Pattern of expression of adiponectin receptors in human adipose tissue depots and its relation to the metabolic state

OBJECTIVE: To investigate whether adiponectin receptor genes (AdipoR1 and AdipoR2) expression in human subcutaneous (SAT) and visceral (VAT) adipose tissue in severely obese patients with or without diabetes is related to adiponectin gene (APM1) expression and in vivo metabolic parameters. DESIGN: Cross-sectional, clinical research study. SUBJECTS: Total RNA was extracted from SAT and VAT tissue obtained during surgery from 13 lean controls, 30 obese diabetic patients, 19 obese glucose-intolerant patients and 54 obese subjects with normal glucose tolerance. MEASUREMENTS: Tissue expression of APM1, AdipoR1 and AdipoR2, tissue concentration of adiponectin (ApN), and metabolic variables. RESULTS: APM1 expression was higher in SAT than VAT (1.06+/-0.76 vs 0.69+/-0.52, P<0.0001) as was AdipoR1 (1.17+/-0.70 vs 0.66+/-0.38, P<0.0001) and AdipoR2 (7.02+/-6.19 vs 0.75+/-0.64, P<0.0001). In SAT, APM1 and AdipoR1 expression tended to be lower - by 0.38+/-0.22 and 0.35+/-0.22, respectively - and AdipoR2 expression was markedly depressed - by 4.82+/-1.93 - in association with obesity, whereas presence of diabetes had no additional effect. In VAT, APM1 and AdipoR1 expressions were also reduced - by 0.36+/-0.16 and 0.30+/-0.11, respectively - in association with obesity. Within both SAT and VAT, expression levels of APM1, AdipoR1 and AdipoR2 were all positively interrelated. Tissue ApN concentrations in SAT were similar across groups, whereas ApN levels in VAT were substantially lower in association with obesity (by an average of 63+/-12 ng/mg total protein, P<0.0001). In multivariate models adjusting for sex, age and obesity, serum triglyceride concentrations were reciprocally related to APM1 (r=-0.27, P<0.02), AdipoR1 (r=-0.37, P<0.002 and AdipoR2 expression (r=-0.37, P<0.002) in VAT. Likewise, plasma insulin concentrations were inversely related only to APM1 in VAT (r=-0.25, P<0.03). CONCLUSIONS: Severe obesity is associated with suppressed expression of both ApN and its receptors in both SAT and VAT, the expression levels in VAT are specifically linked with hyperinsulinemia and dyslipidemia.     

Cardiomyogenic differentiation potential of human adipose precursor cells

DNA demethylation agent 5-azacytidine has been widely described in literature as an effective chemical stimulus used to promote cardiomyogenic differentiation in various cell types, ranging from embryonic stem cells, P19 cells, bone marrow-derived mesenchymal stem cells, and recently to adipose-derived stem cells. The purpose of this study was to examine the effects of 5-azacytidine on human adipose precursor cell differentiation along the cardiomyogenic lineage.     

Normal and malignant human plasma cells: proliferation, differentiation, and expansions in relation to CD45 expression

In this study, we have evaluated the proliferation and the phenotype of human plasma cells of different origins, i.e., from tonsil, peripheral blood, bone marrow as well as plasma cells generated in vitro from memory B cells. We have demonstrated that plasma cells from tonsil, peripheral blood, as well as those generated in vitro, were highly proliferating and presented a homogeneous CD45bright phenotype. In contrast, bone marrow plasma cells were heterogeneous for CD45 expression but their proliferation was restricted to the CD45bright compartment. Subsequently, their CD45 expression decreased with proliferation arrest and final maturation. We also studied the proliferation of abnormal plasma cells, i.e., peripheral blood reactive plasmacytoses and multiple myeloma (MM). All reactive plasmacytoses turned out to be homogeneous expansions of CD45bright plasma cells with unusually high labeling index. In contrast, CD45 expression was heterogeneous in MM as in normal bone marrow. However, a minor CD45bright population was also always the most proliferating one as opposed to a major population of less or non-proliferating myeloma cells characterized by a weaker or a lack of CD45 expression. In conclusion, proliferation is linked to plasma-cell generation and a CD45bright phenotype is the hallmark of the most proliferating normal, reactive as well as malignant plasma cells.     

