High-mobility group box 1 protein and its role in severe acute pancreatitis

The high mobility group box 1(HMGB1),which belongs to the subfamily of HMG-1/-2,is a highly conserved single peptide chain consisting of 215 amino acid residues with a molecular weight of approximately 24894 Da.HMGB1 is a ubiquitous nuclear protein in mammals and plays a vital role in inflammatory diseases.Acute pancreatitis is one of the most common causes of acute abdominal pain with a poor prognosis.Acute pancreatitis is an acute inflammatory process of the pancreas(duration of less than six months),for which the severe form is called severe acute pancreatitis(SAP).More and more studies have shown that HMGB1 has a bidirectional effect in the pathogenesis of SAP.Extracellular HMGB1 can aggravate the pancreatic inflammatory process,whereas intracellular HMGB1 has a protective effect against pancreatitis.The mechanism of HMGB1 is multiple,mainly through the nuclear factor-κB pathway.Receptors for advanced glycation endproducts and toll-like receptors(TLR),especially TLR-2 and TLR-4,are two major types of receptors mediating the inflammatory process triggered by HMGB1 and may be also the main mediators in the pathogenesis of SAP.HMGB1 inhibitors,such as ethyl pyruvate,pyrrolidine dithiocarbamate and Scolopendra subspinipes mutilans,can decrease the level of extracellular HMGB1 and are the promising targets in the treatment of SAP.

Scolopendra subspinipes mutilans protected the ceruleininduced acute pancreatitis by inhibiting high-mobility group box protein-1

AIM:To evaluate the inhibitory effects of Scolopendra subspinipes mutilans(SSM) on cerulein-induced acute pancreatitis(AP) in a mouse model.METHODS:SSM water extract(0.1,0.5,or 1 g/kg) was administrated intraperitoneally 1 h prior to the first injection of cerulein.Once AP developed,the stable cholecystokinin analogue,cerulein was injected hourly,over a 6 h period.Blood samples were taken 6 h later to determine serum amylase,lipase,and cytokine levels.The pancreas and lungs were rapidly removed for morphological examination,myeloperoxidase assay,and real-time reverse transcription polymerase chain reaction.To specify the role of SSM in pancreatitis,the pancreatic acinar cells were isolated using collagenase method.Then the cells were pre-treated with SSM,then stimulated with cerulein.The cell viability,cytokine productions and high-mobility group box protein-1(HMGB-1) were measured.Furthermore,the regulating mechanisms of SSM action were evaluated.RESULTS:The administration of SSM significantly attenuated the severity of pancreatitis and pancreatitis associated lung injury,as was shown by the reduction in pancreatic edema,neutrophil infiltration,vacuolization and necrosis.SSM treatment also reduced pancreatic weight/body weight ratio,serum amylase,lipase and cytokine levels,and mRNA expression of multiple inflammatory mediators such as tumor necrosis factor-α and interleukin-1β.In addition,treatment with SSM inhibited HMGB-1 expression in the pancreas during AP.In accordance with in vivo data,SSM inhibited the cerulein-induced acinar cell death,cytokine,and HMGB-1 release.SSM also inhibited the activation of c-Jun NH2-terminal kinase,p38 and nuclear factor(NF)-κB.CONCLUSION:These results suggest that SSM plays a protective role during the development of AP and pancreatitis associated lung injury via deactivating c-Jun NH2-terminal kinase,p38 and NF-κB.

Suppressing high mobility group box-1 release alleviates morphine tolerance via the adenosine5'-monophosphate-activated protein kinase/heme oxygenase-1 pathway

