SVF细胞影响移植脂肪LPL表达的实验研究

目的 探讨脂肪移植后脂蛋白脂肪酶(Lipoprotein lipase,LPL)在SVF(Stromal vascular fraction)辅助移植脂肪组织中的表达模式。方法 本实验以新鲜分离的自体SVF细胞辅助移植的脂肪组织为实验组,以脂肪组织与等量生理盐水的混合物为对照组,随机在裸鼠背部两侧注射建立动物模型,对两组移植物进行脂周蛋白免疫组化染色分析、LPL免疫组化染色分析和LPL蛋白含量测定。结果 TPL蛋白含量测定及LPL免疫组化染色分析显示,随着移植时间的延长,实验组和对照组LPL表达逐渐下降,在第2周、第3周、第4周时,实验组LPL表达明显高于对照组。脂周蛋白免疫组化染色分析显示,两组脂肪移植后的存活细胞均逐渐减少,但第2周、第3周、第4周实验组存活细胞数高于对照组。结论 SVF促进脂肪组织表达LPL,表达差异在移植后的第2周开始出现,这可能与SVF提高脂肪组织存活率和ASCs(Adipose-derived stromal cells)成脂分化有关。     

非酒精性脂肪性肝病发生中Perilipin和ADRP表达变化实验研究

目的：探讨高脂饮食诱导大鼠非酒精性脂肪性肝病（NAFLD）形成过程中，肝组织Perilipin（脂滴包被蛋白）和ADRP（脂肪分化相关蛋白）的表达变化及意义。方法：建立大鼠高脂饮食NAFLD模型并设正常对照组，观察时间点为建模后4w、8w和12w，观察大鼠肝组织病理变化，检测血浆游离脂肪酸及甘油三酯水平，免疫组织化学染色法检测肝组织中Perilipin和ADRP蛋白水平。结果：成功制作大鼠NAFLD实验模型，与正常组相比，NAFLD组大鼠血浆FFA水平和TG水平从第4w起显著升高（P〈0．01），NAFLD组大鼠肝组织Perilipin蛋白表达水平显著低于正常组（P〈0．01），ADRP蛋白表达水平显著高于同期正常组（P〈0．01）。Perilipin及ADRP表达与FFA及TG具有显著相关性。结论：高脂饮食可制作NAFLD大鼠模型，Perilipin和ADRP与NAFLD的形成有关。     

Perilipin Gene 1237 T＞C Polymorphism is not Associated with Obesity Risk in Northern Chinese Han Adults

Objective To identify the association between PLIN 1237 polymorphism and obesity in Chinese Han adults. Methods A total of 994 adults （157 obese subjects, 322 overweight subjects, and 515 normal controls） were recruited from two rural communities. PLIN 1237 polymorphism was genotyped by polymerase chain reaction-restriction-fragment-length-polymorphism （PCR-RFLP）. Association between PLIN polymorphisms and obesity status was estimated by ordinal logistic regression. Results The three genotypes of,PLIN 1237 were detected with a percentage of 54.3%, 37.1%, and 8.6% in TT, TC, and CC genotypes, respectively. For the PLIN 1237 polymorphism locus, the frequency of alleles T and C was 0.73 and 0.27, respectively. The PLIN 1237 polymorphisms were in Hardy-Weinberg equilibrium. PLIN 1237 polymorphism was not associated with obesity. The odds ratio for overweight or obesity for the CC＋TC genotype was 0.8（0.4, 1.4） in women （P=-0.4） and 0.6 （0.3, 1.3） in men （P=-0.2） after adjustment for age, education, household income and alcohol consumption, smoking, and physicalactivity. Conclusion Chinese Han adults have a lower frequency of variant-allele C in PLIN1237. PLIN1237 T〉C polymorphism is not significantly associated with obesity in northern Chinese adults.     

衰老雄性大鼠雄激素缺乏与睾丸间质细胞中Perilipin基因表达间关系的研究

目的 探讨Perilipin基因差异表达与雄激素缺乏之间的关系.方法 采用D-半乳糖注射的方法建立大鼠亚急性衰老雄激素缺乏动物模型,检测血清总睾酮（TT）、超氧化物岐化酶（SOD）、丙二醛（MDA）水平判断造模成功,采用Percoll非连续密度梯度离心法分离睾丸间质细胞（Leydig）,原代培养72h后提取大鼠Leydig细胞总RNA,基因芯片筛选差异表达基因,最后半定量RT-PCR验证芯片结果.结果 与对照组比较,模型组（低剂量组和高剂量组）血清TT浓度、SOD活力下降,MDA含量升高,差异有统计学意义;基因芯片结果显示,脂类代谢途径与雄激素缺乏最为相关,差异表达基因分别为Perilipin,Fabp4,Mgll;半定量RT-PCR结果发现,Perilipin基因及其两个亚型表达显著下调.结论 在RNA水平Perilipin基因表达与雄激素缺乏可能有关.     

