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Article(id=1204800730801611272, tenantId=1146029695717560320, journalId=1192105938417971205, issueId=1204800727341310425, articleNumber=null, orderNo=null, doi=10.13343/j.cnki.wsxb.20250367, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=research-article, receivedDate=1746460800000, receivedDateStr=2025-05-06, revisedDate=null, revisedDateStr=null, acceptedDate=1749657600000, acceptedDateStr=2025-06-12, onlineDate=1765176478338, onlineDateStr=2025-12-08, pubDate=1764777600000, pubDateStr=2025-12-04, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1765176478338, onlineIssueDateStr=2025-12-08, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1765176478338, creator=13701087609, updateTime=1765176478338, updator=13701087609, issue=Issue{id=1204800727341310425, tenantId=1146029695717560320, journalId=1192105938417971205, year='2025', volume='65', issue='12', pageStart='5191', pageEnd='5649', issueExtLink='null', onlineDate='null', pubDate='null', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=null, createTime=1765176477513, creator=13701087609, updateTime=1765176611928, updator=13701087609, preIssue=null, nextIssue=null, ext={EN=IssueExt(id=1204801291189986067, tenantId=1146029695717560320, journalId=1192105938417971205, issueId=1204800727341310425, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1204801291189986068, tenantId=1146029695717560320, journalId=1192105938417971205, issueId=1204800727341310425, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null}, startPage=5352, endPage=5361, ext={EN=ArticleExt(id=1204800731703386645, articleId=1204800730801611272, tenantId=1146029695717560320, journalId=1192105938417971205, language=EN, title=Yeast dietary fiber regulates arsenic-induced apoptosis in yeast cells by modulating the antioxidant system, columnId=1192149543992045670, journalTitle=Acta Microbiologica Sinica, columnName=Research Article, runingTitle=null, highlight=null, articleAbstract=

Objective To investigate the effect of yeast dietary fiber (YDF) on arsenic-induced apoptosis in Saccharomyces cerevisiae and decipher the possible mechanism. Methods The relative survival rate, apoptosis, and antioxidant indicators were determined by the spread plate method, spectrophotometry, fluorescence microscopy, and RT-qPCR. Results The exposure to arsenic significantly decreased the relative survival rate, elevated the intracellular reactive oxygen species (ROS) and malondialdehyde (MDA) levels, and induced apoptosis. However, in the presence of YDF (0.5 mg/mL or 1.0 mg/mL) and arsenic, the arsenic-induced toxic effects were effectively attenuated, which was evidenced by increases in the relative survival rate, content of glutathione, activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPX), and relative expression of antioxidant enzyme genes (SOD1, CTA1, CTT1, and GPX2). Moreover, the treatment with both YDF and arsenic lowered the ROS and the MDA levels, significantly down-regulated the relative expression levels of pro-apoptotic genes (AIF1, NMAⅢ, and NUC1), and significantly reduced apoptotic cells compared with the treatment with arsenic alone. Conclusion YDF regulates the antioxidant system to attenuate the arsenic-induced cytotoxicity, thereby alleviating the arsenic-induced apoptosis.

, correspAuthors=Lihua WU, authorNote=null, correspAuthorsNote=
*E-mail:
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目的 以模式生物酿酒酵母为材料，研究酵母膳食纤维(yeast dietary fiber, YDF)在砷诱导酵母细胞凋亡过程中的缓解作用。 方法 运用平板涂布法、分光光度法、荧光显微技术以及逆转录实时定量聚合酶链式反应(quantitative reverse transcription polymerase chain reaction, RT-qPCR)法检测酵母细胞的相对存活率、细胞凋亡及抗氧化相关指标。 结果 砷可显著降低酵母细胞的相对存活率，提高胞内活性氧(reactive oxygen species, ROS)水平和丙二醛(malondialdehyde, MDA)含量，诱导细胞发生凋亡。当YDF (0.5 mg/mL或1.0 mg/mL)与砷共同作用时，酵母细胞的相对存活率、抗氧化分子谷胱甘肽(glutathione, GSH)含量、抗氧化酶[超氧化物歧化酶(superoxide dismutase, SOD)、过氧化氢酶(catalase, CAT)和谷胱甘肽过氧化物酶(glutathione peroxidase, GPX)]活性以及抗氧化酶基因(SOD1、GPX2、CTA1和CTT1)的相对表达量均高于砷单独处理组；同时，ROS水平和MDA含量下降，促凋亡基因(AIF1、NMAⅢ和NUC1)的相对表达量显著下调，发生凋亡的细胞数量显著减少。 结论 YDF能够通过调节抗氧化系统降低砷胁迫引发的细胞毒性，进而缓解砷诱导的细胞凋亡。