Opioids,such as morphine,are the most potent drugs used to treat pain.Long-term use results in high tolerance to morphine.High mobility group box-1(HMGB1) has been shown to participate in neuropathic or inflammatory pain,but its role in morphine tolerance is unclear.In this study,we established rat and mouse models of morphine tolerance by intrathecal injection of morphine for 7 consecutive days.We found that morphine induced rat spinal cord neurons to release a large amount of HMGB1.HMGB1 regulated nuclear factor κB p65 phosphorylation and interleukin-1β production by increasing Toll-like receptor 4receptor expression in microglia,thereby inducing morphine tolerance.Glycyrrhizin,an HMGB1 inhibito r,markedly attenuated chronic morphine tole rance in the mouse model.Finally,compound C(adenosine 5’-monophosphate-activated protein kinase inhibitor) and zinc protoporphyrin(heme oxygenase-1 inhibitor)alleviated the morphine-induced release of HMGB1 and reduced nuclear factor κB p65 phosphorylation and interleukin-1β production in a mouse model of morphine tolerance and an SH-SY5Y cell model of morphine tole rance,and alleviated morphine tolerance in the mouse model.These findings suggest that morphine induces HMGB1 release via the adenosine 5’-monophosphate-activated protein kinase/heme oxygenase-1 signaling pathway,and that inhibiting this signaling pathway can effectively reduce morphine tole rance.

High mobility group box-1 protein inhibits regulatory T cell immune activity in liver failure in patients with chronic hepatitis B

BACKGROUND: Liver failure in chronic hepatitis B (CHB) patients is a severe, life-threatening condition. Intestinal endotoxemia plays a significant role in the progress to liver failure. High mobility group box-1 (HMGB1) protein is involved in the process of endotoxemia. Regulatory T (Treg) cells maintain immune tolerance and contribute to the immunological hyporesponsiveness against HBV infection. However, the roles of HMGB1 and Treg cells in the pathogenesis of liver failure in CHB patients, and whether HMGB1 affects the immune activity of Treg cells are poorly known at present, and so were explored in this study. METHODS: The levels of HMGB1 expression were detected by ELISA, real-time RT-PCR, and Western blotting, and the percentage of CD4(+)CD25(+)CD127(low) Treg cells among CD4(+) cells was detected by flow cytometry in liver failure patients with chronic HBV infection, CHB patients, and healthy controls. Then, CD4(+)CD25(+)CD127(low) Treg cells isolated from the peripheral blood mononuclear cells from CHB patients were stimulated with HMGB1 at different concentrations or at various intervals. The effect of HMGB1 on the immune activity of Treg cells was assessed by a suppression assay of the allogeneic mixed lymphocyte response. The levels of forkhead box P3 (Foxp3) expression in Treg cells treated with HMGB1 were detected by RT-PCR and Western blotting. RESULTS: A higher level of HMGB1 expression and a lower percentage of Treg cells within the population of CIA(+) cells were found in liver failure patients than in CHB patients (82.6+/-20.1 mu g/L vs. 34.2+/-13.7 mu g/L; 4.55+/-1.34% vs. 9.52+/-3.89%, respectively). The immune activity of Treg cells was significantly weakened and the levels of Foxp3 expression were reduced in a dose- or time-dependent manner when Treg cells were stimulated with HMGB1 in vitro. CONCLUSIONS: The high level of HMGB1 and the low percentage of Treg cells play an important role in the pathogenesis of liver failure in patients with chronic HBV infection. Moreover, HMGB1 can weaken the immune activity of Treg cells. It is suggested that effectively inhibiting HMGB1 expression could be a feasible way to treat liver failure by suppressing endotoxemia and enhancing Treg cell activity.

High mobility group box 1 protein (HMGB1) as an immune-modulating factor for polarization of human T lymphocytes

Objective This study was performed to investigate the effect of high mobility group box-1 protein (HMGB1) on immune function of human T lymphocytes in vitro and explore its potential role in cell-mediated immune dysfunction. Methods Fresh blood was obtained from healthy adult volunteers and peripheral blood mononuclear cells (PBMCs) were isolated, then rhHMGB1 was added to PBMCs. Four-color flow cytometric (FCM) analysis was used for the measurement of intracellular cytokine including interleukin IL-4 and interferon IFN- ? ELISA kits were employed for the determination of IL-2 and sIL-2R protein levels in cell culture supernatants. Results (1) Different stimulating time and dosage of rhHMGB1 did not alter the number of IFN- αpositive cells (Th1). rhHMGB1 stimulation provoked a dose-dependent and time-dependent increase in Th2 subset and decrease in ratio of Th1 to Th2. (2) Compared with the untreated cells, when the cells were coincubated with rhHMGB1 (10-100ng/ml) for 12 hrs, protein release of IL-2 and sIL-2R were significantly up-regulated. At 48 hrs, in contrast, protein production was relatively lower in cells after exposure to 100-1000 ng/ ml rhHMGB1. Conclusions These findings demonstrated that HMGB1 has a dual influence on immune functions of human T lymphocytes.

