129R inhibited PRL-driven proliferation and migration of tumor cells in vitro. Further, we found that increasing concentrations of hPRL-G129R inhibited the nuclear localization of PRLR and the levels of signal transducer and activator of transcription (STAT) 5 in tumor-bearing mice and hPRL-G129R also exerted an antiproliferative effect in vivo. These results indicate that hPRL-G129R is indeed a dual-function antagonist. This study lays a foundation for exploring and developing highly effective agents against the proliferation and progression of breast malignancies.
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Neoplasias da Mama , Prolactina , Animais , Feminino , Humanos , Camundongos , Neoplasias da Mama/metabolismo , Proliferação de Células , Prolactina/farmacologia , Receptores da Prolactina/antagonistas & inibidores , Células Tumorais CultivadasRESUMO
Aflatoxin B1 (AFB1) is a recognized hazard environmental contaminant mainly found in cereal and fruits. The toxicity of AFB1 exposure to various organs has been revealed in some literature. In current study, we explored the effect of AFB1 exposure on premature aging/senescence of skin. In vivo, 8-week-old C57 mice were used as models to evaluate the effect of dietary AFB1 exposure on premature skin aging. The results showed that AFB1 exposure caused premature skin aging by testing aging markers. Additionally, AFB1 led to oxidative stress and inflammatory response. In vitro, AFB1 exposure triggered premature cellular senescence in mouse skin fibroblasts cells (L929 cells) by assessing a range of cellular senescence-related markers. Further, the potential molecular mechanism by which AFB1 induce the premature skin aging was studied. ROS and Ca2+ is proven to be the key molecules in AFB1-induced cellular senescence. Further, through eliminating Ca2+, AFB1-caused oxidative stress and cellular senescence were both attenuated, suggesting that Ca2+ overload in the mitochondria results in cellular senescence by increasing ROS production. Next, we analyzed the causes of Ca2+ overload, and results showed that AFB1 exposure induces Ca2+ overload through increasing the formation of mitoguardin (Miga) and vesicle-associated membrane protein (VAMP)-associated protein (Vap33)-mediated endoplasmic reticulum (ER)-mitochondria contact sites (ERMCS). AFB1 exposure also inhibited mitophagy, leading to accelerate L929 cell senescence. In short, combining in vivo and in vitro results, we demonstrate that exposure to AFB1 causes premature skin aging, which is dependent on ERMCS/Ca2+/ROS/ signaling axis. The current study suggests that prolonged exposure to AFB1 makes skin more vulnerable to damage.
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Senilidade Prematura , Envelhecimento da Pele , Animais , Camundongos , Senilidade Prematura/induzido quimicamente , Espécies Reativas de Oxigênio/metabolismo , Aflatoxina B1/toxicidade , Aflatoxina B1/metabolismo , Estresse Oxidativo , InflamaçãoRESUMO
BACKGROUND: Plastic pollution is greatly serious in the ocean and soil. Microplastics (MPs) degraded from plastic has threatened animals and humans health. The accumulation of MPs in the tissues and blood in animals and humans has been found. There is therefore a need to assess the toxicological effects of MPs on the reproductive system. RESULTS: In this study, we explored the effect of polystyrene microplastics (PS-MPs) on premature testicular aging in vitro and in vivo. In vitro, we found that testicular sertoli cells (TM4 cells) was prematurely senescent following PS-MPs treatment by the evaluation of a range of aging marker molecules (such as Sa-ß-gal, p16 and 21). TM4 cells were then employed for in vitro model to study the potential molecular mechanism by which PS-MPs induce the premature senescence of TM4 cells. NF-κB is identified as a key molecule for PS-MPs-induced TM4 cellular senescence. Furthermore, through eliminating reactive oxygen species (ROS), the activation of nuclear factor kappa B (NF-κB) was blocked in PS-MPs-induced senescent TM4 cells, indicating that ROS triggers NF-κB activation. Next, we analyzed the causes of mitochondrial ROS (mtROS) accumulation induced by PS-MPs, and results showed that Ca2+ overload induced the accumulation of mtROS. Further, PS-MPs exposure inhibits mitophagy, leading to the continuous accumulation of senescent cells. In vivo, 8-week-old C57 mice were used as models to assess the effect of PS-MPs on premature testicular aging. The results illustrated that PS-MPs exposure causes premature aging of testicular tissue by testing aging markers. Additionally, PS-MPs led to oxidative stress and inflammatory response in the testicular tissue. CONCLUSION: In short, our experimental results revealed that PS-MPs-caused testicular premature aging is dependent on Ca2+/ROS/NF-κB signaling axis. The current study lays the foundation for further exploration of the effects of microplastics on testicular toxicology.