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Mitochondrial oxidative stress in a rat model of chronic intraocular pressure elevation and manganese superoxide dismutase gene therapy[D]. Changsha: Central South University, 2012 (in Chinese)., articleTitle=null, refAbstract=null)], funds=[Fund(id=1217784601666306405, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, awardId=21307087, language=EN, fundingSource=National Natural Science Foundation of China(21307087), fundOrder=null, country=null), Fund(id=1217784601804718444, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, awardId=21307087, language=CN, fundingSource=国家自然科学基金(21307087), fundOrder=null, country=null), Fund(id=1217784601922158964, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, awardId=2019L0820, language=EN, fundingSource=Scientific and Technological Innovation Program of Higher Education Institutions in Shanxi Province(2019L0820), fundOrder=null, country=null), Fund(id=1217784602018627960, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, awardId=2019L0820, language=CN, fundingSource=山西省高校科技创新项目(2019L0820), fundOrder=null, country=null), Fund(id=1217784602152845696, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, awardId=20251297, language=EN, fundingSource=Innovative Training Program for College Students of Shanxi Province(20251297), fundOrder=null, country=null), Fund(id=1217784602270286218, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, awardId=20251297, language=CN, fundingSource=山西省大学生创新项目(20251297), fundOrder=null, country=null)], companyList=[AuthorCompany(id=1217784595630703513, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, xref=null, ext=[AuthorCompanyExt(id=1217784595655869339, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, companyId=1217784595630703513, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=College of Biological Sciences and Technology, Taiyuan Normal University, Jinzhong, Shanxi, China), AuthorCompanyExt(id=1217784595668452252, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, companyId=1217784595630703513, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=太原师范学院 生物科学与技术学院，山西 晋中)])], figs=[ArticleFig(id=1217784599690789082, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, language=EN, label=Figure 1, caption=Effect of YDF on relative survival rate in yeast cells under arsenic stress. Bars with different lowercase letters indicate that the same index is significantly different (P<0.05)., figureFileSmall=az9v/dmCPp1CR5O3OWcaWA==, figureFileBig=XRLU3AN7ybLX9PQzXxfbCQ==, tableContent=null), ArticleFig(id=1217784599829201124, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, language=CN, label=图1, caption=YDF对砷胁迫下酵母细胞相对存活率的影响, figureFileSmall=az9v/dmCPp1CR5O3OWcaWA==, figureFileBig=XRLU3AN7ybLX9PQzXxfbCQ==, tableContent=null), ArticleFig(id=1217784599976001776, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, language=EN, label=Figure 2, caption=Effect of YDF on sodium arsenite-induced oxidative damage in yeast cells. A: The level of intracellular ROS; B: MDA content. Bars with different lowercase letters indicate that the same index is significantly different (P<0.05)., figureFileSmall=xP806O28usZT+KLBgeQrSg==, figureFileBig=XvjUZC+Jeayc6qniPLxwQQ==, tableContent=null), ArticleFig(id=1217784600135385335, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, language=CN, label=图2, caption=YDF对砷引起酵母细胞氧化损伤的影响, figureFileSmall=xP806O28usZT+KLBgeQrSg==, figureFileBig=XvjUZC+Jeayc6qniPLxwQQ==, tableContent=null), ArticleFig(id=1217784600257020158, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, language=EN, label=Figure 3, caption=Effect of YDF on antioxidant enzymes and GSH content in yeast cells under arsenic stress. A: SOD activity; B: CAT activity; C: GPX activity; D: GSH content. Bars with different lowercase letters indicate that the same index is significantly different (P<0.05)., figureFileSmall=DrwxHU8CACrjKWqsw/yskQ==, figureFileBig=v5IAE8zUIS53ShYBlrD8MA==, tableContent=null), ArticleFig(id=1217784600382849290, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, language=CN, label=图3, caption=YDF对砷胁迫下酵母细胞抗氧化酶活性和GSH含量的影响, figureFileSmall=DrwxHU8CACrjKWqsw/yskQ==, figureFileBig=v5IAE8zUIS53ShYBlrD8MA==, tableContent=null), ArticleFig(id=1217784600479318289, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, language=EN, label=Figure 4, caption=Effect of YDF on relative expression of antioxidant enzyme genes in yeast cells under arsenic stress. A: SOD1; B: GPX2; C: CTA1; D: CTT1. Bars with different lowercase letters indicate that the same index is significantly different (P<0.05)., figureFileSmall=QFiVMvkFcMYtNN8c27qZVQ==, figureFileBig=VO7iJh10r0wCXjg5xMVSAQ==, tableContent=null), ArticleFig(id=1217784600584175899, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, language=CN, label=图4, caption=YDF对砷胁迫下酵母细胞抗氧化酶相关基因表达的影响, figureFileSmall=QFiVMvkFcMYtNN8c27qZVQ==, figureFileBig=VO7iJh10r0wCXjg5xMVSAQ==, tableContent=null), ArticleFig(id=1217784600672256291, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, language=EN, label=Figure 5, caption=Effect of YDF on apoptosis in yeast cells under arsenic stress., figureFileSmall=C8x2oKHN7Bfr86oUOcLGVw==, figureFileBig=GTfIF/vaebZ+6Il3ypD1Gg==, tableContent=null), ArticleFig(id=1217784600802279725, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, language=CN, label=图5, caption=YDF对砷胁迫下酵母细胞凋亡的影响, figureFileSmall=C8x2oKHN7Bfr86oUOcLGVw==, figureFileBig=GTfIF/vaebZ+6Il3ypD1Gg==, tableContent=null), ArticleFig(id=1217784600890360118, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, language=EN, label=Figure 6, caption=Effect of YDF on relative expression of pro-apoptotic genes in yeast cells under arsenic stress. A: AIF1; B: NMAⅢ; C: NUC1. Bars with different lowercase letters indicate that the same index is significantly different (P<0.05)., figureFileSmall=dGDceJWctexVkhSlg4WFWg==, figureFileBig=uR18VcMCuyN1763tUBzx3g==, tableContent=null), ArticleFig(id=1217784601041355068, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, language=CN, label=图6, caption=YDF对砷胁迫下酵母细胞促凋亡基因表达的影响, figureFileSmall=dGDceJWctexVkhSlg4WFWg==, figureFileBig=uR18VcMCuyN1763tUBzx3g==, tableContent=null), ArticleFig(id=1217784601158795593, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, language=EN, label=Table 1, caption=