Inhibition of LOX-1 alleviates the proinflammatory effects of high-mobility group box 1 in Aspergillus fumigatus keratitis

AIM: To investigate the inflammatory amplification effect of high-mobility group box 1(HMGB1) in Aspergillus fumigatus(A. fumigatus) keratitis and the relationship between lectin-like oxidized low-density lipoprotein receptor 1(LOX-1) and HMGB1 in keratitis immune responses.METHODS: Phosphate buffer saline(PBS), and Boxb were injected into BALB/c mice subconjunctivally before the corneas were infected with A. fumigatus. RAW264.7 macrophages and neutrophils were pretreated with PBS and Boxb to determine the HMGB1 inflammatory amplification effects. Abdominal cavity extracted macrophages were pretreated with Boxb and Poly(I)(a LOX-1 inhibitor) before A. fumigatus hyphae stimulation to prove the the relationship between the two molecules. LOX-1, interleukin-1β(IL-1β), tumor necrosis factor-α(TNF-α), macrophage inflammatory protein-2(MIP-2) and IL-10 were assessed by polymerase chain reaction and Western blot.RESULTS: Pretreatment with Boxb exacerbated corneal inflammation. In macrophages and neutrophils, A. fumigatus induced LOX-1, IL-1β, TNF-α and MIP-2 expression in Boxb group was higher than those in PBS group. Poly(I) treatments before infection alleviated the proinflammatory effects of Boxb in abdominal cavity extracted macrophages. Pretreatment with Boxb did not influence Dectin-1 mRNA levels in macrophages and neutrophils.CONCLUSION: In fungal keratitis, HMGB1 is a proinflammatory factor in the first line of immune response. HMGB1 mainly stimulates neutrophils and macrophages to produce inflammatory cytokines and chemokines during the immune response. LOX-1 participates in HMGB1 induced inflammatory exacerbation in A. fumigatus keratitis.

Novel insights for high mobility group box 1 proteinmediated cellular immune response in sepsis:A systemic review

BACKGROUND:High mobility group box 1 protein(HMGB1) is a highly conserved,ubiquitous protein in the nuclei and cytoplasm of nearly all cell types.HMGB1 is secreted into the extracellular milieu and acts as a proinflammatory cytokine.In this article we reviewed briefly the cellular immune response mediated by HMGB1 in inflammation and sepsis.METHODS:This systemic review is mainly based on our own work and other related reports.RESULTS:HMGB1 can actively affect the immune functions of many types of cells including T lymphocytes,regulatory T cells(Tregs),dendritic cells(DCs),macrophages,and natural killer cells(NK cells).Various cellular responses can be mediated by HMGB1 which binds to cell-surface receptors[e.g.,the receptor for advanced glycation end products(RAGE),Toll-like receptor(TLR)2,and TLR4].Anti-HMGB1 treatment,such as anti-HMGB1 polyclonal or monoclonal antibodies,inhibitors(e.g.,ethyl pyruvate) and antagonists(e.g.,A box),can protect against sepsis lethality and give a wider window for the treatment opportunity.CONCLUSION:HMGB1 is an attractive target for the development of new therapeutic strategies in the treatment of patients with septic complications.

Effect on proliferation and apoptosis of retinoblastoma cell by RNA inhibiting high mobility group protein box-1 expression

AIM： To investigate the effect of high mobility group protein box-1 （HMGB1） siRNA on proliferation and apoptosis of retinoblastoma （Rb） cells.METHODS： The expression of HMGB1 in Rb cells were detected by real-time polymerase chain reaction （RT-PCR） and Western blot. Chemically synthesized HMGB1 siRNA was transfected into Y79 cells. The inhibitory rate was also examined by RT-PCR and Western blot. After HMGB1 siRNA transfection, the cell proliferation was analyzed by MTT, and cell apoptosis was detected by Caspase-3 active detection kit. Cell cycle distribution and apoptosis were detected by flow cytometry. RESULTS： The expression of HMGB1 significantly elevated in Rb cells （P〈0.01）. After transfected by siRNA, the HMGB1 protein level of Y79 cells was significantly reduced （P〈0.01）. After siRNA interference HMGB1, the proportion of proliferating cells reduced, and the proportion of quiescent cells increased （P〈0.05）. In addition, apoptosis rate of Y79 cells increased from 2.03% to 9.10% after interfering with HMGB1 siRNA （P〈0.05）.CONCLUSION： Specific HMGB1 siRNA can inhibit the expression of HMGB1. The effect may be attributed to inhibit the proliferation and promote cell apoptosis.