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Senilidade Prematura , Humanos , Masculino , Animais , Camundongos , Microplásticos/toxicidade , Poliestirenos/toxicidade , Plásticos , NF-kappa B , Espécies Reativas de OxigênioRESUMO
The dynamic balance of Ca2+ in oocytes promotes the recovery of the meiotic arrest phase, consequently promoting oocyte maturation. Hence, the analysis of the maintenance and role of calcium homeostasis in oocytes has important guiding significance for obtaining high-quality eggs and maintaining the development of preimplantation embryos. Inositol 1,4,5-trisphosphate receptors (IP3Rs) are calcium channel proteins that regulate the dynamic balance between the endoplasmic reticulum (ER) and mitochondrial Ca2+. Nevertheless, the expression and role of IP3R in normal pig oocytes have not been reported, and other studies have focused on the role of IP3R in damaged cells. The purpose of this study was to investigate the potential role of IP3R in regulating calcium homeostasis in oocyte maturation and early embryonic development. Our results showed that IP3R1 is stably expressed at different stages of porcine oocyte meiosis, IP3R1 gradually converges to the cortex, and cortical clusters are formed in MII stages. The loss of IP3R1 activity contributeds to the failure of porcine oocyte maturation and cumulus cell expansion, as well as the obstruction of polar body excretion. Further analysis showed that IP3R1 plays an important role in affecting calcium balance by regulating the IP3R1-GRP75-VDAC1 channel between mitochondria and the endoplasmic reticulum (ER) during porcine oocyte maturation. Inhibiting IP3R1 expression-induced ER dysfunction, contributeding to ER calcium concentration ([Ca2+]ER) release outwards into mitochondria and causing mitochondrial free calcium concentration ([Ca2+]m) overload and mitochondrial oxidative stress, which was confirmed by the increase in the level of reactive oxygen species (ROS) and apoptosis. Thereby, IP3R1 plays an important role in affecting calcium balance by regulating the IP3R1-GRP75 -VDAC1 channel between mitochondria and the ER during porcine oocyte maturation, inhibiting IP3R1 expression-induced calcium overload and mitochondrial oxidative stress, and increasing ROS levels and apoptosis.
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Cálcio , Oogênese , Animais , Suínos , Cálcio/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Oócitos/fisiologia , Cálcio da Dieta , Desenvolvimento EmbrionárioRESUMO
This experiment was conducted to evaluate the growth performance, growth regulating factors, and liver morphology of chicks hatched from egg-laying breeding hens dietary supplemented with additives (ß-carotene). Hy-line breeding hens were allocated into three groups with three replicates/group. The dietary treatments were as follows: basal diet as a control (Con), basal diet supplemented with 120 (ßc-L) or 240 (ßc-H) mg/kg of ß-carotene diet. After 6 weeks, the eggs were collected and incubated. The hatched chicks were fed the same diet. The results showed that chicks in the ßc-L group increased in body weight at 21 days (p < 0.01). At 42 days, chicks in the ßc-H group showed a significant increase in tibia length (p < 0.05). The liver index increased in the ßc-L and ßc-H groups at 7 days (p < 0.05). Serum HGF (7, 14, 21, and 42 days) and leptin (14 days) were significantly increased in the group supplemented with ßc. Hepatic GHR (14 days), IGF-1R (14 days), and LEPR (21 days) mRNA expression were significantly increased. In addition, there was an increase in PCNA-positive cells in the liver of chicks in the ßc group. In conclusion, the addition of ß-carotene to the diet of laying breeder hens was more advantageous in terms of growth performance and liver development of the offspring.