Primers for RT-qPCR

, figureFileSmall=null, figureFileBig=null, tableContent=
Gene namePrimer sequences (5′→3′)
ACT1

F: TACTCTTTCTCCACCACTGCTGA

R: CTTGACCATCTGGAAGTTCGTAG

SOD1

F: CGAGCCAACCACTGTCTCTT

R: ACGTTACCCATGTCACCGAC

GPX2

F: TAATGTTGCCTCCAAGTGCG

R: GTTCATCAGAGCCCGGTTCC

CTA1

F: GTTCTGGTGCCTTCGGCTAT

R: GAATTTGGTGGCAAACCCCC

CTT1

F: CATGCCAAAGGTGGTGGTTG

R: CAGTCATGGTTCCCCCACTC

AIF1

F: GCTGGTGTGTTTGGTGTGTC

R: TAACGGCAGGATCGACTTGG

NMAⅢ

F: AGTTTGGCTAAGGTCGGCTC

R: TTGAACCGCCAGAAGCAGAT

NUC1

F: AGAGGATATTGCTGTCGCGG

R: GTTTGATCGCCGCGTTAGAG

), ArticleFig(id=1217784601288819028, tenantId=1146029695717560320, journalId=1192105938417971205, articleId=1204800730801611272, language=CN, label=表1, caption=

RT-qPCR引物

, figureFileSmall=null, figureFileBig=null, tableContent=
Gene namePrimer sequences (5′→3′)
ACT1