High mobility group protein 1: A collaborator in nucleosome dynamics and estrogen-responsive gene expression

High mobility group protein 1(HMGB1) is a multifunctional protein that interacts with DNA and chromatin to influence the regulation of transcription, DNA replication and repair and recombination. We show that HMGB1 alters the structure and stability of the canonical nucleosome(N) in a nonenzymatic,adenosine triphosphate-independent manner. As a result, the canonical nucleosome is converted to two stable, physically distinct nucleosome conformers. Although estrogen receptor(ER) does not bind to its consensus estrogen response element within a nucleosome, HMGB1 restructures the nucleosome to facilitate strong ER binding. The isolated HMGB1-restructured nucleosomes(N' and N'') remain stable and exhibit a number of characteristics that are distinctly different from the canonical nucleosome. These findings complement previous studies that showed(1) HMGB1 stimulates in vivo transcriptional activation at estrogen response elements and(2) knock down of HMGB1 expression by siR NA precipitously reduced transcriptional activation. The findings indicate that a major facet of the mechanism of HMGB1 action involves a restructuring of aspects of the nucleosome that appear to relax structural constraints within the nucleosome. The findings are extended to reveal the differences between ER and the other steroid hormone receptors. A working proposal outlines mechanisms that highlight the multiple facets that HMGB1 may utilize in restructuring the nucleosome.

Expression of high mobility group protein B1 in the lungs of rats with sepsis

BACKGROUND： Vibrio vulnifi cus inside the body could activate the NF-!B signaling pathwayand initiate the inflammatory cascade. The lung is one of the earliest organs affected by sepsisassociated with acute lung injury. High mobility group protein B1 （HMGB1） is an important late-actingpro-infl ammatory cytokine involving in the pathophysiology of sepsis. It is also involved in the injuryprocess in the lung, liver and intestine. There has been no report on the involvement of HMGB1 inVibrio vulnifi cus sepsis-induced lung injury.METHODS： Sixty rats were randomly divided into a normal control group （group A, n=10） anda Vibrio vulnificus sepsis group （group B, n=50）. Sepsis was induced in the rats by subcutaneousinjection of Vibrio vulnificus （concentration 6×108 cfu/mL, volume 0.1 mL/100g）） into the left lowerlimbs. The rats in group B were sacrifi ced separately 1, 6, 12, 24, and 48 hours after the infection.Their lungs were stored as specimens, lung water content was measured, and lung pathology wasobserved under a light microscope. The expressions of the HMGB1 gene and protein in the lungswere detected by RT-PCR and Western blot. Data were analyzed with one-way analysis of variance（ANOVA） and the LSD method for pair-wise comparison between the two groups. P〈0.05 wasconsidered statistically signifi cant.RESULTS： Compared to group A （0.652±0.177）, HMGB1 mRNA expression in the lungs ofgroup B was signifi cantly higher at 0 hour （1.161±0.358, P=0.013）, 24 hours （1.679±0.235, P=0.000）,and 48 hours （1.258±0.274, P=0.004） （P〈0.05）, and peaked at 24 hours. Compared to group A（0.594±0.190）, HMGB1 protein expression at 6 hours （1.408±0.567, P=0.026） after infection wassignificantly increased （P〈0. 05）, and peaked at 24 hours （2.415±1.064, P=0.000） after infection.Compared to group A （0.699±0.054）, lung water content was significantly increased at 6 hours（0.759±0.030, P=0.001）,12 hours （0.767±0.023, P=0.000）, 24 hours （0.771±0.043, P=0.000） and 48hours （0.789±0.137, P=0.000） after infection （P〈0.05）. Compared to group A, pathological changesat 12 hours in group B indicate marked pulmonary vascular congestion, interstitial edema andinfl ammatory infi ltration. Alveolar cavity collapse and boundaries of the alveolar septum could not beclearly identifi ed.CONCLUSION： Vibrio vulnifi cus sepsis can lead to injury in rat lungs, and increased HMGB1expression in lung tissue may be one of the mechanisms for injury from Vibrio vulnifi cus sepsis.

