Role of PCK2 in the proliferation of vascular smooth muscle cells in neointimal hyperplasia

Vascular smooth muscle cell (VSMC) proliferation is a hallmark of neointimal hyperplasia (NIH) in atherosclerosis and restenosis post-balloon angioplasty and stent insertion. Although numerous cytotoxic and cytostatic therapeutics have been developed to reduce NIH, it is improbable that a multifactorial disease can be successfully treated by focusing on a preconceived hypothesis. We, therefore, aimed to identify key molecules involved in NIH via a hypothesis-free approach. We analyzed four datasets ({"type":"entrez-geo","attrs":{"text":"GSE28829","term_id":"28829"}}GSE28829, {"type":"entrez-geo","attrs":{"text":"GSE43292","term_id":"43292"}}GSE43292, {"type":"entrez-geo","attrs":{"text":"GSE100927","term_id":"100927"}}GSE100927, and {"type":"entrez-geo","attrs":{"text":"GSE120521","term_id":"120521"}}GSE120521), evaluated differentially expressed genes (DEGs) in wire-injured femoral arteries of mice, and determined their association with VSMC proliferation in vitro. Moreover, we performed RNA sequencing on platelet-derived growth factor (PDGF)-stimulated human VSMCs (hVSMCs) post-phosphoenolpyruvate carboxykinase 2 (PCK2) knockdown and investigated pathways associated with PCK2. Finally, we assessed NIH formation in Pck2 knockout (KO) mice by wire injury and identified PCK2 expression in human femoral artery atheroma. Among six DEGs, only PCK2 and RGS1 showed identical expression patterns between wire-injured femoral arteries of mice and gene expression datasets. PDGF-induced VSMC proliferation was attenuated when hVSMCs were transfected with PCK2 siRNA. RNA sequencing of PCK2 siRNA-treated hVSMCs revealed the involvement of the Akt-FoxO-PCK2 pathway in VSMC proliferation via Akt2, Akt3, FoxO1, and FoxO3. Additionally, NIH was attenuated in the wire-injured femoral artery of Pck2-KO mice and PCK2 was expressed in human femoral atheroma. PCK2 regulates VSMC proliferation in response to vascular injury via the Akt-FoxO-PCK2 pathway. Targeting PCK2, a downstream signaling mediator of VSMC proliferation, may be a novel therapeutic approach to modulate VSMC proliferation in atherosclerosis.     

牡丹根部内生细菌的分离鉴定及脂肽类物质的拮抗活性研究

【目的】从牡丹(Paeonia suffruticosa Andr.)根部组织中分离鉴定内生细菌,测定拮抗菌株脂肽类活性物质的体外抑菌活性。【方法】采用平板对峙法筛选出对牡丹灰霉病菌(Botrytis paeoniae Oadem)、牡丹炭疽病菌(Gloeosporium sp.)、牡丹黑斑病菌(Altenaria sp.)、牡丹黄斑病菌(Phyllosticta commonsii)有拮抗作用的内生细菌。基于形态特征、生理生化特性和16S rRNA基因序列同源性鉴定拮抗菌株。根据脂肽类抗菌物质合成相关基因序列对拮抗菌株进行基因扩增检测,采用酸沉淀法提取拮抗菌株的脂肽类物质,平板对峙法测定脂肽类物质的体外抑菌活性。【结果】从牡丹根部组织中共分离获得62株内生细菌,其中菌株Md31和Md33对4种病原菌均有较明显的抑制作用。Md31和Md33被鉴定为解淀粉芽孢杆菌(Bacillus amyloliquefaciens)。通过对菌株Md31和Md33进行5个脂肽类合成功能基因bmyB、fenD、ituC、srfAA和srfAB的检测,序列同源性分析,表明两个菌株具有合成脂肽类物质的能力。菌株Md31和Md33的脂肽类粗提物对所测试的牡丹病原真菌均具有不同程度的抑制作用。【结论】获得了2株对牡丹病原菌有良好抑制效果的解淀粉芽孢杆菌Md31和Md33,两个菌株的脂肽类粗提物也具有较强的体外抑菌活性,该研究为牡丹内生细菌的进一步开发应用奠定了基础。     

