Tissu adipeux : glande endocrine polyvalente

Research on the adipose tissue secretions has undergone a major inflection for a few years. The investigators became aware that the biology of the various cells of the stroma-vascular fraction of adipose tissue was to be taken into account to include/understand the diversity of the secretions. The adipose tissue is a site of production of adipokines (strictly synthesized and secreted by the adipocyte), of pro- or anti-inflammatory cytokines and of acute phase reactant proteins. Cells of the stroma-vascular fraction (monocytes/macrophages, microvascular endothelial cells…) have a secretory importance neglected up to now. The action of both major adipokines (leptin and adiponectin) will be more particularly evoked while considering the diversity of action of the other most current factors. Finally some secreted products more recently discovered (apelin, autotaxin, visfatin.), whose properties are still in the course of exploration, will be also mentioned.     

A novel adipokine WISP1 attenuates lipopolysaccharide-induced cell injury in 3T3-L1 adipocytes by regulating the PI3K/Akt pathway

BackgroundTo study the effect of WISP1 on lipopolysaccharide (LPS)-induced cell injury in 3T3-L1 adipocytes.MethodLentivirus was transiently transfected into log phase 3T3-L1 adipocytes, which were then treated with LPS at a concentration of 10 μg/mL for 24 h. The cells were divided into the following groups: group A (control, untreated cells); group B (LPS-treated cells); group C (GFP), cells transfected with lentivirus-containing GFP; group D (GFP+LPS), group C treated with LPS;group E (WISP1OE), cells transfected with lentivirus, group F (shNC+LPS), cells transfected with lentivirus-containing nshRNA treated with LPS; group G (shWISP1 +LPS), cells transfected with lentivirus-containing shRNA treated with LPS; group H (WISP1OE+LPS), group E treated with LPS; group I (WISP1OE+LPS+LY294002), group E treated with LPS followed by LY294002 for 24 h.ResultsWISP1 overexpression notably ameliorated cell apoptosis, accompanied with the increased expression of bcl-2, the decreased expressions of bax and cleaved-caspase-3, and promoted the release of inflammatory factors, such as tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in LPS-treated 3T3-L1 adipocytes. WISP1 knockdown exhibited the opposite results. In addition, WISP1 stimulated Akt phosphorylation and reduced nuclear translocation of Fork head box protein O3 (FoxO3a) in 3T3-L1 adipocytes treated by LPS. The inhibition of the PI3K/Akt signaling pathway diminished the protective effect of WISP1.ConclusionWISP1 prevents 3T3-L1 adipocytes from being injured by LPS by regulating the PI3K/Akt pathway.     

Apelin-13对高尿酸诱导的脂肪细胞氧化应激的作用

目的:探讨apelin-13对高尿酸诱导的3T3-L1脂肪细胞氧化应激的作用及其机制。方法:3T3-L1脂肪细胞予以10 mg/dL尿酸刺激,部分细胞予以1μmol/L apelin-13预处理,以100μmol/L H₂O₂刺激的细胞为阳性对照。48 h后,流式细胞术检测活性氧族(ROS)含量,生化试剂盒检测细胞及培养液上清抗氧化酶[超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)和过氧化氢酶(CAT)]和促氧化酶NADPH氧化酶(NOX)的活性以及丙二醛(MDA)含量,real-time PCR法检测细胞局部肾素-血管紧张素系统(RAS)各组份血管紧张素原(AGT)、血管紧张素转化酶I(ACE1)、血管紧张素II 1型受体(AT1R)和AT2R及血管紧张素II 1型受体相关蛋(APJ)的mRNA表达水平,ELISA法检测细胞及培养液中血管紧张素II(AngⅡ)浓度。结果:10 mg/dL的尿酸明显降低3T3-L1脂肪细胞SOD、GSH-Px和CAT的活性,升高NOX的活性,增加MDA的含量,细胞内ROS的含量相应升高;apel...     

丹参酮ⅡA在3T3-L1脂肪细胞中促进脂联素的组装(英文)

目的:研究丹参酮IIA对3T3-L1脂肪细胞中脂联素表达和蛋白多聚化的作用。方法:用油红染色鉴定3T3-L1脂肪细胞的分化状态。用Real-time PCR检测基因的mRNA表达水平。用2%15%梯度SDS-PAGE及Western blot检测脂联素三种聚体。用siRNA技术进行基因沉默。结果:本研究发现传统中药丹参中的萘醌二萜类成分丹参酮IIA在脂肪细胞中激活AMPK通路,同时抑制脂肪细胞分化,降低脂联素的表达。而在前体脂肪细胞分化过程或成熟脂肪细胞中加入丹参酮IIA进行处理,均明显促进脂联素的多聚化,增加高聚体的浓度。抑制AMPK通路能够解除丹参酮IIA对脂联素的作用。结论:丹参酮IIA通过激活AMPK在3T3-L1脂肪细胞中促进脂联素的组装。     