Epigenetic Repression of SATB1 by Polycomb Group Protein EZH2 in Epithelial Cells

Objective To study the regulatory mechanism of SATB1 repression in cells other than T cells or erythroid cells, which have high expression level of SATB1. Methods HeLa epithelial cells were treated with either histone deacetylase inhibitor (HDACi) trichostatin A (TSA) or DNA methylation inhibitor 5-Aza-C before detecting SATB1 expression. Luciferase reporter system was applied to measure effects of EZH2 on SATB1 promoter activity. Over-expression or knockdown of EZH2 and subsequent quantitative reverse transcription-polymerase chain reaction were performed to determine the effect of this Polycomb group protein on SATB1 transcription. Chromatin immunoprecipitation (ChIP) assay was applied to measure enrichment of EZH2 and trimethylated H3K27 (H3K27me3) at SATB1 promoter in HeLa cells. K562 cells and Jurkat cells, both having high-level expression of SATB1, were used in the ChIP experiment as controls. Results Both TSA and 5-Aza-C increased SATB1 expression in HeLa cells. Over-expression of EZH2 reduced promoter activity as well as the mRNA level of SATB1, while knockdown of EZH2 apparently enhanced SATB1 expression in HeLa cells but not in K562 cells and Jurkat cells. ChIP assay results suggested that epigenetic silencing of SATB1 by EZH2 in HeLa cells was mediated by trimethylation modification of H3K27. In contrast, enrichment of EZH2 and H3K27me3 was not detected within proximal promoter region of SATB1 in either K562 or Jurkat cells. Conclusion SATB1 is a bona fide EZH2 target gene in HeLa cells and the repression of SATB1 by EZH2 may be mediated by trimethylation modification on H3K27.

Effect of high mobility group box-1 protein on immune cells and its regulatory mechanism

High mobility group box-1 protein(HMGB1),which is a nuclear protein,participates in chromatin architecture and transcriptional regulation.When released from cells,HMGB1 also plays a well-established role as a pro-inflammatory mediator during innate immune responses to injury.In the initial stage of injury,there is a release of large quantities of early pro-inflammatory mediators to initiate or perpetuate immune responses against pathogens,but this pro-inflammatory period is transient,and it is followed by a prolonged period of immune suppression.At present,several lines of evidences have suggested that HMGB1 is a late cytokine provoking delayed endotoxin morbidity,which may enhance the production of early proinflammatory mediators,and it can contribute potently to the activation of different immune cells and play a role in the development of host cell-mediated immunity.The biology of HMGB1 has been extensively studied as a pro-inflammatory cytokine of systemic inflammation,however,this review will attempt to provide a summary of the effects of HMGB1 on different immune cells and its regulatory mechanism in acute insults.

Assessment of Sulf hydryl Group in Individual Rat Lens Protein Subunits During Galactose Cataract Development

A specific reagent DACM [N-( 7-Dimethylamino-4-methyl-3-coumarinyl) maleimide] is used to study the -SH groups in lens proteins of normal and galactose cataractous rats. DACM when reacts readily with -SH groups form strong fluorescent adducts. The two -dimensional electrophoresis with DACM pre-labeled proteins is a simple and sensitive method for detecting -SH groups of protein subunit. In the present study, based on IEF/SDS-PAGE electrophoretically characterized soluble crystallins, describes specific ...