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Galinhas , beta Caroteno , Animais , Feminino , Galinhas/genética , beta Caroteno/farmacologia , beta Caroteno/metabolismo , Dieta/veterinária , Suplementos Nutricionais , Ração Animal/análise , FígadoRESUMO
BACKGROUND: Nutrition drives immunity and health in animals, and maternal immunity benefits offspring. In our previous study, a nutritional intervention strategy was found to promote the immunity of hens, which subsequently improved immunity and growth in offspring chicks. Maternal effects clearly exist, but how are mothers' immune advantages transferred to their offspring, and how do they benefit them? RESULTS: Here, we traced the beneficial effects back to the process of egg formation in the reproductive system, and we focused on the embryonic intestinal transcriptome and development, as well as on maternal microbial transfer in offspring. We found that maternal nutritional intervention benefits maternal immunity, egg hatching, and offspring growth. The results of protein and gene quantitative assays showed that the transfer of immune factors into egg whites and yolks depends on maternal levels. Histological observations indicated that the promotion of offspring intestinal development begins in the embryonic period. Microbiota analyses suggested that maternal microbes transfer to the embryonic gut from the magnum to the egg white. Transcriptome analyses revealed that offspring embryonic intestinal transcriptome shifts are related to development and immunity. Moreover, correlation analyses showed that the embryonic gut microbiota is correlated with the intestinal transcriptome and development. CONCLUSIONS: This study suggests that maternal immunity positively influences offspring intestinal immunity establishment and intestinal development beginning in the embryonic period. Adaptive maternal effects might be accomplished via the transfer of relatively large amounts of maternal immune factors and by shaping of the reproductive system microbiota by strong maternal immunity. Moreover, reproductive system microbes may be useful resources for the promotion of animal health. Video Abstract.
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Galinhas , Microbioma Gastrointestinal , Animais , Feminino , Humanos , Herança Materna , Desenvolvimento Infantil , Perfilação da Expressão GênicaRESUMO
In vitro-cultured oocytes are separated from the follicular micro-environment in vivo and are more vulnerable than in vivo oocytes to changes in the external environment. This vulnerability disrupts the homeostasis of the intracellular environment, affecting oocyte meiotic completion, and subsequent embryonic developmental competence in vitro. Glycine, one of the main components of glutathione (GSH), plays an important role in the protection of porcine oocytes in vitro. However, the protective mechanism of glycine needs to be further clarified. Our results showed that glycine supplementation promoted cumulus cell expansion and oocyte maturation. Detection of oocyte development ability showed that glycine significantly increased the cleavage rate and blastocyst rate during in vitro fertilization (IVF). SMART-seq revealed that this effect was related to glycine-mediated regulation of cell membrane structure and function. Exogenous addition of glycine significantly increased the levels of the anti-oxidant GSH and the expression of anti-oxidant-related genes (glutathione peroxidase 4 [GPX4], catalase [CAT], superoxide dismutase 1 [SOD1], superoxide dismutase 2 [SOD2], and mitochondrial solute carrier family 25, member 39 [SLC25A39]), decreased the lipid peroxidation caused by reactive oxygen species (ROS) and reduced the level of malondialdehyde (MDA) by enhancing the functions of mitochondria, peroxisomes and lipid droplets (LDs) and the levels of lipid metabolism-related factors (peroxisome proliferator activated receptor coactivator 1 alpha [PGC-1α], peroxisome proliferator-activated receptor γ [PPARγ], sterol regulatory element binding factor 1 [SREBF1], autocrine motility factor receptor [AMFR], and ATP). These effects further reduced ferroptosis and maintained the normal structure and function of the cell membrane. Our results suggest that glycine plays an important role in oocyte maturation and later development by regulating ROS-induced lipid metabolism, thereby protecting against biomembrane damage.