萎缩芽胞杆菌CKL1促盐胁迫下燕麦生长活性及其功能基因分析

【背景】青海省特殊生境孕育了特殊微生物资源。【目的】探究适合生活于高原生境的芽胞杆菌菌源。【方法】采用平板对峙法、显色法对萎缩芽胞杆菌（Bacillus atrophaeus） CKL1的拮抗、产吲哚乙酸活性进行测定,并检测耐低温、耐盐性及菌株对盐胁迫下燕麦品种（Avena sativa）“青燕1号”种子萌发、幼苗生长效应及叶绿素、脯氨酸、丙二醛的含量变化,利用二代测序技术对菌株进行基因组测序并分析相关功能基因。【结果】菌株CKL1对禾谷镰孢菌（Fusarium graminearum）、锐顶镰孢菌（Fusarium acuminatum）表现出显著的拮抗活性（抑菌圈直径>15 mm）;与Salkowski比色液反应变红,能在NaCl浓度为13%的LB培养基及4℃低温下生长,表现出一定的产吲哚乙酸、耐盐及耐低温活性;盐胁迫下,菌株CKL1对“青燕1号”种子萌发及幼苗生长具有显著促进作用,叶绿素及脯氨酸含量显著增加,丙二醛含量下降,增强了燕麦的抗盐性。菌株CKL1基因组全长为14 281 280 bp,与GO功能数据库比对注释到3 303个功能基因;基因组编码与脂肽类化合物itur...     

北京山区黄栌叶片水分和光能利用效率季节变化及影响因子

未来气候变化将影响光合环境资源供给,尤其是水分和光能。为深入了解植物对气候变化的适应性,使用LI-6800便携式光合仪,于2021年5—10月份(完全展叶期)测定了北京山区广布灌木黄栌(Cotinus coggygria)叶片的光响应曲线,分析其水分利用效率(WUE=最大净光合速率[P_nmax]/气孔导度[g_s])和光能利用效率(LUE)的季节变化特征及影响因子。结果显示：黄栌叶片WUE在5—6月份呈下降趋势,7—10月份比较稳定;LUE在5—7月份呈上升趋势,8—10月份比较稳定。WUE和LUE的生长季平均值分别为98.25μmol/mol和0.06 mol/mol,变异系数分别为22%和17%,两者呈负相关(R²=0.86;P<0.01)。环境因子中,WUE和LUE主要受土壤含水量(SWC)影响,WUE随SWC增加呈线性降低趋势,而LUE随SWC增加呈线性增加趋势。SWC每增加0.1 m³/m³,P_nmax和g_s分别线性增加...     

六种海桑属红树植物根际真菌特征及其影响因子

真菌多样性是植物根际生态系统的重要构成与植物健康稳定的重要指标。海桑属是红树林的先锋物种,采用真菌ITS1区高通量测序方法,分析了六种海桑属红树根际真菌的组成和多样性,结合土壤理化性质探讨影响不同植物根际真菌群落组成差异的因素。结果显示,根际真菌隶属于7门、96科、155属,子囊菌门作为优势菌门在海桑属不同红树中相对丰度无显著差异,都超过27%,但次优势的担子菌门丰度含量有差异;属水平上,优势菌属的丰度含量不同,曲霉属在卵叶海桑的丰度最高(29.57%),在海南海桑最低(3.47%)。六种红树植物根际存在特有的代表类群,如无瓣海桑的马拉色菌(9.31%)和毛腐菌属(10.05%),海南海桑中的Talaromyces(19.61%)和Acremonium(13.58%)。比较多样性指数Simpson和Shannon,发现拟海桑是六种植物中丰度最高的,卵叶海桑最低。RDA分析发现子囊菌门与全磷含量呈显著负相关,担子菌门与速效钾呈明显正相关。六种海桑属红树植物根际核心物种分析表明,优势真菌类群曲霉属和一些低丰度的真菌类群,通过降解有机质参与碳循环,对根际土壤生态系统的稳定起重要作用。六种海桑...     

