Comparative analysis of MAPK and PI3K/AKT pathway activation and inhibition in human and canine melanoma

The lack of advanced animal models of human cancers is considered a barrier to developing effective therapeutics. Canine and human melanomas are histologically disparate but show similar disease progression and response to therapies. The purpose of these studies was to compare human and canine melanoma tumours and cell lines regarding MAPK and PI3K/AKT signalling dysregulation, and response to select molecularly targeted agents. Pathway activation was investigated via microarray and mutational analysis. Growth inhibition and cell cycle effects were assessed for pathway inhibitors AZD6244 (MAPK) and rapamycin (PI3K/AKT) in human and canine melanoma cells. Human and canine melanoma share similar differential gene expression patterns within the MAPK and PI3K/AKT pathways. Constitutive pathway activation and similar sensitivity to AZD6244 and rapamycin was observed in human and canine cells. These results show that human and canine melanoma share activation and sensitivity to inhibition of cancer‐related signalling pathways despite differences in activating mutations.     

Biphasic activation of PI3K/Akt and MAPK/Erk1/2 signaling pathways in bovine herpesvirus type 1 infection of MDBK cells

ABSTRACT: Many viruses have been known to control key cellular signaling pathways to facilitate the virus infection. The possible involvement of signaling pathways in bovine herpesvirus type 1 (BoHV-1) infection is unknown. This study indicated that infection of MDBK cells with BoHV-1 induced an early-stage transient and a late-stage sustained activation of both phosphatidylinositol 3-kinase (PI3K)/Akt and mitogen activated protein kinases/extracellular signal-regulated kinase 1/2 (MAPK/Erk1/2) signaling pathways. Analysis with the stimulation of UV-irradiated virus indicated that the virus binding and/or entry process was enough to trigger the early phase activations, while the late phase activations were viral protein expression dependent. Biphasic activation of both pathways was suppressed by the selective inhibitor, Ly294002 for PI3K and U0126 for MAPK kinase (MEK1/2), respectively. Furthermore, treatment of MDBK cells with Ly294002 caused a 1.5-log reduction in virus titer, while U0126 had little effect on the virus production. In addition, the inhibition effect of Ly294002 mainly occurred at the post-entry stage of the virus replication cycle. This revealed for the first time that BoHV-1 actively induced both PI3K/Akt and MAPK/Erk1/2 signaling pathways, and the activation of PI3K was important for fully efficient replication, especially for the post-entry stage.     

ROS介导的氧化应激与自噬

自噬是真核细胞所特有的细胞内物质成分被溶酶体降解过程的统称。生命体借此清除细胞内的废物，重建结构从而维持蛋白质代谢平衡及细胞内环境稳定。氧化应激是指体内氧化与抗氧化作用失衡，倾向于氧化，导致中性粒细胞炎性浸润，蛋白酶分泌增加，产生大量活性氧中介物（ROS），而ROS直接参与细胞存活和死亡调节。大量研究表明，氧化应激中产生的ROS在多种条件下都是自噬的重要调节因子，它能诱导自噬发生，而自噬能通过不同的信号通路来缓解氧化应激造成的损伤，从而保护细胞存活。ROS在多种条件下都是自噬的重要调节因子。作者主要对自噬的形成过程、氧化应激诱导自噬产生机制（包括调控mTOR信号通路、丝裂原活化蛋白激酶（MAPK）信号通路机制）及自噬缓解氧化应激的途径（mTOR信号通路、PI3K介导的信号通路和调控p53等）进行综述，以期为畜牧生产中通过调控自噬缓解动物氧化应激的措施提供理论依据。     

Electrolysed reduced water decreases reactive oxygen species-induced oxidative damage to skeletal muscle and improves performance in broiler chickens exposed to medium-term chronic heat stress

1. The present study was designed to achieve a reduction of reactive oxygen species (ROS)-induced oxidative damage to skeletal muscle and to improve the performance of broiler chickens exposed to chronic heat stress.