Doxorubicin toxicity in relation to the proliferative state of human hematopoietic cells

The relation between the proliferative state of normal human hematopoietic cells and their sensitivity to doxorubicin was studied. T-lymphocytes were stimulated with phytohaemagglutinin/interleukin 2 before or after a 2-h exposure to doxorubicin (range 0-2 microgram/ml). The doxorubicin concentration that inhibited DNA synthesis in 50% of the lymphocytes, measured qualitatively with 5-iodo-2'-deoxyuridine incorporation, was significantly lower (a factor of 2.5) in case of drug exposure of stimulated lymphocytes compared to nonstimulated lymphocytes. These proliferation-dependent differences were not related to differences in cellular drug concentrations, as was determined with flow cytometry. Bone marrow cells were stimulated for 2 days with human placenta-conditioned medium before or after exposure to doxorubicin (range 0-2 microgram/ml), after which they were cultured in a bone marrow clonogenic assay. In analogy with the lymphocyte experiments, proliferation-dependent differences in drug sensitivity were found. The drug concentration that inhibited the growth of granulocyte-macrophage colonies (granulocyte-macrophage colony-forming units, CFU-GM) to 50% appeared significantly lower (a factor of 3.4) with drug exposure of stimulated bone marrow cells compared to nonstimulated bone marrow cells. The relative insensitivity of quiescent, but potentially proliferative cells to doxorubicin might explain the recovery of hematopoiesis after doxorubicin-induced bone marrow hypoplasia.     

Hypoxia increases leptin expression in human PAZ6 adipose cells

AIMS/HYPOTHESIS: Leptin, an adipose tissue-derived cytokine involved in the control of body weight, also participates in a variety of biological functions, including angiogenesis. Because reduced oxygen availability is a major inducer of angiogenesis, we hypothesized that low cellular oxygen tension could regulate leptin expression in adipose cells. METHODS: Differentiated PAZ6 adipocytes were cultured for 48 h in the presence of chemical inducers of cellular hypoxia (cobalt chloride or desferrioxamine) or in an atmosphere containing only 6% oxygen. The effect of hypoxia on the expression of leptin and several adipose genes was assessed by semi-quantitative RT-PCR. The effect of hypoxia on leptin promoter activity was tested in PAZ6 cells transiently transfected with a luciferase reporter construct, containing 1.87 kb of the human leptin promoter. Leptin secretion in the culture medium was determined by radioimmunoassay. RESULTS: Hypoxia increased leptin mRNA expression, leptin promoter activity and leptin secretion in the culture medium by two- to threefold ( p<0.05). The expression of the glucose transporter isoform 1 (GLUT-1) mRNA, a well known hypoxia inducible gene, was also increased. In contrast, glucose transporter isoform 4 (GLUT-4), hormone sensitive lipase (HSL), fatty acid binding protein (aP2) and uncoupling protein 2 (UCP2) mRNAs were markedly reduced by hypoxia. In addition, a similar hypoxia-induced increase in leptin mRNA and secretion was observed in primary rat adipose cells. CONCLUSION/INTERPRETATION: Hypoxia markedly and specifically increased leptin gene expression through activation of the leptin gene promoter, and this resulted in an increased leptin production by human PAZ6 adipocytes.     

脂肪间充质干细胞诱导向肝系细胞分化

大多数肝脏疾病终末期并发严重肝功能障碍，肝移植是治愈的唯一办法。但是临床可供移植肝来源短缺、移植伦理以及移植后排异问题，限制了肝移植在临床的广泛应用。细胞疗法的出现为肝脏疾病治疗提供了另一种可选途径。干细胞替代治疗或者刺激体内肝脏细胞再生是细胞治疗的主要目标。哺乳动物肝脏可以自身修复或再生，卵圆细胞在肝脏修复过程中扮演重要作用。他们可以向肝实质细胞以及向胆管细胞分化。     