凋落物和根系对沂蒙山区三种森林恢复方式土壤酸性磷酸酶动力学特征的影响

酸性磷酸酶动力学特征可反映不同底物供应下土壤磷转化状况。以人工恢复的引进种黑松人工林和本地种栓皮栎人工林以及自然恢复的天然次生林为研究对象,开展凋落物添加去除和根系去除对土壤酸性磷酸酶动力学特征的影响研究。结果表明:(1)三种森林恢复方式下土壤酸性磷酸酶活性和最大反应速度(V_max)均为双倍凋落物＞对照＞去除凋落物＞去除根系＞无输入;(2)改变凋落物和根系输入对酸性磷酸酶的半饱和常数(K_m)和催化效率(V_max/K_m)影响不大;(3)酸性磷酸酶活性受速效磷含量的反馈调节;土壤氮可利用性和含水量显著影响酸性磷酸酶活性。改变凋落物和根系输入对引进种黑松人工林土壤有机磷转化能力影响最大,天然次生林次之,本地种栓皮栎人工林最稳定。根系对土壤酸性磷酸酶动力学特征的影响大于凋落物。其结果可为暖温带森林恢复,应对气候变化和森林防火、收集凋落物等管理措施提供理论依据。     

Metal migration and subunit swapping in ALS-linked SOD1: Zn2+ transfer between mutant and wild-type occurs faster than the rate of heterodimerization

The heterodimerization of WT Cu, Zn superoxide dismutase-1 (SOD1), and mutant SOD1 might be a critical step in the pathogenesis of SOD1-linked amyotrophic lateral sclerosis (ALS). Rates and free energies of heterodimerization (ΔG_Het) between WT and ALS-mutant SOD1 in mismatched metalation states—where one subunit is metalated and the other is not—have been difficult to obtain. Consequently, the hypothesis that under-metalated SOD1 might trigger misfolding of metalated SOD1 by “stealing” metal ions remains untested. This study used capillary zone electrophoresis and mass spectrometry to track heterodimerization and metal transfer between WT SOD1, ALS-variant SOD1 (E100K, E100G, D90A), and triply deamidated SOD1 (modeled with N26D/N131D/N139D substitutions). We determined that rates of subunit exchange between apo dimers and metalated dimers—expressed as time to reach 30% heterodimer—ranged from t_30% = 67.75 ± 9.08 to 338.53 ± 26.95 min; free energies of heterodimerization ranged from ΔG_Het = -1.21 ± 0.31 to -3.06 ± 0.12 kJ/mol. Rates and ΔG_Het values of partially metalated heterodimers were more similar to those of fully metalated heterodimers than apo heterodimers, and largely independent of which subunit (mutant or WT) was metal-replete or metal-free. Mass spectrometry and capillary electrophoresis demonstrated that mutant or WT 4Zn-SOD1 could transfer up to two equivalents of Zn²⁺ to mutant or WT apo-SOD1 (at rates faster than the rate of heterodimerization). This result suggests that zinc-replete SOD1 can function as a chaperone to deliver Zn²⁺ to apo-SOD1, and that WT apo-SOD1 might increase the toxicity of mutant SOD1 by stealing its Zn²⁺.     

Pygopus2 ameliorates mesenteric adipocyte poor differentiation to alleviate Crohn's disease ‐like colitis via the Axin2/GSK3β pathway

ObjectivesCrohn''s disease (CD) mesenteric adipose tissue (MAT) inflammation affects enteritis through the interaction between the mesentery and intestine, and we previously found that poorly differentiated mesenteric adipocytes were related to its inflammatory features. Pygopus2 (Pygo2) is a key negative regulator of adipocyte differentiation. We aimed to determine whether Pygo2 participates in CD mesenteric lesions and whether Pygo2 knockdown would be beneficial in a CD model (Il‐10 ^−/− mice).MethodsPygo2 expression in MAT from control and CD patients and Il‐10 ^−/− mice was measured by immunohistochemistry. Lentiviral transfection was used to regulate Pygo2 expression in Il‐10 ^−/− mice, and the effects on mesenteric adipocyte differentiation, inflammation, and dysfunction during spontaneous colitis, as well as the possible mechanism, were investigated.ResultsPygo2 expression was increased in MAT from CD patients and Il‐10 ^−/− mice, and its expression correlated with poor adipocyte differentiation and inflammation. Pygo2 knockdown significantly ameliorated colitis in Il‐10 ^−/− mice. Moreover, the downregulation of Pygo2 gene expression could promote adipocyte differentiation and inhibit adipocyte inflammation in vivo and in vitro, and the effects were at least partly mediated by the Axis inhibition protein 2 (Axin2)/glycogen synthase kinase 3 beta (GSK3β) pathway.ConclusionsThe increase in Pygo2 may be related to mesenteric adipocyte poor differentiation and inflammatory features of CD, and Pygo2 inhibition could alleviate CD‐like colitis by improving mesenteric lesions by regulating the Axin2/GSK3β pathway.