2. Chickens were given a control diet with normal drinking water, or diets supplemented with cashew nut shell liquid (CNSL) or grape seed extract (GSE), or a control diet with electrolysed reduced water (ERW) for 19 d after hatch. Thereafter, chickens were exposed to a temperature of either 34°C continuously for a period of 5 d, or maintained at 24°C, on the same diets.

3. The control broilers exposed to 34°C showed decreased weight gain and feed consumption and slightly increased ROS production and malondialdehyde (MDA) concentrations in skeletal muscle. The chickens exposed to 34°C and supplemented with ERW showed significantly improved growth performance and lower ROS production and MDA contents in tissues than control broilers exposed to 34°C. Following heat exposure, CNSL chickens performed better with respect to weight gain and feed consumption, but still showed elevated ROS production and skeletal muscle oxidative damage. GSE chickens did not exhibit improved performance or reduced skeletal muscle oxidative damage.

4. In conclusion, this study suggests that ERW could partially inhibit ROS-induced oxidative damage to skeletal muscle and improve growth performance in broiler chickens under medium-term chronic heat treatment.     

Role of intracellular kinases in the regulation of equine eosinophil migration and actin polymerization

Inappropriately activated eosinophils can contribute to disease pathogenesis and intracellular signalling pathways that regulate functional responses may represent a therapeutic target. Little is known about intracellular signalling in equine eosinophils and this study examined the role of phospholipase C (PLC) and a range of protein kinases on responses to histamine and CCL11. Histamine (10(-4) M) or CCL11 (5.6 x 10(-9) M)-induced actin polymerization, migration and superoxide production by eosinophils from healthy horses were compared in the presence and absence of selective kinase inhibitors. Inhibition of phosphatidylinositol-3 kinase (PI3K) significantly reduced the response in each assay. In contrast, whilst inhibition of PLC decreased actin polymerization and superoxide production, an increase in migration was observed; the latter effect was also seen when protein kinase C (PKC) was inhibited. With the exception of histamine-induced migration, which was significantly reduced by blocking extracellular regulated kinase (ERK)1/2, activation of ERK1/2, p38 MAPK and tyrosine kinase did not appear to play an important role in the responses studied. These results suggest that equine eosinophil activation by histamine and CCL11 is mediated through PI3K. Whilst PLC activation is required for actin polymerization and superoxide production, migration may be negatively regulated by PLC and PKC. These kinases represent potential targets for modulating eosinophil activation by multiple stimuli.     

Regulation of platelet activating factor-induced equine platelet activation by intracellular kinases

Lipopolysaccharide (LPS) can activate equine platelets directly or indirectly, via leukocyte-derived platelet activating factor (PAF). Thromboxane (Tx) production by LPS-stimulated equine platelets requires p38 MAPK and this kinase has been suggested as a therapeutic target in endotoxaemia. The present study has utilised selective inhibitors to investigate the role of p38 MAPK and two other kinases, phosphatidylinositol-3 kinase (PI3K) and protein kinase C (PKC), in regulating PAF-induced Tx production, aggregation and 5-HT release in equine platelets, and the modification of these responses by LPS. LPS enhanced PAF-induced 5-HT release, an effect that was reduced by the p38 MAPK inhibitor, SB203580 (60 ± 8% reduction; n  = 6). SB203580 did not affect responses to PAF alone; whereas inhibition of PKC reduced PAF-induced 5-HT release, Tx production and aggregation (maximal inhibition by the PKCδ inhibitor, rottlerin: 69 ± 13%, 63 ± 14% and 97 ± 1%, respectively; n  = 6). Wortmannin and LY249002, which inhibit PI3K, also caused significant inhibition of PAF-induced aggregation (maximal inhibition 78 ± 3% and 88 ± 2%, respectively; n  = 6). These data suggest that inhibition of platelet p38 MAPK may be of benefit in equine endotoxaemia by counteracting some of the effects of LPS. However, detrimental effects of platelet activation mediated by PAF and not enhanced by LPS are unlikely to be markedly affected.     