人脂肪干细胞向内皮分化最佳诱导体系的建立

目的 探讨人脂肪干细胞(human Adipose-derived stem cells,hADSCs)体外分化为内皮细胞的最佳诱导体系.方法 利用胶原酶消化法和贴壁筛选法从人脂肪组织中分离、培养及扩增hADSCs,分3组分别进行内皮诱导:即纤维连接蛋白(fibronectin,FN)包被组、明胶包被组和未铺组;同时诱导液分为两组:单纯内皮细胞生长培养基(EGM2-MV)组及内皮细胞生长培养基(EGM2-MV)+50 ng/ml血管内皮细胞生长因子(VEGF_(165))组.诱导15 d后,通过流式细胞术检测各组内皮细胞特异性表面抗原CD34的表达;通过相差显微镜观察诱导前后细胞的一般形态学特征.结果 体外分离、培养出高度同源性的hADSCs,CD29、CD44、CD90、CD105及CD166呈阳性表达,而CD34、CD45及HLA-DR呈阴性表达;诱导后细胞流式结果显示单纯EGM-2MV诱导的3组细胞内皮细胞特异性标志CD34阴性表达,而EGM2-MV+50 ng/ml VEGF_(165)诱导的细胞,其中FN包被组CD34表达率为8.4%,明胶包被组为5.4%,未铺组为4.2%;相差显微镜下可见诱导后细胞呈三角形或多边形,融合处呈铺路石样生长.结论 FN作为细胞外基质与EGM2-MV+50 ng/ml VEGF_(165)组成的诱导液协同作用,构成了hADSCs体外分化为内皮细胞的最佳诱导体系,可为临床上缺血性疾病的自体细胞移植治疗提供大量的种子细胞来源.     

PD-1 expression on human CD8 T cells depends on both state of differentiation and activation status

OBJECTIVE AND DESIGN: PD-1 expression on HIV-specific CD8 T cells was recently reported to reflect functional exhaustion, resulting in uncontrolled HIV-1 replication. Assessing PD-1 expression on T cells may be highly relevant in T-cell immunology and vaccine monitoring. However, this requires us to gain further insights into the significance of PD-1 expression on CD8 T cells in humans. METHODS: We performed a detailed analysis of PD-1 expression pattern on various CD8 T cell subsets from healthy or HIV infected donors. RESULTS: PD-1 expression has two facets in vivo. On the one hand, it is linked to T-cell differentiation: PD-1 is up-regulated on early/intermediate differentiated subsets, which include HIV and Epstein-Barr virus-specific CD8 T-cell populations, but is down-regulated during late stages of differentiation. On the other hand, it is linked to T-cell activation: on PD-1 positive cells, PD-1 over-expression occurs along with the up-regulation of activation markers such as CD38 or HLA-DR. CONCLUSIONS: PD-1 expression on CD8 T cells, including those specific for HIV, can be related both to their differentiation stage and their activation status. It is important to consider these findings when assessing the expression of PD-1 on T cells.     

Characteristic expression of γ-aminobutyric acid and glutamate decarboxylase in rat jejunum and its relation to differentiation of epithelial cells

AIM: To investigate the expression between γ-aminobutyric acid (GABA) and glutamate decarboxylase and its relation with differentiation and maturation of jejunal epithelial cells in rat jejunum.METHODS: Immunohistochemical expression of GABA and glutamate decarboxylase (GAD, including two isoforms,GAD65 and GAD67) was investigated in rat jejunum.Meanwhile, double staining was performed with GAD65 immunohistochemistry, followed by lectin histochemistry of fluorescent wheat germ agglutinin. Furthermore,evaluation of cell kinetics in jejunum was conducted by ^3Hthymidine autoradiography and immunohistochemistry using a monoclonal antibody to proliferating cell nuclear antigen (PCNA).RESULTS: The cells showing positive immunoreactivity GABA and GAD65 were mainly distributed in the villi in rat jejunum, while jejunal epithelial cells were negative for GAD67. Positive GABA or GAD65 staining was mainly located in the cytoplasm and along the brush border of epithelial cells in the middle and upper portions. In addition, a few GABA and GAD65 strongly positive cells were scattered in the upper two thirds of jejunal villi. Double staining showed that GAD65 immunoreactivity was not found in goblet cells.^3H-thymidine-labeled nuclei were found in the lower and middle portions of jejunal crypts, which was consistent with PCNA staining. Therefore, GABA and GAD65 were expressed in a maturation or functional zone.CONCLUSION: The characteristic expression of GABA and GAD suggests that GABA might be involved in regulation of differentiation and maturation of epithelial cells in rat jejunum.     