Production of high-quality gametes is the premise of livestock reproduction and conservation of germplasm resources, especially high-quality oocytes, as oocyte quality determines the quality of offspring. Due to the limitations in approaches and the number of mature oocytes in vivo, in vitro maturation (IVM) culture has become an important way to obtain mature oocytes. However, IVM-cultured oocytes are separated from the follicular microenvironment in vivo and are, thus, more vulnerable than in vivo oocytes to changes in the external environment. Our study was conducted to determine if exogenous supplementation of glycine, the highest content of amino acids in oviduct fluid and follicular fluid, can improve oocyte maturation efficiency in vitro, and analyze the mechanism of glycine. This study demonstrated that glycine can maintain redox balance and block reactive oxygen species-induced lipid peroxidation, thereby protecting against biomembrane damage and reducing the occurrence of ferroptosis to maintain normal oocyte development function. This study will provide a theoretical basis for preventing and improving oxidative damage during oocyte culture in vitro.
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Antioxidantes , Técnicas de Maturação in Vitro de Oócitos , Gravidez , Feminino , Suínos , Animais , Espécies Reativas de Oxigênio/metabolismo , Técnicas de Maturação in Vitro de Oócitos/veterinária , Antioxidantes/metabolismo , Peroxidação de Lipídeos , Glicina/farmacologia , Desenvolvimento Embrionário , Oócitos/fisiologia , Blastocisto , Glutationa/metabolismoRESUMO
The liver is the largest digestive organ in the human body. The increasing incidence of chronic liver fibrosis is one of the major health challenges in the world. Liver fibrosis is a wound-healing response to acute or chronic cellular damage of liver tissue. At present, despite a series of research progress on the pathophysiological mechanism of fibrosis that has been made, there is still a gap in identifying antifibrotic targets and converting them into effective treatments. Therefore, it is extremely important to seek a molecular target that can alleviate or reverse liver fibrosis, which has important scientific and clinical significance. In the current study, to evaluate the therapeutic effect of HBO1 as a molecular target on liver aging and fibrosis, naturally-aged mice and CCL4-induced liver fibrosis mice were used as animal models, and multiple experiments were performed. Experimental results showed that HBO1 knockdown could strongly mitigate the accumulation of hepatic collagen by Masson and Sirius Red staining. Further study showed that HBO1 knockdown reduced the expression of fibrosis-related marker molecules (α-SMA, collagen type I (ColI), and fibronectin). Further work showed that HBO1 knockdown could significantly alleviate HSC activation. On this basis, we analyzed the underlying mechanism by which HBO1 alleviates liver fibrosis. It was found that HBO1 knockdown may modulate liver fibrosis by regulating the processes of EMT, inflammation, and oxidative stress. We further studied the effect of HBO1 knockdown on liver aging and aging-related liver fibrosis, and the results showed that HBO1 knockdown could significantly reduce the level of aging-related liver fibrosis and relieve liver aging. In conclusion, we systematically investigated the potential of HBO1 as a therapeutic target to attenuate liver fibrosis and liver aging. The current study found a crucial target for liver fibrosis and liver-aging therapy, which has laid a solid foundation for the liver fibrosis-related research.