Possible involvement of phosphatidylinositol 3-kinase in the maintenance of metaphase II attest in porcine oocytes matured in vitro

It has been reported that phosphatidylinositol 3-kinase (PI3K)-protein kinase B (PKB) pathway plays a crucial role in the meiotic resumption and progression to the metaphase II (MII) stage of oocytes. However, the role of this pathway in meiotic arrest at the MII stage (cytostatic activity) is not well understood. In this study the effect of a PI3K inhibitor, LY294002, on the MAPK and p34^cdc2 kinase activities of matured porcine oocytes was examined. After maturation culture, both the MAPK and p34^cdc2 kinase activities in the oocytes were gradually decreased in a time-dependent manner. Although 25 µmol/L LY294002 did not affect either the MAPK or p34^cdc2 kinase activities, 50 µmol/L LY294002 suppressed the PKB phosphorylation and slightly decreased MAPK activity, but not the p34^cdc2 kinase activity. Therefore the effect of 10 µmol/L Ca²⁺ ionophore which was reported as inducing a transient decrease of p34^cdc2 kinase but not MAPK activities, was also examined in LY294002-treated oocytes. By additional treatment with LY294002 after Ca²⁺ ionophore, both the MAPK and p34^cdc2 kinase activities were decreased in a time-dependent manner, concomitantly with improvement of pronuclear formation. Therefore, we concluded that PI3K is involved in the maintenance of MAPK activity in matured porcine oocytes.     

Regulation of COX-2 expression in canine prostate carcinoma: increased COX-2 expression is not related to inflammation

BACKGROUND: Cyclooxygenase-2 (COX-2) expression has been documented in human and canine prostate carcinoma (PCA). Canine PCA is a histologically heterogeneous tumor, sometimes including inflammatory infiltrates. However, it is unknown whether COX-2 expression in canine PCA is related to the histologic type of tumor, to the presence of inflammation, or to both. Moreover, little is known about the mechanisms regulating COX-2 expression in neoplastic tissue. HYPOTHESIS: COX-2 expression is related to the presence of inflammation in canine PCA and correlates with the degree of tumor differentiation. METHODS: The expression of COX-2 was examined in 28 cases of canine PCA by immunohistochemistry. In addition, a neoplastic and a nonneoplastic canine prostatic cell line were used to investigate the effects of interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), phorbol 12-myristate 13-acetate (PMA), epithelial growth factor (EGF), and specific signal transduction pathway inhibitors on COX-2 expression. RESULTS: Twenty-four of the 28 prostate tumors showed COX-2 expression. The presence of inflammatory infiltrates in tumor tissue was associated with lower COX-2 expression scores. In vitro, TNF-alpha, IL-6, and EGF increased COX-2 expression in nonneoplastic cells but not in PCA cells, where baseline expression was high. COX-2 expression in PCA cells could be suppressed by means of specific phosphatidyl inositol-3 kinase (PI3K), protein kinase C (PKC), or inhibitor of extracellular signal-related kinase (ERK/MAPK) inhibitors. CONCLUSIONS AND CLINICAL IMPORTANCE: COX-2 is expressed in canine PCA; however, expression is not related to the presence of inflammatory infiltrates. This conclusion is further supported by the finding that the cytokines TNF-alpha and IL-6 and their involved signaling pathways do not stimulate COX-2 expression in malignant canine prostate cells.     