Bmi-1,a Polycomb Group Protein,Plays an Essential Role in Tumorigenesis and Metastasis

Dysregulation of polycomb group protein Bmi-1 expression has been linked with an invasive phenotype of certain human cancers and poor prognosis of patients; however, the underlying mechanisms are

Relationship of plasma homocysteine, soluble intercellular adhesion molecule-1, high mobility group box 1 protein with carotid intima-media thickness in elderly patients with type 2 diabetes mellitus

Objective:To explore the relationship of plasma homocysteine(Hcy),soluble intercellular adhesion molecule-1(sICAM-1)and high mobility group box 1 protein(HMGB1)with carotid intima-media thickness(c-IMT)in elderly patients with type 2 diabetes mellitus.Methods:A total of 100 elderly patients who were diagnosed as type 2 diabetes mellitus in Baogang Hospital of Inner Mongolia from June 2017 to May 2020 were chosen as research objects.According to c-IMT,they were divided into the normal group(n=35),the mild to moderate group(n=41)and the severe group(n=24).The expression levels of plasma Hcy,sICAM-1 and HMGB1 were compared between groups respectively.Pearson’s correlation coefficient was used to analyze the relationship of plasma Hcy,sICAM-1,HMGB1 with c-IMT.Results:The comparison in plasma Hcy,sICAM-1,HMGB1 and c-IMT among the three groups of patients was of statistical significance(p<.05).The results of correlation analysis showed that the expression levels of plasma Hcy,sICAM-1 and HMGB1 were positively correlated with c-IMT in elderly patients with type 2 diabetes mellitus(r=.627,.598,.614;p<.05).Conclusions:The expression levels of plasma Hcy,sICAM-1 and HMGB1 are abnormally increased in elderly patients with type 2 diabetes mellitus,and related to c-IMT,which can provide a strong evidence for clinical diagnosis and treatment by detecting their levels in clinical practice.

血清HMGB1、CCL20、HSP27水平与慢性牙周炎患者牙周病变程度的相关性分析

目的探讨血清高迁移率族蛋白1(HMGB1)、CC趋化因子配体20(CCL20)、热休克蛋白27(HSP27)水平与慢性牙周炎(CP)患者牙周病变程度的相关性。方法选取2021年9月至2023年8月在新乡医学院第一附属医院诊治的60例CP患者纳入观察组,根据1∶1原则,另选取同期牙周健康者60例纳入对照组。比较两组及不同牙周病变程度CP患者血清HMGB1、CCL20、HSP27水平,分析各指标水平与CP牙周病变程度的相关性及联合诊断价值,并分析不同血清水平患者发生CP的危险度。结果观察组血清HMGB1、CCL20、HSP27水平高于对照组(P<0.05);不同牙周病变程度CP患者血清HMGB1、CCL20、HSP27水平比较:轻度<中度<重度,且各指标水平与牙周病变程度均呈正相关(P<0.05);入院时HMGB1、CCL20、HSP27水平联合诊断CP的曲线下面积(AUC)为0.905,最佳诊断敏感度为91.67%,特异度为88.33%,约登指数0.800,且各指标高水平患者发生CP的危险度是低水平的1.105倍、1.034倍、1.105倍(P<0.05)。结论HMGB1、CCL20、HSP27在CP患者血清中呈异常高表达,各指标水平与牙周病变程度均呈正相关,且联合检测对CP具有较高诊断价值,可作为临床诊断CP、评估牙周病变程度的有效指标。

Hmo1:A versatile member of the high mobility group box family of chromosomal architecture proteins

Eukaryotic chromatin consisting of nucleosomes connected by linker DNA is organized into higher order structures,which is facilitated by linker histone H1.Formation of chromatin compacts and protects the genome,but also hinders DNA transactions.Cells have evolved mechanisms to modify/remodel chromatin resulting in chromatin states suitable for genome functions.The high mobility group box(HMGB)proteins are non-histone chromatin architectural factors characterized by one or more HMGB motifs that bind DNA in a sequence nonspecific fashion.They play a major role in chromatin dynamics.The Saccharomyces cerevisiae(yeast hereafter)HMGB protein Hmo1 contains two HMGB motifs.However,unlike a canonical HMGB protein that has an acidic C-terminus,Hmo1 ends with a lysine rich,basic,C-terminus,resembling linker histone H1.Hmo1 exhibits characteristics of both HMGB proteins and linker histones in its multiple functions.For instance,Hmo1 promotes transcription by RNA polymerases I and II like canonical HMGB proteins but makes chromatin more compact/stable like linker histones.Recent studies have demonstrated that Hmo1 destabilizes/disrupts nucleosome similarly as other HMGB proteins in vitro and acts to maintain a common topological architecture of genes in yeast genome.This minireview reviews the functions of Hmo1 and the underlying mechanisms,highlighting recent discoveries.