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Tetracloreto de Carbono , Cirrose Hepática , Camundongos , Humanos , Animais , Idoso , Tetracloreto de Carbono/efeitos adversos , Cirrose Hepática/terapia , Cirrose Hepática/tratamento farmacológico , Fígado/metabolismo , Estresse Oxidativo , Envelhecimento , Células Estreladas do FígadoRESUMO
Senescence is associated with a decline in physiological function, which is accompanied by onset of diseases. Growth hormone (GH) is a class of growth-promoting cytokines with reduced secretion in aging populations. However, the effect of senescence on GH bioactivity is not fully understood in human mesenchymal stem cells (hMSCs). In this work, GH-induced cellular behavior and intracellular signaling transduction were explored in senescent hMSCs. Therefore, hMSCs were used to establish a senescence model by H2O2 treatment for this study. First, we investigated the effects of cellular senescence on the cell behavior of GH. The experimental results suggested that GH could not be internalized into the nucleus, and a significant reduction in GH internalization into the cytoplasm was observed in senescent hMSCs compared to the control group. Second, the effect of cellular senescence on GH-mediated intracellular signaling pathways was investigated by Western blotting. For this, the signaling molecule activation of Janus kinase 2 (JAK2)/signal transducer and activator transcription (STAT) stimulated by GH was detected. Our data indicated that the signaling intensity of p-JAK2, p-STAT5, p-STAT3 and p-STAT1 was considerably weakened. Taken together, these findings provide important insights into the impaired effects of cellular senescence on the biological activity of GH.
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Hormônio do Crescimento , Células-Tronco Mesenquimais , Humanos , Peróxido de Hidrogênio/farmacologia , Senescência Celular , Ciclo CelularRESUMO
Monobutyl phthalate (MBP) is the main metabolite of dibutyl phthalate (DBP) in vivo. MBP has a stable structure, can continuously accumulate in living organisms, and has the potentially to harm animal and human reproductive function. In the ovarian follicle microenvironment, MBP may lead to defects in follicular development and steroid production, abnormal meiotic maturation, impaired ovarian function and other reproductive deficits. In this study, SMART-seq was used to investigate the effects of MBP exposure on the in vitro maturation (IVM) and development of porcine oocytes. The results showed that differentially expressed genes after MBP exposure were enriched in the biological processes cytoskeleton, cell apoptosis, endoplasmic reticulum (ER) and mitochondria. Glycine (Gly) improved the developmental potential of porcine oocytes by regulating mitochondrial and ER function. The effect of Gly in protecting oocytes against MBP-induced damage was studied. The results showed that the addition of Gly significantly decreased the rate of MBP-induced spindle abnormalities, decreased the frequency of MBP-induced mitochondria-associated ER membrane (MAM) interactions, and downregulated the protein and gene expression of the linkage molecules Mitofusin 1 (MFN1) and Mitofusin 2 (MFN2) in the MAM. Additionally, treatment with Gly restored the distribution of the 1,4,5-triphosphate receptor 1 (IP3R1) and voltage-dependent anion channel 1 (VDAC1), further decreasing the intracellular free calcium concentration ([Ca2+]i) levels and mitochondrial Ca2+ ([Ca2+]m) , increasing the ER Ca2+ ([Ca2+]ER) levels, and thus significantly increasing the ER levels and mitochondrial membrane potential (ΔΨ m). Gly also decreased the levels of reactive oxygen species (ROS) and increased the levels of Glutathione (GSH), oocyte apoptosis-related indicators (Caspase-3 activity and Annexin V) and oocyte apoptosis-related genes (BAX, Caspase 3 and AIFM1). Our results suggest that Gly can ameliorate microtubule cytoskeleton abnormalities and improve oocyte maturation by reducing the defective mitochondrial-ER interactions caused by MBP exposure in vitro.