The increase in Pygopus2 (Pygo2) may be related to mesenteric adipocyte poor differentiation and inflammatory features of CD, and Pygo2 inhibition could alleviate adipocyte differentiation and mesenteric lesions by regulating the Axin2/GSK3β pathway.     

Mutant C/EBPα p30 alleviates immunosuppression of CD8+ T cells by inhibiting autophagy‐associated secretion of IL‐1β in AML

ObjectivesMutant C/EBPα p30 (mp30), the product of C/EBPα double mutations (DM), lacks transactivation domain 1 and has C‐terminal loss‐of‐function mutation. Acute myeloid leukaemia (AML) patients harbouring C/EBPα DM could be classified as a distinct subgroup with favourable prognosis. However, the underlying mechanism remains elusive.Materials and MethodsAutophagy regulated by mp30 was detected by western blot and immunofluorescence. Immune infiltration analysis and GSEA were performed to investigate autophagic and inflammatory status of AML patients from the {"type":"entrez-geo","attrs":{"text":"GSE14468","term_id":"14468"}}GSE14468 cohort. Flow cytometry was applied to analyse T cell activation.ResultsMp30 inhibited autophagy by suppressing nucleus translocation of NF‐κB. Autophagy‐associated secretion of IL‐1β was decreased in mp30‐overexpressed AML cells. Bioinformatic analysis revealed that inflammatory status was attenuated, while CD8⁺ T cell infiltration was upregulated in C/EBPα DM AML patients. Consistently, the proportion of CD8⁺CD69⁺ T cells in peripheral blood mononuclear cells (PBMCs) was upregulated after co‐culture with mp30 AML cell conditional culture medium. Knock‐out of IL‐1β in AML cells also enhanced CD8⁺ T cell activation. Accordingly, IL‐1β expression was significantly reduced in the bone marrow (BM) cells of C/EBPα DM AML patients compared to the wildtype, while the CD8⁺CD69⁺ T cell proportion was specifically elevated.ConclusionsC/EBPα DM alleviates immunosuppression of CD8⁺ T cells by inhibiting the autophagy‐associated secretion of IL‐1β, which elucidated that repression of autophagy‐related inflammatory response in AML patients might achieve a favourable clinical benefit.

Mp30 suppresses autophagy‐associated IL‐β secretion, which ultimately alleviates the immunosuppression of CD8⁺ T cells in the microenvironment, contributing to favourable prognosis of AML patients.     

Black soldier fly (Hermetia illucens) larvae oil as an alternative fat ingredient to soybean oil in laying hen diets

ObjectiveThe objective of this study was to determine whether dietary black soldier fly (Hermetia illucens, HI) larvae oil (HILO) could serve as an alternative fat source to soybean oil (SBO) in laying hen diets.MethodsWe randomly assigned 25-week-old Hy-line Brown laying hens (n = 144) to receive (n = 6 hens/group; eight replicates) a control or an experimental diet in which SBO was replaced with 50% (50HILO) or 100% HILO (100HILO).ResultsDietary HILO did not negatively affect body weight or productive performance during the study. The eggs also had similar quality parameters, proximate composition, and cholesterol levels. However, the yolk color index was significantly higher (p<0.01) in the 100HILO than in the other groups. Dietary HILO significantly altered the composition of fatty acids (FAs) in abdominal fat and eggs. Total saturated fatty acids (SFAs) and total polyunsaturated FAs (PUFAs) were significantly increased and decreased in the 50HILO and 100HILO groups, respectively, compared with those in the control group (p<0.001 and p<0.0001, respectively). Specifically, the medium-chain FAs lauric and myristic acids were remarkably increased in the abdominal fat of laying hens fed HILO (p<0.0001), whereas only myristic acid increased in eggs (p<0.0001). Undesirable heavy metal (aluminum, fluorine, arsenic, lead, mercury, and cadmium) concentrations were below permissible limits in eggs.ConclusionWe considered that HILO could be an alternative dietary fat to SBO for laying hens with maintained productive performance and good egg quality.