Roles of IGF-I and the estrogen, androgen and IGF-I receptors in estradiol-17beta- and trenbolone acetate-stimulated proliferation of cultured bovine satellite cells

Although numerous studies have shown that both androgenic and estrogenic steroids increase rate and efficiency of muscle growth in steers, there is little consensus as to their mechanism of action. A combined estradiol 17beta (E2)/trenbolone acetate (TBA) implant causes a significant increase in muscle IGF-I mRNA and both E2 and TBA stimulate a significant increase in IGF-I mRNA level in bovine satellite cell (BSC) cultures in media containing 10% fetal bovine serum (FBS). Consequently, increased IGF-I expression may play a role in anabolic-steroid-enhanced muscle growth. However, even though treatment of cultured BSC with E2 or TBA in media containing 1% IGFBP-3-free swine serum (SS) results in increased proliferation there is no effect on IGF-I mRNA expression, suggesting that increased IGF-I expression may not be responsible for anabolic-steroid-enhanced BSC proliferation. To further examine the role of estrogen, androgen and IGF-I receptors and their respective ligands in E2- and TBA-stimulated BSC proliferation, we assessed the effects of specific inhibitors on E2- or TBA-stimulated proliferation of BSC. Both ICI 182 780 (an estrogen receptor blocker) and flutamide (an inhibitor of androgen receptor) suppressed (p<0.05) E2- and TBA-stimulated BSC proliferation, respectively. JB1 (a competitive inhibitor of IGF-I binding to type I IGF receptor) reduced (p<0.05) both E2- and TBA-stimulated proliferation in BSC cultures. Both the Raf-1/MAPK kinase (MEK)1/2/ERK1/2, and the phosphatidylinositol 3-kinase (PI3K)/Akt pathways play significant roles in the actions of IGF-I on proliferation and differentiation of myogenic cells. PD98059, an inhibitor of the MAPK pathway, and wortmannin, an inhibitor of the PI3K pathway, both suppressed (p<0.05) E2- and TBA-stimulated proliferation of cultured BSC. Our data suggest that IGF-I plays a role in E2- and TBA-stimulated proliferation of cultured BSC even in the absence of increased IGF-I expression.     

Measurement of phagocytosis and oxidative burst of canine neutrophils: high variation in healthy dogs

Quantitative analysis of phagocytosis and oxidative burst in canine polymorphonuclear (PMN) cells was performed by flow cytometry techniques. Different concentrations of phorbol myristate acetate (PMA) were used to modulate PMN phagocytosis. A low concentration of PMA (3 nmol) resulted in increased phagocytic activity of canine PMN, which could not be enhanced by higher dosages. Experiments with a reference cell population showed high losses of PMN, most probably by adherence to plastic material. It was possible to avoid this loss by layering all ingredients on cushions of Histopaque. However, Histopaque had a negative influence on the phagocytic activity of canine PMN. The use of PMA led to a dosage-dependent increase in the oxidative burst measured by the production of reactive oxygen species (ROS). Cushions of Histopaque were used to avoid cell loss. There was no negative influence of Histopaque on ROS formation. Storage of canine PMN for 24 h at room temperature had no negative influence on phagocytosis or oxidative burst measurements. Variations in the ROS assays conducted by two different examiners could be eliminated by use of a Histopaque-cushion.     

Involvement of the ERK1/2 MAPK pathway in insulin-induced S6K1 activation in avian cells

In mammals, insulin regulates S6K1, a key enzyme involved in the control of protein synthesis, via the well-documented phosphoinositide-3'kinase (PI3K) pathway. Conversely, S6K1 is activated by insulin in avian muscle despite the relative insulin insensitivity of the PI3K pathway in this tissue. Mitogen-activated protein kinase (MAPK) cascade is another insulin sensitive pathway. The aim of this study was to explore the potential involvement of the ERK1/2 MAPK pathway in the control of p70 S6 kinase (S6K1) in avian species. Firstly, we characterized ERK1/2 MAPK in various chicken tissues. ERK2 was the only isoform detected in avian species whatever the tissue studied. We also showed that ERK2 is activated in vivo by insulin in chicken muscle. The regulation and the role of ERK2 in insulin signaling were next investigated in chicken hepatoma cells (LMH) and primary myoblasts. Insulin stimulation led to ERK2 and S6K1 phosphorylation, and concomitantly increased kinase activity. U0126, an inhibitor of the ERK MAPK pathway, completely abolished insulin-induced S6K1 phosphorylation and activity in chicken myoblasts, whereas its effect was only partial in LMH cells. In conclusion, these results show that ERK1/2 MAPK is involved in the control of S6K1 by insulin in chicken cells, particularly myoblasts.     