Expression of fatty-acid-handling proteins in human adipose tissue in relation to obesity and insulin resistance

Aims/hypothesis Protein-mediated trans-membrane and intracellular fatty acid trafficking are becoming increasingly recognised as biochemically and physiologically important concepts. Obesity and insulin resistance are polygenic disorders, heavily influenced by environmental and life-style factors, and are virtually always associated with disturbed fatty acid metabolism in adipose and other tissues. The aim of this study was to investigate mRNA expression levels of fatty-acid-handling proteins in adipose tissue in relation to markers of genetic and acquired obesity and insulin resistance.Methods We quantified mRNA expression of subcutaneous adipose tissue fatty-acid-handling proteins (ALBP, KLBP, FATP1, FATP4, CD36, ACS1) in 17 monozygotic twin-pairs with a range of intra-pair differences () in BMI and detailed measures of obesity and insulin resistance, allowing influences of genetic and non-genetic factors to be distinguished.Results In acquired obesity FATP4 expression was up-regulated independently of genetic background (FATP4 versus BMI; r=0.50, p=0.04; FATP4 versus body fat; r=0.59, p=0.01). Similarly, CD36 and FATP1 expression correlated with acquired differences in HDL cholesterol and non-esterified fatty acid concentrations respectively. Moreover, FATP4 and CD36 expression levels correlated with measures of obesity and insulin resistance that are influenced by both genetic and non-genetic factors (FATP4 versus BMI: r=0.53, p=0.0001; FATP4 versus body fat: r=0.51, p=0.002; FATP4 versus homeostasis model assessment [HOMA]: r=0.49, p=0.001; CD36 versus BMI: r=0.50, p=0.02; CD36 versus body fat: r=0.63, p=0.001; CD36 versus HOMA: r=0.34, p=0.06).Conclusions/interpretation These findings indicate that expression of specific adipose tissue fatty-acid-handling proteins is related to obesity and insulin resistance, and that, in particular, FATP4 plays a role in acquired obesity. Our results suggest that facilitated fatty acid trafficking is a physiologically and pathologically relevant phenomenon in man.Abbreviations ACS acyl-CoA synthase - ALBP adipocyte lipid-binding protein - FABP fatty-acid-binding protein - FATP fatty acid transport protein - HOMA homeostasis model assessment - KLBP keratinocyte lipid-binding protein - MZ monozygotic - TBP TATA-box binding protein - TG triglycerideThis revised version was published online in June 2004 with corrections to title and author addresses.     

Hepatogenic differentiation of human mesenchymal stem cells from adipose tissue in comparison with bone marrow mesenchymal stem cells

INTRODUCTIONMost liver diseases lead to hepatocyte dysfunction with the possibility of eventual organ failure. The replacement of diseased hepatocytes and the stimulation of endogenous or exogenous regeneration by stem cells are the main aims of liver-dir…     

Spontaneous cardiomyocyte differentiation from adipose tissue stroma cells

Cardiomyocyte regeneration is limited in adult life. Thus, the identification of a putative source of cardiomyocyte progenitors is of great interest to provide a usable model in vitro and new perspective in regenerative therapy. As adipose tissues were recently demonstrated to contain pluripotent stem cells, the emergence of cardiomyocyte phenotype from adipose-derived cells was investigated. We demonstrated that rare beating cells with cardiomyocyte features could be identified after culture of adipose stroma cells without addition of 5-azacytidine. The cardiomyocyte phenotype was first identified by morphological observation, confirmed with expression of specific cardiac markers, immunocytochemistry staining, and ultrastructural analysis, revealing the presence of ventricle- and atrial-like cells. Electrophysiological studies performed on early culture revealed a pacemaker activity of the cells. Finally, functional studies showed that adrenergic agonist stimulated the beating rate whereas cholinergic agonist decreased it. Taken together, this study demonstrated that functional cardiomyocyte-like cells could be directly obtained from adipose tissue. According to the large amount of this tissue in adult mammal, it could represent a useful source of cardiomyocyte progenitors.     