基于转录组比较的燕麦类似贮藏蛋白在小麦面团强度性状中的作用

面团强度是影响小麦加工应用的主要品质指标之一。为鉴定与面团强度性状相关的贮藏蛋白编码基因,本研究以高相对分子量麦谷蛋白亚基组合完全相同的2个品种(郑麦366:高面团强度品种;郑麦366杂交后代品种郑麦369:低面团强度品种)为研究材料,比较开花后14 d、21 d、28 d的贮藏蛋白编码基因表达差异,评估基因编码蛋白的面团强度贡献值,以及面粉的巯基含量差异等。结果表明,在25个显著差异表达的贮藏蛋白编码基因中,无高相对分子量麦谷蛋白亚基、低相对分子量麦谷蛋白亚基基因;其中郑麦366显著上调表达基因14个,包含8个燕麦类似蛋白编码基因和6个醇溶蛋白编码基因;显著下调表达基因11个,包括10个醇溶蛋白编码基因和1个燕麦类似蛋白编码基因。贮藏蛋白面团强度评价模型的评分结果显示,差异表达基因编码的燕麦类似贮藏蛋白对面团强度性状的贡献值不低于优质麦谷蛋白亚基,暗示燕麦类似贮藏蛋白可能也是影响面团强度性状的重要蛋白质类型。

circLRP6调节miR-31-5p/HMGA1轴对高糖诱导的肾小管上皮细胞损伤的影响

目的分析circLRP6调节miR-31-5p/高迁移率族蛋白A1(HMGA1)轴对高糖诱导的肾小管上皮细胞损伤的影响。方法体外培养人肾小管上皮HK-2细胞,分为对照组、高糖组、高糖+si-NC组、高糖+si-circLRP6组、高糖+si-circLRP6+miR-NC组、高糖+si-circLRP6+miR-31-5p inhibitor组、高糖+si-circLRP6+miR-31-5p inhibitor+si-NC组、高糖+si-circLRP6+miR-31-5p inhibitor+si-HMGA1组,对照组置于含5.5 mmol/L葡萄糖的DMEM培养基内培养,其余各组置于含25 mmol/L葡萄糖的DMEM培养基内培养,并通过Lipofectamine 2000将相应质粒转染至HK-2细胞内,实时定量PCR测定细胞circLRP6、miR-31-5p、HMGA1 mRNA水平,试剂盒测定细胞上清液白细胞介素-6(IL-6)水平、肿瘤坏死因子α(TNF-α)水平、乳酸脱氢酶(LDH)活性以及丙二醛(MDA)含量;流式细胞仪观察细胞凋亡;Western blotting测定细胞HMGA1、Bax、Bcl-2蛋白表达;双荧光素酶报告基因实验分析miR-31-5p与circLRP6、HMGA1的靶向关系。结果与对照组相比,高糖组HK-2细胞circLRP6水平升高,细胞增殖抑制率、IL-6、TNF-α、LDH、MDA、细胞凋亡率、Bax蛋白表达升高,Bcl-2蛋白表达降低(均P<0.05);与高糖组相比,高糖+si-circLRP6组HK-2细胞增殖抑制率、IL-6、TNF-α、LDH、MDA、细胞凋亡率、Bax蛋白表达降低,Bcl-2蛋白表达升高(均P<0.05);与高糖+si-circLRP6组相比,高糖+si-circLRP6+miR-31-5p inhibitor组HK-2细胞增殖抑制率、IL-6、TNF-α、LDH、MDA、细胞凋亡率、Bax蛋白表达升高,Bcl-2蛋白表达降低(均P<0.05);与高糖+si-circLRP6+miR-31-5p inhibitor组相比,高糖+si-circLRP6+miR-31-5p inhibitor+si-HMGA1组HK-2细胞增殖抑制率、IL-6、TNF-α、LDH、MDA、细胞凋亡率、Bax蛋白表达降低,Bcl-2蛋白表达升高(均P<0.05)。miR-31-5p与circLRP6、HMGA1均具有靶向关系。结论沉默circLRP6可能通过上调miR-31-5p,抑制HMGA1表达,对高糖诱导的肾小管上皮细胞损伤发挥保护作用。