Ankyrin repeat and suppressor of cytokine signaling (SOCS) box-containing protein (ASB) 15 alters differentiation of mouse C2C12 myoblasts and phosphorylation of mitogen-activated protein kinase and Akt

Ankyrin repeat and suppressor of cytokine signaling box-containing protein (ASB) 15 is a novel ASB gene family member predominantly expressed in skeletal muscle. We have previously reported that overexpression of ASB15 delays differentiation and alters protein turnover in mouse C(2)C(12) myoblasts. However, the extent of ASB15 regulation of differentiation and molecular pathways underlying this activity are unknown. The extracellular signal-regulated kinase (Erk) 1/2 and phosphatidylinositol-3 kinase-Akt (PI3K/Akt; Akt is also known as protein kinase B) signaling pathways have a role in skeletal muscle growth. Activation (phosphorylation) of the Erk1/2 signaling pathway promotes proliferation, whereas activation of the PI3K/Akt signaling pathway promotes myoblast differentiation. Accordingly, we tested the hypothesis that ASB15 controls myoblast differentiation through its regulation of these kinases. Stably transfected myoblasts overexpressing ASB15 (ASB15+) demonstrated decreased differentiation, whereas attenuation of ASB15 expression (ASB15-) increased differentiation. However, ASB15+ cells had less abundance of the phosphorylated mitogen-activated protein kinase (active) form, despite decreased differentiation relative to control myoblasts (ASB15Con). The mitogen-activated protein kinase kinase inhibitor, U0126, effectively decreased mitogen-activated protein kinase phosphorylation and stimulated differentiation in ASB15- and ASB15Con cells. However, inhibition of the Erk1/2 pathway was unable to overcome the inhibitory effect of overexpressing ASB15 on differentiation (ASB15+), suggesting that the Erk1/2 pathway is likely not the predominant mediator of ASB15 activity on differentiation. Expression of ASB15 also altered phosphorylation of the PI3K/Akt pathway, as ASB15+ and ASB15- cells had decreased and increased Akt phosphorylation, respectively. These data were consistent with observed differences in differentiation. Administration of IGF-I, a PI3K/Akt activator, in ASB15+ was able to partially override the previously observed phenotype of delayed differentiation, whereas administration of the PI3K/ Akt inhibitor, LY294002, decreased phosphorylation of Akt and differentiation of all cell lines similar to the untreated ASB15+ myoblasts. These results provide initial evidence that ASB15 has a role in early myoblast differentiation and that its effects may be mediated in part by the PI3K/Akt signal transduction pathway.     

The 5‐lipoxygenase inhibitor tepoxalin induces oxidative damage and altered PTEN status prior to apoptosis in canine osteosarcoma cell lines

The 5‐lipoxygenase (5‐LOX) inhibitor tepoxalin has been shown to slow canine osteosarcoma (OSA) tumour xenografts growth, yet the mechanisms are poorly elucidated. Further examination of tepoxalin in canine OSA cell lines shows that tepoxalin treated cells undergo apoptosis through caspase‐3 activation and annexin staining. Interestingly, apoptosis is superseded by an increase in reactive oxygen species (ROS), as measured by activation of dihydrorhodamine 123 and mitosox. This increase in ROS appears to be related to the 5‐LOX inhibitor regardless of cellular 5‐LOX status, and was not observed after treatment with the tepoxalin metabolite RWJ20142. Additionally, 5‐LOX inhibition by tepoxalin appears to increase phosphatase and tensin (PTEN) homolog activity by preventing its alkylation or oxidation. PTEN modification or inhibition allows phosphoinositide‐3 (PI3) kinase activity thereby heightening activation of protein kinase B (AKT) phosphorylation. Our data suggest that off target oxidation and LOX inhibition play roles in the apoptotic response.     