脂滴包被蛋白PLIN基因多态性与代谢综合征易感性的关联性研究

目的研究脂滴包被蛋白PUN基因多态性与代谢综合征的关系。方法从河南省信阳市选取代谢综合征及对照共2068例，病例组1021例，对照组1047例。所有研究对象记录其病史、体检等临床资料及吸烟、饮酒等流行病学资料，采用聚合酶链式反应．限制性片段长度多态性（PCR—RFLP）技术检测各组PLIN基因rs8179078和rs6496589两个SNPs位点的多态基因型并统计各组的多态性频率。结果rs6496589在女性中CC基因型的频率病例组明显高于对照组（P〈0．01）。与对照组比较，代谢综合征组rs6496589cc基因型的频率显著高于对照组（代谢综合征组：rs6496589CC为56．6％；对照组rs6496589CC为48．8％；P=0．002）。校正了年龄、性别、高血压史、糖尿病史和高脂血症史等传统危险因素的影响后，这种相关性仍然存在。rs8179078的等位基因和基因型频率在病例组和对照组的分布无显著的统计学差异。结论PUN基因rs6496589多态性中CC基因型是中国汉族人群代谢综合征发病的独立危险因素之一，rs8179078多态性与中国汉族人群的代谢综合征发病无显著相关性。     

Perilipin2 inhibits diabetic nephropathy-induced podocyte apoptosis by activating the PPARγ signaling pathway

Podocyte apoptosis plays a pivotal role in the pathogenesis of diabetic nephropathy (DN). The main purpose of this study was to investigate the effects of perilipin2 on high glucose (HG)-induced podocyte apoptosis and associated mechanisms. Differentially expressed genes (DEGs) in BTBR ob/ob mice vs. nondiabetic mice kidneys were obtained from GSE106841 dataset and picked out using the ‘limma’ package. The protein-protein interaction (PPI) network was constructed using the Search Tool for the Retrieval of Interacting Genes (STRING) and was visualized by Cytoscape. Perilipin2 was a hub gene using the cytoHubba plug-in from Cytoscape. Gene ontology (GO) analysis revealed that the 126 overlapping DEGs were mainly enriched in ‘oxidation reduction’ [biological process, (BP)], metal ion binding’ [molecular function, (MF)] and ‘extracellular region’ [cellular component, (CC)]. KEGG pathway analysis revealed that perilipin2 was mainly involved in ‘PPAR signaling pathway’. DN inhibited perilipin2 expression and PPARγ expression, as by both in vitro and in vivo studies. In vitro experiments demonstrated that perilipin2 inhibition could not only reduced PPARγ expression in podocytes, it could also promote the apoptosis, and inhibit the viability in HG treated podocytes using western blot, CCK8 and flow cytometry assays. Perilipin2 overexpression reversed the effects of HG on inhibiting podocalyxin, nephrin, precursor (pro)-caspase-3/-9 and PPARγ protein expression and increasing cleaved caspase-3/-9 protein expression. Furthermore, the functions of perilipin2 overexpression reversing HG-induced podocyte apoptosis were inhibited by PPARγ inhibitor. In conclusion, the functions of DN-induced podocyte apoptosis were inhibited by activation of the PPARγ signaling pathway caused by perilipin2 overexpression.     

Do estrogens regulate lipid status in testicular steroidogenic Leydig cell?