F: TACTCTTTCTCCACCACTGCTGA

R: CTTGACCATCTGGAAGTTCGTAG

SOD1

F: CGAGCCAACCACTGTCTCTT

R: ACGTTACCCATGTCACCGAC

GPX2

F: TAATGTTGCCTCCAAGTGCG

R: GTTCATCAGAGCCCGGTTCC

CTA1

F: GTTCTGGTGCCTTCGGCTAT

R: GAATTTGGTGGCAAACCCCC

CTT1

F: CATGCCAAAGGTGGTGGTTG

R: CAGTCATGGTTCCCCCACTC

AIF1

F: GCTGGTGTGTTTGGTGTGTC

R: TAACGGCAGGATCGACTTGG

NMAⅢ

F: AGTTTGGCTAAGGTCGGCTC

R: TTGAACCGCCAGAAGCAGAT

NUC1

F: AGAGGATATTGCTGTCGCGG

R: GTTTGATCGCCGCGTTAGAG

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酵母膳食纤维通过调节酵母抗氧化系统缓解砷引起的细胞凋亡
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吴丽华 * , 葛文君 , 姚夏 , 董娓伶 , 赵文婧
微生物学报 | 研究报告 2025,65(12): 5352-5361
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微生物学报 | 研究报告 2025, 65(12): 5352-5361
酵母膳食纤维通过调节酵母抗氧化系统缓解砷引起的细胞凋亡
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吴丽华* , 葛文君, 姚夏, 董娓伶, 赵文婧
作者信息
  • 太原师范学院 生物科学与技术学院，山西 晋中
Yeast dietary fiber regulates arsenic-induced apoptosis in yeast cells by modulating the antioxidant system
Lihua WU* , Wenjun GE, Xia YAO, Weiling DONG, Wenjing ZHAO
Affiliations
  • College of Biological Sciences and Technology, Taiyuan Normal University, Jinzhong, Shanxi, China
出版时间: 2025-12-04 doi: 10.13343/j.cnki.wsxb.20250367
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目的 以模式生物酿酒酵母为材料，研究酵母膳食纤维(yeast dietary fiber, YDF)在砷诱导酵母细胞凋亡过程中的缓解作用。 方法 运用平板涂布法、分光光度法、荧光显微技术以及逆转录实时定量聚合酶链式反应(quantitative reverse transcription polymerase chain reaction, RT-qPCR)法检测酵母细胞的相对存活率、细胞凋亡及抗氧化相关指标。 结果 砷可显著降低酵母细胞的相对存活率，提高胞内活性氧(reactive oxygen species, ROS)水平和丙二醛(malondialdehyde, MDA)含量，诱导细胞发生凋亡。当YDF (0.5 mg/mL或1.0 mg/mL)与砷共同作用时，酵母细胞的相对存活率、抗氧化分子谷胱甘肽(glutathione, GSH)含量、抗氧化酶[超氧化物歧化酶(superoxide dismutase, SOD)、过氧化氢酶(catalase, CAT)和谷胱甘肽过氧化物酶(glutathione peroxidase, GPX)]活性以及抗氧化酶基因(SOD1、GPX2、CTA1和CTT1)的相对表达量均高于砷单独处理组；同时，ROS水平和MDA含量下降，促凋亡基因(AIF1、NMAⅢ和NUC1)的相对表达量显著下调，发生凋亡的细胞数量显著减少。 结论 YDF能够通过调节抗氧化系统降低砷胁迫引发的细胞毒性，进而缓解砷诱导的细胞凋亡。