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Glicina , Oócitos , Animais , Apoptose , Retículo Endoplasmático , Feminino , Glutationa/metabolismo , Glicina/metabolismo , Glicina/farmacologia , Glicina/uso terapêutico , Humanos , Mitocôndrias , Ácidos Ftálicos/toxicidade , Espécies Reativas de Oxigênio/metabolismo , SuínosRESUMO
Epidermal growth factor (EGF) is an effective cytoprotective peptide. It is the main nutritional factor involved in the development of the intestinal tract. It has many important biological effects on the intestinal mucosa. After binding to epidermal growth factor receptor (EGFR), it initiates a signal transduction cascade to jointly promote the migration, proliferation, and differentiation of various cell types. Heat stress severely affects the intestinal health of livestock and is becoming increasingly prevalent due to the yearly increase in ambient temperature and intestinal diseases. However, the effect of heat stress on the activity and signaling of EGF/EGFR in intestinal cells is still unclear. Therefore, rat intestinal crypt epithelial cell line (IEC6) was used as a model to explore this issue, and the results showed that EGF/EGFR is internalized into IEC6 cells in a time-dependent manner under physiological conditions. However, the activity of EGF/EGFR was altered under heat stress. Furthermore, we explored the effect of heat stress on EGF/EGFR-activated signaling transduction in IEC6 cells, and the results showed that levels of factors involved in EGFR-mediated intracellular signaling (such as EGFR, signal transducers and activators of transcription 3/protein kinase B, and extracellular regulatory kinase 1/2) were downregulated under heat stress. In summary, this study shows that heat stress could damage the biological activity and intracellular signaling of EGF/EGFR. These findings have scientific importance in the field of animal husbandry; and lay the foundation for the further study of the biological activities of EGF/EGFR in the intestine.
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Fator de Crescimento Epidérmico , Receptores ErbB , Animais , Fator de Crescimento Epidérmico/farmacologia , Receptores ErbB/metabolismo , Resposta ao Choque Térmico , Mucosa Intestinal/metabolismo , Fosforilação , Ratos , Transdução de SinaisRESUMO
At present, heat stress caused by the thermal environment is the main factor that endangers the reproductive function of animals. Growth hormone (GH) is a polypeptide hormone, the biological function of reproductive organs has been reported, and it has many important physiological functions in the body. However, so far, the behavior and signal transduction of GH in testicular cells under heat stress are still unclear. To this end, in the current work, we use a swine testicular cell line (ST) as an in vitro model to explore the cell behavior and intracellular signaling profile of porcine growth hormone (pGH) under heat stress; the results showed that when cells were under heat stress, pGH and GHR were basically not internalized, and a large number of them accumulated on the cell membrane. In addition, we also studied the effect of pGH on the JAK2-STATs signaling pathway and IGF-1 expression under heat stress, we found that the ability of pGH to activate the JAK-STATs signaling pathway and IGF-1 under heat stress was greatly reduced (p < 0.05). In conclusion, our research shows that when cells undergo heat stress, the internalization of pGH and GHR were inhibited, and the activation of the JAK2-STATs signaling pathway and IGF-1 expression were reduced; this lays a solid foundation for further research on the effect of pGH on swine testicular tissue under thermal environment.
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Hormônio do Crescimento , Receptores da Somatotropina , Animais , Hormônio do Crescimento/metabolismo , Hormônio do Crescimento/farmacologia , Resposta ao Choque Térmico , Fator de Crescimento Insulin-Like I , Receptores da Somatotropina/metabolismo , Transdução de Sinais , SuínosRESUMO
Porcine growth hormone (pGH) has many important biological functions and roles, and the biological activity of pGH is closely related with its cell behavior and characteristics. However, so far, the behavior of pGH in swine testicular cell remains unclear. For this, in the current work, the swine testicular cell line (ST) was used as an in vitro model, and CLSM (Confocal laser scanning microscope), IFA (Indirect immunofluorescence assay), FCM (Flow cytometry) and WB (Western-blotting) were used to explore the pGH's cell behivior and function, and the results showed that pGH and GHR could internalize into ST cell and transported to the nucleus. Furthermore, we studied the internalization kinetics of pGH and GHR on ST cell, and found that pGH and GHR internalizes into ST cell in a time-dependent manner. More importantly, we also investigated the potential molecular functions of pGH-GHR after it entered into the cell nuclei. The results indicated that nuclear-localized GHR could participate in cell proliferation by regulating the signal intensity of STAT5. In summary, our current research shows that the nuclear-localized pGH-GHR participates in the cell proliferation of ST cell, which lays a solid foundation for further research on the regulatory effect of pGH on testicular tissue.