不同来源人脂肪细胞脂联素分泌比较及干预研究

目的分析不同部位(腹部皮下、内脏网膜)人脂肪细胞脂联素分泌有无差异及游离脂肪酸(FFAs)、罗格列酮(RSG)干预后的改变。方法取6例外科手术女性患者的网膜及皮下脂肪组织,胶原酶消化分离纯化成熟脂肪细胞。观察两部位来源细胞在基础状态下不同时点(4、6、12、24h)脂联素分泌浓度的差异及加入FFAs(PA0.5mmol/L+OA1.0mmol/L)、FFAs+RSG(PA0.5mmol/L+OA1.0mmol/L+RSG10μmol/L)培养24h后脂联素浓度差异。结果整体上,网膜脂肪细胞基础脂联素分泌较皮下高,平均高5%,但差异无统计学意义。以基础脂联素分泌作对照,培养24h后,在网膜脂肪细胞,FFAs抑制脂联素分泌,脂联素浓度降至对照组66%,加入RSG联合培养后,逆转至对照组97%(P<0.05);而在皮下脂肪细胞,FFAs对脂联素分泌无抑制作用,RSG亦无促进脂联素分泌。结论人网膜脂肪细胞与皮下脂肪细胞在基础状态下脂联素分泌无差别,但在FFAs和(或)RSG介导的脂联素分泌中存在异质性。     

人骨髓基质干细胞体外诱导和鉴定

目的建立人自体骨髓基质干细胞(human menchymal stem cells,hMSCs)体外分离、鉴定体系,建立经诱导表达成骨、软骨、脂肪细胞表型的体外培养体系,探讨作为骨组织工程种子细胞的可能性,为组织工程技术应用打下基础。方法抽取患者骨髓5ml,以密度梯度法分离hMSCs,使用流式细胞仪鉴定细胞表型;应用免疫组化和分子生物学技术,对hMSCs进行成骨、软骨、脂肪细胞的诱导培养和表型鉴定。结果第2代hMSCs阳性细胞表达率CD105(78±6)%,CD166(43±7)%,CD29(69±12)%;hMSCs成骨诱导培养第3代平均扩增(163.4±13.4)倍,形成钙结节,骨细胞转录因子、骨钙素、骨桥蛋白和I型胶原免疫荧光阳性,RT-PCR证实有I型胶原、骨钙素、骨桥蛋白和骨结合素mRNA表达,对照组阴性;成软骨细胞诱导培养后II型胶原、SOX9(SRY-type HMGbox9,是在哺乳动物性别决定和软骨生成中起着关键调控作用的一个基因)。免疫荧光阳性,RT-PCR证实II型胶原、软骨聚集蛋白聚糖mRNA表达,对照组阴性;成脂肪细胞诱导培养后,油红-O染色阳性,RT-PCR证实PPAR2 mRNA...     

Recombinant C3adesArg/acylation stimulating protein (ASP) is highly bioactive: A critical evaluation of C5L2 binding and 3T3-L1 adipocyte activation

C5L2 is a recently identified receptor for C5a/C5adesArg, C3a and C3adesArg (ASP). C5a/C5adesArg bind with high affinity, with no identified activation. By contrast, some studies demonstrate C3a/ASP binding/activation to C5L2; others do not. Our aim is to critically evaluate ASP/C3adesArg-C5L2 binding and bioactivity.Cell-associated fluorescent-ASP (Fl-ASP) binding to C5L2 increased from transiently transfected < stably transfected < Fl-ASP-sorted C5L2-HEK for both human C5L2 and mouse C5L2. Transfected C5L2-CHO cells had similar results. Endogenous C5L2 expression increased from 3T3-L1 preadipocytes < 3T3-L1 adipocytes < primary mouse adipocytes. Non-transfected cells ± Fl-ASP demonstrated background fluorescence only.In adherent C5L2-HEK (Fl-ASP sorted) and 3T3-L1 cells, blocking with 10% fetal calf serum, protamine sulfate or ovalbumin prevented ¹²⁵I-ASP non-specific binding (NSB, no cells), while albumin increased NSB. Binding to non-transfected HEK was comparable to NSB. Optimal specific binding was obtained at 20 °C (vs. 4 °C) in PBS or serum-free medium with K_d 83.7 ± 23.7 nM (C5L2-HEK), 66 ± 15 nM (C5L2-CHO) and 76 ± 14.3 nM (3T3-L1 preadipocytes); ¹²⁵I-C5a binding had greater affinity. Fl-ASP-C5L2 binding was comparable and concentration dependent (K_d 31 nM (direct binding) and IC₅₀ 35 nM (competition binding) regardless of conditions).Recombinant ASP (rASP) produced in modified Escherichia coli Origami (DE3) (allowing folding and disulphide bridge formation), purified under non-denaturing conditions demonstrated 10× greater bioactivity vs. proteolytically derived plasma ASP for triglyceride synthesis and fatty acid uptake in 3T3-L1 adipocytes and preadipocytes while adipose tissue from C5L2 KO mice was non-responsive. rASP stimulation of adipocyte BODIPY-fatty acid uptake demonstrated EC₅₀ 115 ± 93 nM and maximal stimulation of 413 ± 33%, p < 0.001. ASP binding has distinct characteristics that lead to C5L2 activation and increased bioactivity.