酵母膳食纤维  /  酿酒酵母  /  亚砷酸钠  /  凋亡  /  抗氧化系统

Objective To investigate the effect of yeast dietary fiber (YDF) on arsenic-induced apoptosis in Saccharomyces cerevisiae and decipher the possible mechanism. Methods The relative survival rate, apoptosis, and antioxidant indicators were determined by the spread plate method, spectrophotometry, fluorescence microscopy, and RT-qPCR. Results The exposure to arsenic significantly decreased the relative survival rate, elevated the intracellular reactive oxygen species (ROS) and malondialdehyde (MDA) levels, and induced apoptosis. However, in the presence of YDF (0.5 mg/mL or 1.0 mg/mL) and arsenic, the arsenic-induced toxic effects were effectively attenuated, which was evidenced by increases in the relative survival rate, content of glutathione, activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPX), and relative expression of antioxidant enzyme genes (SOD1, CTA1, CTT1, and GPX2). Moreover, the treatment with both YDF and arsenic lowered the ROS and the MDA levels, significantly down-regulated the relative expression levels of pro-apoptotic genes (AIF1, NMAⅢ, and NUC1), and significantly reduced apoptotic cells compared with the treatment with arsenic alone. Conclusion YDF regulates the antioxidant system to attenuate the arsenic-induced cytotoxicity, thereby alleviating the arsenic-induced apoptosis.