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Hormônio do Crescimento , Receptores da Somatotropina , Animais , Núcleo Celular/metabolismo , Hormônio do Crescimento/metabolismo , Hormônio do Crescimento/farmacologia , Hepatócitos/metabolismo , Receptores da Somatotropina/metabolismo , Transdução de Sinais , SuínosRESUMO
The endoplasmic reticulum (ER) is a multifunctional organelle in the cytoplasm that plays important roles in female mammalian reproduction. The endoplasmic reticulum and mitochondria interact to maintain the normal function of cells by maintaining intracellular calcium homeostasis. As proven by previous research, glycine (Gly) can regulate the intracellular free calcium concentration ([Ca2+]i) and enhance mitochondrial function to improve oocyte maturation in vitro. The effect of Gly on ER function during oocyte in vitro maturation (IVM) is not clear. In this study, we induced an ER stress model with thapsigargin (TG) to explore whether Gly can reverse the ER stress induced by TG treatment and whether it is associated with calcium regulation. The results showed that the addition of Gly could improve the decrease in the average cumulus diameter, the first polar body excretion rate caused by TG-induced ER stress, the cleavage rate and the blastocyst rate. Gly supplementation could reduce the ER stress induced by TG by significantly improving the ER levels and significantly downregulating the expression of genes related to ER stress (Xbp1, ATF4, and ATF6). Moreover, Gly also significantly alleviated the increase in reactive oxygen species (ROS) levels and the decrease in mitochondrial membrane potential (ΔΨ m) to improve mitochondrial function in porcine oocytes exposed to TG. Furthermore, Gly reduced the [Ca2+]i and mitochondrial Ca2+ ([Ca2+]m) levels and restored the ER Ca2+ ([Ca2+]ER) levels in TG-exposed porcine oocytes. Moreover, we found that the increase in [Ca2+]i may be caused by changes in the distribution and expression of inositol 1,4,5-triphosphate receptor (IP3R1) and voltage-dependent anion channel 1 (VDAC1), while Gly can restore the distribution and expression of IP3R1 and VDAC1 to normal levels. Apoptosis-related indexes (Caspase 3 activity and Annexin-V) and gene expression Bax, Cyto C, and Caspase 3) were significantly increased in the TG group, but they could be restored by adding Gly. Our results suggest that Gly can ameliorate ER stress and apoptosis in TG-exposed porcine oocytes and can further enhance the developmental potential of porcine oocytes in vitro.
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Soybean agglutinin (SBA) has a toxic effect on most animals. The anti-nutritional mechanisms of SBA are not fully understood, in terms of cell survival activity and metabolism of intestinal cells. This study aims to investigate the effects of SBA on the cell cycle, apoptosis, and to verify the mechanism of SBA anti-nutritional characters based on proteomic-based analysis. The IPEC-J2 cell line was cultured with medium containing 0.0, 0.5, or 2.0 mg/mL SBA. With increasing SBA levels, the percentage of the cells at G0/G1 phase, cell apoptosis rates, expressions of Bax and p21, and the activities of Casp-3 and Casp-9 were increased, while cyclin D1 and Bcl-2 expressions were declined (p < 0.05). The proteomic analysis showed that the numbers of differentially expressed proteins, induced by SBA, were mainly enriched in different pathways including DNA replication, base excision repair, nucleus excision repair, mismatch repair, amide and peptide biosynthesis, ubiquitin-mediated proteolysis, as well as structures and functions of mitochondria and ribosome. In conclusion, the anti-nutritional mechanism of SBA is a complex cellular process. Such process including DNA related activities; protein synthesis and metabolism; signal-conducting relation; as well as subcellular structure and function. This study provides comprehensive information to understand the toxic mechanism of SBA in monogastrics.