In this study mouse Leydig cell (MA-10) were treated with G-protein coupled membrane estrogen receptor antagonist (G-15; 10 nM). Cells were analyzed by Western blotting for expression of estrogen-related receptors (ERRα, β and γ), steroidogenic markers (lutropin receptor; LHR and 3β-hydroxysteroid dehydrogenase; 3β-HSD) and lipid droplet markers (perilipin; PLIN and microtubule-associated protein 1 A/1B-light chain 3; LC3). Concomitantly, microscopic analyses by light microscope (immunofluorescent staining for lipid droplets, PLIN and LC3) as well as by electron microscope (for lipid droplet ultrastructure) were utilized. For analysis of cholesterol content, cAMP level and progesterone secretion, G-15, estrogen receptor (ER) antagonist (ICI 182,780; 10 μM), 17β-estradiol (10 mM) and, bisphenol A (BPA; 10 nM) were used alone or in combinations.We revealed no changes in ERRs expression but alterations in ERRβ and γ localization in G-15-treated cells when compared to control. Partial translocation of ERRβ and γ from the cell nucleus to cytoplasm was observed. Decreased expression of LHR, 3β-HSD, PLIN and LC3 was detected. Moreover, in treated cells large lipid droplets and differences in their distribution were found. Very strong signal of co-localization for PLIN and LC3 was found in treated cells when compared to control. In ultrastructure of treated cells, degenerating lipid droplets and double membrane indicating on presence of lipophagosome were observed. We found, that only (i) BPA and G-15 did not effect on cholesterol content, (ii) BPA, G-15 and ICI did not effect on cAMP level and (iii) BPA, ICI alone and in combination, and BPA with G-15 did not modulate progesterone secretion.These findings showed complex and diverse estrogen effects on mouse Leydig cells at various steps of steroid hormone production (cholesterol storage, release and processing). Lipid homeostasis and metabolism in these cells were affected by endogenous and exogenous estrogen, interactions of receptors (GPER, ER and ERR) and GPER and ER antagonists.     

Intragenic linkage disequilibrium structure of the human perilipin gene (PLIN) and haplotype association with increased obesity risk in a multiethnic Asian population

Perilipin is a lipid droplet surface protein present in adipocytes and steroidogenic cells. We examined five common single nucleotide polymorphisms (SNPs) at the perilipin (PLIN) locus (PLIN 6209C>T, 10171A>T, 11482G>A, 13041A>G, and 14995A>T) to investigate their association with obesity risk. The study population included 4,131 subjects of three ethnic groups (Chinese, Malay, and Indian) from Singapore. The prevalence of obesity in Malays and Indians was much higher than in Chinese. Moreover, in these groups the prevalence of obesity was three times higher in women than in men. Crude analysis indicated that haplotype 11212 (CAAAT) is shared by Malays and Indians and is significantly associated with increased obesity risk as compared to the most common haplotype 21111 (TAGAA): OR 1.65 (95% CI 1.11–2.46) in Malays and 1.94 (95% CI 1.06–3.53) in Indians. No associations between PLIN haplotypes and obesity risk were found in Chinese. To simplify the haplotype analyses we used a subgroup of three SNPs (11482G>A, 13041A>G, and 14995A>T) in positive linkage disequilibrium. These analyses revealed similar associations, showing that haplotypes XX212 (XXAAT) and XX222 (XXAGT) are associated with increased obesity risk in Malays OR 2.04 (95% CI 1.28–3.25) and 2.05 (95% CI 1.35–3.12) respectively, and that haplotype XXX212 (XXAAT) is significantly associated with increased obesity risk in Indians OR 2.16 (95% CI 1.10–4.26) after adjusting for covariates including age, sex, smoking, alcohol consumption, exercise, and diabetes status. Moreover, individual SNP analyses demonstrated that the PLIN 14995A>T SNP is the most informative single genetic marker for the observed haplotype association, being significantly associated with increased obesity risk in both Malays OR 2.28 (95% CI 1.45–3.57) and Indians OR 2.04 (95% CI 1.08–3.64). These results support the role of the PLIN locus as an ethnically dependent modulator of obesity risk in humans.     

Perilipin‐1 in Hemodialyzed Patients: Association With History of Coronary Heart Disease and Lipid Profile

Perilipin‐1 surrounds lipid droplets in both adipocytes and in atheroma plaque foam cells and controls access of lipases to the lipid core. In hemodialysis (HD) patients, dyslipidemia, malnutrition, inflammation and atherosclerosis are common. Thirty‐six HD patients and 28 healthy volunteers were enrolled into the study. Ten HD patients suffered from coronary heart disease (CHD). Perilipin‐1, triglycerides, total cholesterol, low‐density lipoprotein cholesterol, high‐density lipoprotein cholesterol (HDL‐C), body mass index, albumin, geriatric nutritional risk index, normalized protein catabolic rate, interleukin‐6 (IL‐6) and tumor necrosis factor‐α (TNF‐α) were measured. Perilipin‐1 did not differ between HD patients and healthy volunteers. IL‐6 and TNF‐α were higher in HD patients. The evaluated nutritional markers and the markers of inflammation did not differ between HD patients with high perilipin‐1 levels and HD patients with low perilipin‐1 levels. Regarding the lipid profile, only HDL‐C differed between HD patients with high perilipin‐1 levels and HD patients with low perilipin‐1 levels, and it was higher in the first subgroup. Perilipin‐1 was significantly higher in HD patients without CHD. Perilipin‐1 is detectable in the serum of HD patients and it is associated with increased HDL‐C and decreased incidence of CHD.