yeast dietary fiber  /  Saccharomyces cerevisiae  /  sodium arsenite  /  apoptosis  /  antioxidant system
吴丽华, 葛文君, 姚夏, 董娓伶, 赵文婧. 酵母膳食纤维通过调节酵母抗氧化系统缓解砷引起的细胞凋亡. 微生物学报, 2025 , 65 (12) : 5352 -5361 . DOI: 10.13343/j.cnki.wsxb.20250367
Lihua WU, Wenjun GE, Xia YAO, Weiling DONG, Wenjing ZHAO. Yeast dietary fiber regulates arsenic-induced apoptosis in yeast cells by modulating the antioxidant system[J]. Acta Microbiologica Sinica, 2025 , 65 (12) : 5352 -5361 . DOI: 10.13343/j.cnki.wsxb.20250367
正文
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膳食纤维(dietary fiber, DF)，是一类在人体小肠中无法被完全水解的碳水化合物，具有降血糖、降血脂、抗肿瘤等多种功效[1]。Tan等[2]调查发现，增加DF的摄入量可降低肿瘤幸存者的死亡风险。Román-Ochoa等[3]研究发现，DF能够减轻砷暴露对肠道微生物菌群的不利影响。酵母膳食纤维(yeast dietary fiber, YDF)是从酵母细胞壁中提取的主要由甘露聚糖和β-葡聚糖组成的功能性物质，具有抑菌、抗氧化及提高机体免疫力的功效[4-5]。实验表明，酵母甘露聚糖可提高断奶仔猪的抗氧化能力，抵御大肠杆菌的侵袭[6]；酵母β-葡聚糖具有抗病毒作用，在一定程度上可控制病毒传播[7]；酵母甘露聚糖和β-葡聚糖协同作用能够显著增强免疫细胞的吞噬活性，缓解脂多糖(lipopolysaccharide, LPS)诱导的细胞炎症反应[8]。廖凯威等[5]在蛋鸡日粮中添加YDF后蛋鸡的生产性能和免疫力显著提高，鸡蛋品质明显改善。根据《GB 2760—2014食品安全国家标准食品添加剂使用标准》[9]，酵母及酵母类制品可直接食用，且无添加量限制。因此，探究YDF的抗毒性作用对保护人群健康具有重要意义。
砷是一种广泛存在于自然界中的环境污染物，其可穿过膜脂双分子层进入细胞，通过抑制抗氧化酶活性使超氧阴离子(O2-)、羟自由基(∙OH)和过氧化氢(H2O2)等活性氧自由基异常积累，引发细胞氧化应激，造成一系列损伤：(1) 改变DNA修复机制，导致DNA损伤；(2) 使蛋白折叠错误并聚集，致使酶蛋白结构变性、功能丧失；(3) 攻击细胞膜脂类物质，造成脂质过氧化，最终引起细胞损伤[10-11]。现有研究显示，高浓度砷可改变动物肠道微生物菌群[12]，引发人体皮肤、肾脏、肝脏等器官组织发生病变，甚至导致癌症发生[13-15]。吴丽华等[16-17]的研究结果显示，砷可引起酵母发生细胞凋亡和氧化应激。然而，YDF是否可缓解砷引起的细胞凋亡尚不明确。因此，本研究以模式生物酿酒酵母为材料，研究YDF对砷引起酵母凋亡的影响，以期阐明部分YDF缓解砷毒性的作用机制。
1 材料与方法
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1.1 菌株和培养基
供试菌株为酿酒酵母(BY4741)，保存于太原师范学院微生物实验室；YDF购自安琪酵母股份有限公司。
YPD液体培养基(g/L)：酵母粉10.0，蛋白胨20.0，葡萄糖20.0，pH自然。
YPD固体培养基(g/L)：酵母粉10.0，蛋白胨20.0，葡萄糖20.0，琼脂粉15.0-20.0，pH自然。
1.2 酵母细胞的培养与处理
在YPD液体培养基中接种酵母单菌落，置于恒温振荡培养箱中28 ℃、180 r/min培养至对数期(OD600为0.8-1.2)后进行药物处理，并使各处理组初始OD600一致。处理组包括对照组、YDF组(0.5、1.0、5.0 mg/mL)、砷处理组(1 mmol/L)和YDF+砷处理组(1 mmol/L)。处理12 h后进行各指标检测，每个处理设置3次重复。
1.3 细胞相对存活率测定
将数量相等的对数期酵母细胞均匀涂布至各处理组平板中，28 ℃恒温培养48 h后进行菌落计数，并计算相对存活率，如公式(1)所示，该结果用于筛选YDF的实验浓度，其中YDF浓度范围为0.1-15.0 mg/mL。
相对存活率=(处理组菌落数/对照组菌落数)×100%
1.4 酵母胞内活性氧水平、丙二醛和谷胱甘肽含量的测定
胞内活性氧(reactive oxygen species, ROS)检测参考文献[17]，装载探针后在显微镜下观察并拍照；丙二醛(malondialdehyde, MDA)含量检测参照文献[18]进行；谷胱甘肽(glutathione, GSH)含量检测参照GSH测定试剂盒(南京建成生物工程研究所)说明书进行。
1.5 酵母细胞抗氧化酶活性检测
收集酵母细胞，使用新鲜配制的破壁酶液破壁30 min后，参照文献[19]方法提取粗酶液，根据相关试剂盒(南京建成生物工程研究所)说明书分别检测酵母细胞抗氧化酶超氧化物歧化酶(superoxide dismutase, SOD)、过氧化氢酶(catalase, CAT)和谷胱甘肽过氧化物酶(glutathione peroxidase, GPX)的活性。