RESUMO
ß-Carotene displays antioxidant and anti-inflammatory activities and prevents the development of cancer. Ulcerative colitis (UC) is a kind of inflammatory bowel disease that is accompanied by a certain risk of colon cancer. However, the role of ß-carotene in the modulation of gut microbiota and UC improvement is unclear. In this research, the properties of ß-carotene on anti-inflammatory and the composition of gut microbiota were evaluated in a rat model of UC induced by dextran sulfate sodium (DSS). The results revealed that ß-carotene significantly (p < 0.05) decreased the severity of colitis in rats, as assessed using body weight (6.00 ± 1.73%), colon length (22.23 ± 0.53%), and disease activity index, and improved the structure of the colon damaged. Moreover, colonic levels of proinflammatory cytokines were significantly lower following ß-carotene supplementation. ß-Carotene intervention also lowered the expression levels of phosphorylated p65 (0.60 ± 0.02), p38 (0.57 ± 0.00), Erk (0.63 ± 0.04), and JNK (0.70 ± 0.00). The result of the relative abundance of gut microbiota showed that DSS administration significantly changed the microbial structure at the phylum and genus levels of rats. Furthermore, ß-carotene treatment significantly increased the abundance of Faecalibacterium, the levels of which negatively correlated with the levels of inflammatory cytokines. Faecalibacterium may be a potential target in the alleviation of DSS-induced UC. ß-Carotene can alleviate DSS-induced UC through the regulation of gut microbiota. This study provides a reference for the rational use of ß-carotene in the treatment of UC. PRACTICAL APPLICATION: ß-Carotene can relieve ulcerative colitis and regulate the gut microbiota; the nutritional intervention of ß-carotene enhancing animal health.
Assuntos
Anti-Inflamatórios/farmacologia , Colite Ulcerativa/tratamento farmacológico , Sulfato de Dextrana/toxicidade , Modelos Animais de Doenças , Microbioma Gastrointestinal/efeitos dos fármacos , beta Caroteno/farmacologia , Animais , Colite Ulcerativa/induzido quimicamente , Colite Ulcerativa/microbiologia , Citocinas/metabolismo , Masculino , Provitaminas/farmacologia , Ratos , Ratos Sprague-DawleyRESUMO
Mitochondria play an important role in controlling oocyte developmental competence. Our previous studies showed that glycine (Gly) can regulate mitochondrial function and improve oocyte maturation in vitro. However, the mechanisms by which Gly affects mitochondrial function during oocyte maturation in vitro have not been fully investigated. In this study, we induced a mitochondrial damage model in oocytes with the Bcl-2-specific antagonist ABT-199. We investigated whether Gly could reverse the mitochondrial dysfunction caused by ABT-199 exposure and whether it is related to calcium regulation. Our results showed that ABT-199 inhibited cumulus expansion, decreased the oocyte maturation rate and the intracellular glutathione (GSH) level, caused mitochondrial dysfunction, which was confirmed by decreased mitochondrial membrane potential (ΔΨm) and the expression of mitochondrial function-related genes PGC-1α, and increased reactiveoxygenspecies (ROS) levelsand the expression of apoptosis-associated genes Bax, Caspase-3, and Cyto C.More importantly, ABT-199-treated oocytes showed an increase in the intracellular free calcium concentration ([Ca2+]i) and had impaired cortical type 1 inositol 1,4,5-trisphosphate receptors (IP3R1) distribution. Nevertheless, treatment with Gly significantly ameliorated mitochondrial dysfunction, oxidative stress, and apoptosis, and Gly also regulated [Ca2+]i levels and IP3R1 cellular distribution, which further protects oocyte maturation in ABT-199-induced porcine oocytes.Taken together, our results indicate that Gly has a protective action against ABT-199-induced mitochondrial dysfunction in porcine oocytes.