Ultrastructural differentiation and CEA expression of butyrate-treated human pancreatic carcinoma cells

The effects of butyrate (a biological response modifier) on cellular morphologic features and carcinoembryonic antigen (CEA) expression of human pancreatic carcinoma cells were studied and compared in a well-differentiated, CEA-producing cell line (CAPAN-1), and a poorly differentiated cell line (PANC-1). Butyrate treatment resulted in the acquisition of phenotypic traits commonly attributed to increased "differentiation," including a twofold increase in doubling time, decreased saturation densities, and approximately 50% reduction in colony forming efficiency in both cell lines. Elongation and flattening of cells with extending cellular processes were seen by light microscopy. Significant ultrastructural changes were seen only in the PANC-1 cells, including an increased number of intercellular desmosomes, tonofilaments, and lipid droplets. In contrast, to the coarsely clumped nuclear chromatin (heterochromatin) of untreated PANC-1 cells, the nuclei of the butyrate-treated cells consisted of finely dispersed chromatin (euchromatin). CAPAN-1 cells responded to butyrate with increased CEA synthesis and release. This effect was greatest in the stationary growth phase. Butyrate had no effect on the already low rate of CEA synthesis by PANC-1 cells. These studies suggest that CEA synthesis and state of differentiation are affected independently by butyrate treatment and that the original tumor phenotype plays an important role in response to such treatment.     

Characteristic expression of γ-aminobutyric acid and glutamate decarboxylase in rat jejunum and its relation to differentiation of epithelial cells

AIM: To investigate the expression between γ-aminobutyric acid (GABA) and glutamate decarboxylase and its relation with differentiation and maturation of jejunal epithelial cells in rat jejunum.METHODS: Immunohistochemical expression of GABA and glutamate decarboxylase (GAD, including two isoforms,GAD65 and GAD67) was investigated in rat jejunum.Meanwhile, double staining was performed with GAD65 immunohistochemistry, followed by lectin histochemistry of fluorescent wheat germ agglutinin. Furthermore,evaluation of cell kinetics in jejunum was conducted by 3Hthymidine autoradiography and immunohistochemistry using a monoclonal antibody to proliferating cell nuclear antigen (PCNA).RESULTS: The cells showing positive immunoreactivity GABA and GAD65 were mainly distributed in the villi in rat jejunum, while jejunal epithelial cells were negative for GAD67. Positive GABA or GAD65 staining was mainly located in the cytoplasm and along the brush border of epithelial cells in the middle and upper portions. In addition, a few GABA and GAD65 strongly positive cells were scattered in the upper two thirds of jejunal villi. Double staining showed that GAD65 immunoreactivity was not found in goblet cells.3H-thymidine-labeled nuclei were found in the lower and middle portions of jejunal crypts, which was consistent with PCNA staining. Therefore, GABA and GAD65 were expressed in a maturation or functional zone.CONCLUSION: The characteristic expression of GABA and GAD suggests that GABA might be involved in regulation of differentiation and maturation of epithelial cells in rat jejunum.     

Growth hormone and adipose differentiation: growth hormone-induced antimitogenic state in 3T3-F442A preadipose cells.

An additional activity for pituitary growth hormone is described--i.e., the in vitro induction of an antimitogenic state in murine 3T3-F442A preadipocyte fibroblasts. We previously developed a serum-free, hormonally defined medium permissive for the adipose differentiation of 3T3-F442A cells. When 3T3-F442A fibroblasts were maintained in serum-free medium without insulin but with growth hormone (2 nM), typical adipose differentiation did not occur. However, we found that growth hormone induced a state of cellular refractoriness to the mitogenic stimulus of fetal bovine serum as assayed by de novo DNA synthesis. The mitogen refractory condition (i.e., the antimitogenic state) was time-dependent (half maximal at approximately 2.5 days) and growth hormone concentration-dependent (half maximal and maximal at approximately 0.05 and 2.0 nM, respectively). The antimitogenic state was specifically induced by growth hormone and was not mediated by insulin-like growth factor I or prolactin. The growth hormone-induced antimitogenic state was completely reversible. The antimitogenic state was not induced by growth hormone in 3T3-C2 cells, a sister clone of 3T3 cells that exhibits essentially no adipose conversion. The kinetics for growth hormone-dependent commitment to adipose differentiation and induction of the antimitogenic state were similar. We suggest a relationship of growth hormone-induced antimitogenic state and the growth hormone-induced adipose differentiation of 3T3-F442A cells.     

脂肪分化及其相关进展

脂肪组织不仅是体内最大的能量储存器官,而且作为内分泌器官产生多种脂肪因子,参与重要的生理和病理过程,包括食欲、血压、生殖、血管发生、胰岛素敏感性、炎症等诸多环节.