Oxidative Stress and Interaction of Endothelin Receptors in Airways of Clinically Healthy Horses

Endothelin-1 (ET-1), a physiological as well as an inflammatory mediator, causes contraction of airway smooth muscles by binding to endothelin-A (ETA) and endothelin-B (ETB) receptors. We investigated the interaction between endothelin receptors on contractions and oxidative stress of bronchial rings of clinically healthy horses. Rings for response studies were set in tissue chambers containing oxygenated Tyrode’s solution. Concentration-response relationships were determined for ET-1 on control rings, rings incubated with ETA antagonist (BQ123), rings incubated with ETB antagonists (IRL 1038 and BQ788), and rings incubated with all antagonists combined. For oxidative stress studies, rings were incubated for 30 minutes separately with ET-1, ETA antagonist + ET-1, ETB antagonists + ET-1, and all three antagonists + ET-1. The control rings were not treated with any agents. Electron paramagnetic resonance (EPR) method was used to measure total reactive oxygen species (ROS), superoxide, and peroxynitrite. Results showed that ET-1 caused dose-dependent contractions which were increased by ETA blockade and decreased by ETB blockade. Combined blockade significantly inhibited the response to ET-1. Regarding the oxidative stress, ET-1 and its antagonists significantly reduced the total ROS. Superoxide production was significantly decreased by ETA antagonist and completely abolished when ETA and ETB antagonists were combined. Peroxynitrite production was not changed significantly. The study suggested that an interaction between ET receptors exists in the expression of contractile responses and oxidative stress. ET-1 attenuated oxidative stress by decreasing total ROS. ETA receptors may be primarily responsible for superoxide production. It appears that ET-1 does not affect peroxynitrite production in normal equine tissues.     

氨基酸在炎症性肠病中的作用及其信号通路

肠道是营养物质消化、吸收的主要器官，但易受外界环境刺激，导致炎症性肠病（inflammatory bowel disease，IBD）的发生，严重危害动物肠道健康。膳食氨基酸在促进肠道发育、维持肠道健康方面发挥重要作用，其主要通过调控腺苷酸活化蛋白激酶（AMP-activated kinase，AMPK）、雷帕霉素靶蛋白（mammalian target of rapamycin，mTOR）、丝裂原活化蛋白激酶（mitogen-activated protein kinase，MAPK）、Toll样受体（Toll-like receptors，TLRs）、核苷酸结合寡聚化结构域（nucleotide binding oligomerization domain，NOD）/核因子-κB（nuclear factor kappa-B，NF-κB）等信号通路影响肠上皮细胞生理活动、改善肠道屏障功能、减轻肠道氧化损伤、调节炎性因子的产生、提高内源抗菌肽表达，进而预防和治疗IBD。本文综述了IBD的基本特征、氨基酸在IBD中的作用及其信号通路，以及氨基酸在畜禽生长中维持肠道健康的作用与应用，为膳食营养素防治IBD提供有效线索与策略。     