1.6 细胞凋亡检测
将酵母细胞用预冷的PBS洗涤后去壁，使用细胞凋亡检测试剂盒(北京博奥森生物技术有限公司)避光孵育20 min后，取适量细胞在荧光显微镜下观察并拍照。
1.7 RT-qPCR检测基因表达水平
参照吴丽华等[20]建立的方法提取酵母细胞RNA，反转录为cDNA后使用RT-qPCR技术检测抗氧化酶相关基因和酵母细胞促凋亡基因的表达情况，然后采用2-∆∆Ct法计算基因的相对表达量。其中，ACT1为本研究的内参基因，所用引物的序列见表1。
1.8 数据统计分析
实验数据使用SPSS 26.0软件统计分析，图中不同小写英文字母代表不同处理组间具有显著性差异(P<0.05)。数据以平均值±标准差(mean±SD)表示，并采用Origin 2021软件制图。
2 结果与分析
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2.1 YDF对砷胁迫下酵母细胞相对存活率的影响
从图1可知，砷单独处理组中酵母细胞的相对存活率为对照组的15.37%。在砷+YDF (0.5-15.0 mg/mL)组中细胞相对存活率均高于砷单独处理组，当5.0 mg/mL或10.0 mg/mL的YDF与砷共同作用时细胞相对存活率显著高于砷单独处理组。结果表明，砷处理可引起酵母细胞生长抑制和细胞死亡，而一定浓度的YDF可缓解砷引起的酵母细胞毒性。由于5.0 mg/mL的YDF即可显著缓解砷对酵母细胞的毒性作用，因此本研究选取高(5.0 mg/mL)、中(1.0 mg/mL)和低(0.5 mg/mL) 3个浓度的YDF用于下一步研究YDF对砷毒性的缓解效应。
2.2 YDF对砷胁迫下酵母细胞氧化损伤的影响
ROS水平和MDA含量是衡量细胞氧化应激的重要指标。由图2可知，经砷处理后酵母细胞中代表ROS的绿色荧光强度和MDA含量均显著高于对照组；添加YDF后细胞荧光强度和MDA含量均低于砷单独处理组。结果表明，砷可引起酵母细胞内ROS水平和MDA含量升高，而YDF在一定程度上可缓解酵母细胞中砷引起的氧化损伤，减轻砷对酵母细胞的毒性。
2.3 YDF对砷胁迫下酵母细胞抗氧化酶活性及GSH含量的影响
如图3所示，砷可显著降低酵母细胞的抗氧化酶SOD、CAT和GPX的活性及胞内抗氧化分子GSH的含量。添加0.5 mg/mL或1.0 mg/mL的YDF后，抗氧化酶活性和GSH含量均显著高于砷单独处理组，其中1.0 mg/mL YDF缓解组中CAT活性是砷单独处理组的3.61倍。结果表明，砷可抑制酵母细胞抗氧化酶活性，减少抗氧化分子含量，而YDF可显著拮抗砷引起的酵母细胞抗氧化能力的降低，从而缓解由砷引起的氧化损伤。
2.4 YDF对砷胁迫下酵母细胞抗氧化酶基因表达的影响
从图4可以看出，砷可上调抗氧化酶相关基因(除SOD1外)的相对表达量，而YDF的添加可改变所检测基因的表达量，呈现出先升后降的趋势；当浓度为1.0 mg/mL的YDF与砷共同作用时，所检测抗氧化基因相对表达量达到峰值，其中CTA1的相对表达量为砷单独处理组的3.13倍。结果表明，YDF能够通过调节砷胁迫下酵母细胞抗氧化酶基因表达上调提高抗氧化酶活性，进而提高酵母砷耐受性。
2.5 YDF对砷胁迫下酵母细胞凋亡的影响
从图5可以看出，酵母细胞经砷处理后荧光信号显著增强，出现早期凋亡(绿色荧光)和晚期凋亡细胞(红色荧光)。当YDF与砷共同作用时出现绿色荧光和红色荧光的细胞均显著少于砷单独处理组，当YDF浓度为1.0 mg/mL时凋亡细胞比例最低。结果表明，砷可引起酵母细胞凋亡，而YDF可显著缓解砷引起的细胞凋亡。
2.6 YDF对砷胁迫下酵母细胞促凋亡基因表达的影响
由图6可以看出，砷胁迫下促凋亡基因(AIF1、NMAⅢ和NUC1)相对表达水平均显著高于对照组，YDF与砷共同作用时促凋亡基因的相对表达量随YDF浓度的升高呈现出先降后升的趋势，与抗氧化酶活性及相关基因表达趋势相反(图3和图4)。结果表明，YDF可通过抗氧化系统调节砷胁迫下酵母细胞促凋亡基因表达。
3 讨论与结论
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凋亡是机体中由基因控制的细胞主动死亡的生命现象，在维持生物体稳态、发育和疾病防治方面具有重要意义[21]。自1977年Madeo等[21]在酵母细胞中发现细胞凋亡以来，众多学者发现多种外源性刺激均可诱导酵母细胞凋亡[22-24]。酵母细胞受到凋亡刺激后，AIF1和NUC1从线粒体定位到细胞核中，并引发凋亡应答[25-26]；而ΔNMAⅢ经高温和氧化胁迫处理均不会发生细胞凋亡[27]，这表明AIF1、NUC1和NMAⅢ在酵母细胞凋亡过程中发挥着关键作用。本研究结果显示，砷可诱导酵母细胞凋亡，当0.5-1.0 mg/mL的YDF与砷共同处理时，发生细胞凋亡的酵母细胞数量以及促凋亡基因AIF1、NMAⅢ和NUC1的相对表达量均显著低于砷单独处理组，说明YDF可抑制促凋亡基因表达，提高细胞存活率，减少细胞凋亡的发生。
砷化物是一类毒性较强的环境污染物。氧化应激是砷中毒的重要原因之一，砷可抑制生物体内关键酶系统，诱导ROS过量积累，引发机体氧化损伤，最终导致生物体死亡[28]。本研究结果显示，砷可引起酵母细胞氧化应激反应，使细胞内ROS和MDA含量升高，最终导致细胞凋亡，此结果与其他学者在动物细胞中的研究结果一致[29-30]。酵母细胞具有易培养、代时短、遗传背景简单，且与人类很多疾病相关基因高度保守等优点，因此可作为研究砷毒性的模式生物[16-17]。本研究结果显示，YDF可通过提高酵母抗氧化酶基因表达增强其抗氧化作用，降低砷暴露引起的酵母细胞氧化损伤，从而增强酵母对砷胁迫的耐受性，此结果为进一步探究YDF的食用功效及其在减轻重金属毒性方面的潜在应用提供了实验依据。