Gene expression pattern in canine mammary osteosarcoma

Canine mammary sarcomas are usually very aggressive and easily metastasize. Unfortunately the biology of this type of tumor is not well known because they are a very rare type of tumors. The aim of this study was to find differences in gene expression patterns in canine mammary osteosarcomas (malignant) versus osteomas (benign) using DNA microarrays. Our microarray experiment showed that 11 genes were up-regulated in osteosarcoma in comparison to osteoma whereas 36 genes were down-regulated. Among the up-regulated genes were: PDK1, EXT1, and EIF4H which are involved in AKT/PI3K and GLI/Hedgehog pathways. These genes play an important role in cell biology (cancer cell proliferation) and may be essential in osteosarcoma formation and development. Analyzing the down-regulated genes, the most interesting seemed to be HSPB8 and SEPP1. HSPB8 is a small heat shock protein that plays an important role in cell cycle regulation, apoptosis, and breast carcinogenesis. Also SEPP1 may play a role in carcinogenesis, as its down-regulation may induce oxidative stress possibly resulting in carcinogenesis. The preliminary results of the present study indicate that the up-regulation of three genes EXT1, EIF4H, and PDK1 may play an essential role in osteosarcoma formation, development and proliferation. In our opinion the cross-talk between GLI/Hedgehog and PI3K/AKT pathways may be a key factor to increase tumor proliferation and malignancy.     

Effects of Amino Acids on Inflammatory Bowel Disease and Its Signaling Pathways

The intestine is the primary organ responsible for digestion and absorption of nutrients and is frequently subjected to external environmental stimulations leading to the development of inflammatory bowel disease (IBD), which can cause serious harm to intestinal health in animals. Dietary amino acids play important roles in promoting intestinal development and maintaining intestinal health and exert diverse effects through multiple signaling pathways on the prevention and treatment of IBD, including affecting the physiological activities of intestinal epithelial cells, improving intestinal barrier function, reducing intestinal oxidative damage, regulating the production of inflammatory cytokines, and promoting the expression of endogenous antimicrobial peptides, and involved in main signaling pathways including AMP-activated kinase (AMPK), mammalian target of rapamycin (mTOR), mitogen-activated protein kinase (MAPK), Toll-like receptors (TLRs), nucleotide binding oligomerization domain (NOD)/nuclear factor kappa-B (NF-κB). In this review, basic characteristics of IBD, effects and involved signaling pathways of the amino acids on IBD, and effects and applications of amino acids in maintaining intestinal health in livestock and poultry production were reviewed, so as to provide effective clues and strategies for dietary nutrients in the prevention and treatment of IBD.     

辣木叶多糖对过氧化氢诱导奶牛乳腺上皮细胞氧化损伤的保护作用

奶牛乳腺上皮细胞(BMECs)在奶牛泌乳期代谢旺盛,导致活性氧(ROS)大量产生,从而诱发氧化应激。辣木叶多糖(MLP)能有效清除ROS和自由基,但其是否具有缓解BMECs氧化损伤的潜力尚不清楚。因此,本文以MLP为添加剂,探究其对过氧化氢(H2O2)诱导BMECs氧化损伤的保护作用。本试验首先将分离的BMECs置于含有不同浓度H2O2的培养基中培养2 h建立氧化损伤模型,以确定H2O2的适宜浓度;随后在培养基中加入不同浓度MLP溶液培养BMECs 2 h,以确定MLP适宜浓度;最终选用浓度为500μmol/L的H2O2和4 mg/mL的MLP用于本试验。试验设置4个组,分别为对照组1(BMECs)、对照组2(BMECs+MLP)、损伤组(BMECs+H2O2)、保护组(BMECs+MLP+H2O2),每组3个重复。试验对BMECs中ROS数量、BMECs凋亡以及BMECs中过氧化氢酶(CAT)、谷胱甘肽过氧化物酶(GSH-Px)、超氧化物歧化酶(SOD)活性和丙二醛(MDA)含量进行检测。结果表明:1)ROS检测结果显示,MLP抑制了细胞内ROS的生成。2)Hochest33258染色结果与透射电镜观察结果显示,MLP降低了BMECs的凋亡率,同时保持了细胞膜和细胞结构完整性。3)试剂盒检测结果显示,MLP提高了BMECs中CAT、GSH-Px和SOD活性,同时降低了MDA含量。综上所述,MLP可有效减缓BMECs凋亡,提高其抗氧化能力。