酵母细胞中的氧化防御系统主要包括非酶促系统的谷胱甘肽(GSH)和酶促系统中的超氧化物歧化酶(SOD1和SOD2)、过氧化氢酶(CTT1和CTA1)和谷胱甘肽过氧化物酶(GPXs)。其中，SOD可将O2-催化形成低毒的H2O2和O2；CTT1、CTA1和GPXs可将H2O2进一步分解生成无害的H2O和O2[31]。Rona等[32]发现在酵母细胞中CTT1基因的过量表达可回补SOD1基因的缺失，使胞内MDA含量减少，从而降低细胞凋亡率；吴丽华等[33]在前期的研究中也发现，SOD1基因缺失后砷诱导的酵母细胞凋亡率显著高于野生株，表明抗氧化系统可在一定程度上影响酵母细胞凋亡的发生。本研究中砷可抑制抗氧化酶SOD、CAT和GPX的活性，降低胞内GSH含量，但对基因SOD1、CTT1、CTA1和GPX2的表达却有促进作用，这有可能是因为砷胁迫可引起酵母细胞发生氧化应激，而氧化应激时SOD、CAT和GSH蛋白翻译受阻，导致酶活性下降[34]。然而，在砷处理组中添加0.5-1.0 mg/mL的YDF后，抗氧化酶活性和GSH含量均显著提高，且相关基因的相对表达量呈现出一致的趋势，说明YDF可在酵母细胞中缓解砷引起的氧化损伤，进而调控细胞凋亡。
综上所述，在砷处理组中添加YDF后，酵母的抗氧化酶活性及相关基因表达显著高于砷单独处理组，ROS水平、MDA含量和发生细胞凋亡的细胞数量均显著下降。结果表明，YDF可通过提高酵母细胞的抗氧化系统调控砷诱导的酵母细胞凋亡。
作者贡献声明
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吴丽华：获取基金、项目管理、实验指导、论文撰写和修改；葛文君：实验操作、数据收集和处理；姚夏：实验操作和数据收集；董娓伶：协助实验操作；赵文婧：参与实验讨论。
作者利益冲突公开声明
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作者声明不存在任何可能会影响本文所报告工作的已知经济利益或个人关系。
基金
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  • 国家自然科学基金(21307087)
  • 山西省高校科技创新项目(2019L0820)
  • 山西省大学生创新项目(20251297)
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2025年第65卷第12期
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doi: 10.13343/j.cnki.wsxb.20250367
  • 接收时间：2025-05-06
  • 首发时间：2025-12-08
  • 出版时间：2025-12-04
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  • 收稿日期：2025-05-06
  • 录用日期：2025-06-12
基金
National Natural Science Foundation of China(21307087)
国家自然科学基金(21307087)
Scientific and Technological Innovation Program of Higher Education Institutions in Shanxi Province(2019L0820)
山西省高校科技创新项目(2019L0820)
Innovative Training Program for College Students of Shanxi Province(20251297)
山西省大学生创新项目(20251297)
作者信息
    太原师范学院 生物科学与技术学院，山西 晋中

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2种不同金属材料的力学参数
科
Family		 属数
Number of
genus		 种数
Number of
species		 占总种数比例
Percentage of
total species (%)		 属
Genus		 种数
Number of
species		 占总种数比例
Percentage of total
species (%)
鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
小菇科 Mycenaceae 2 12 5.74 丝盖伞属 Inocybe 5 2.39
多孔菌科 Polyporaceae 8 14 6.70 蜡蘑属 Laccaria 5 2.39
红菇科 Russulaceae 3 23 11.00 小皮伞属 Marasmius 6 2.87
小菇属 Mycena 11 5.26
光柄菇属 Pluteus 5 2.39
红菇属 Russula 17 8.13
栓菌属 Trametes 5 2.39
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