Angiopoietin-like protein 2 promotes chondrogenic differentiation during bone growth as a cartilage matrix factor.

OBJECTIVE: Chondrocyte differentiation is crucial for long bone growth. Many cartilage extracellular matrix (ECM) proteins reportedly contribute to chondrocyte differentiation, indicating that mechanisms underlying chondrocyte differentiation are likely more complex than previously appreciated. Angiopoietin-like protein 2 (ANGPTL2) is a secreted factor normally abundantly produced in mesenchymal lineage cells such as adipocytes and fibroblasts, but its loss contributes to the pathogenesis of lifestyle- or aging-related diseases. However, the function of ANGPTL2 in chondrocytes, which are also differentiated from mesenchymal stem cells, remains unclear. Here, we investigate whether ANGPTL2 is expressed in or functions in chondrocytes. METHODS: First, we evaluated Angptl2 expression during chondrocyte differentiation using chondrogenic ATDC5 cells and wild-type epiphyseal cartilage of newborn mice. We next assessed ANGPTL2 function in chondrogenic differentiation and associated signaling using Angptl2 knockdown ATDC5 cells and Angptl2 knockout mice. RESULTS: ANGPTL2 is expressed in chondrocytes, particularly those located in resting and proliferative zones, and accumulates in ECM surrounding chondrocytes. Interestingly, long bone growth was retarded in Angptl2 knockout mice from neonatal to adult stages via attenuation of chondrocyte differentiation. Both in vivo and in vitro experiments show that changes in ANGPTL2 expression can also alter p38 mitogen-activated protein kinase (MAPK) activity mediated by integrin α5ß1. CONCLUSION: ANGPTL2 contributes to chondrocyte differentiation and subsequent endochondral ossification through α5ß1 integrin and p38 MAPK signaling during bone growth. Our findings provide insight into molecular mechanisms governing communication between chondrocytes and surrounding ECM components in bone growth activities.

Angiopoietin-like protein 2 sensitively responds to weight reduction induced by lifestyle intervention on overweight Japanese men.

OBJECTIVE: Overexpression of Angiopoietin-like protein 2 (Angptl2) in obese adipose tissues promotes adipose tissue inflammation and its-related metabolic abnormalities. In a comparative study with adiponectin, we investigated whether alterations in serum Angptl2 concentrations reflect the effect of lifestyle intervention on weight loss and improved metabolic parameters in overweight subjects. METHODS: A total of 154 Japanese men (age, 40.9±5.1 years; body mass index, 26.9±3.6 kg m(-2); abdominal circumference, 94.1±8.9 cm) underwent a 3-month lifestyle intervention and underwent follow-up for 3 months thereafter. RESULTS: Decreased serum Angptl2 levels, but not increased serum adiponectin levels, were immediately apparent at the end of 3-month lifestyle intervention. Angptl2 levels continued to decrease for 3 months in parallel with body weight loss and improvement in metabolic indicators. In subjects showing 6% weight reduction, markedly reduced Angptl2 levels were detected at the end of 3-month intervention, whereas increased adiponectin levels were detected 3 months after the end of intervention. Multivariate analysis revealed changes in serum Angptl2 levels associated with changes in triglycerides (TGs), aspartate aminotransferase and alanine aminotransferase. In contrast, changes in serum adiponectin levels were associated with altered high-density lipoprotein cholesterol (HDL-C) and fasting plasma glucose levels. CONCLUSION: A 3-month lifestyle intervention promoted weight reduction and improved glucose and lipid metabolism, an effect maintained 3 months later. Notably, our findings indicate that decreased Angptl2 levels are a good indicator of reduced visceral fat and metabolic improvement at early stages of lifestyle intervention. Thus, Angptl2 reflects adiposity and might be a key protein to regulate inflammation and TG metabolism, whereas adiponectin levels could reflect improved glucose and HDL-C metabolism.

Tumour necrosis factor alpha signalling through activation of Kupffer cells plays an essential role in liver fibrosis of non-alcoholic steatohepatitis in mice.

BACKGROUND: While tumour necrosis factor alpha (TNF-alpha) appears to be associated with the development of non-alcoholic steatohepatitis (NASH), its precise role in the pathogenesis of NASH is not well understood. METHODS: Male mice deficient in both TNF receptors 1 (TNFR1) and 2 (TNFR2) (TNFRDKO mice) and wild-type mice were fed a methionine and choline deficient (MCD) diet or a control diet for eight weeks, maintaining isoenergetic intake. RESULTS: MCD dietary feeding of TNFRDKO mice for eight weeks resulted in attenuated liver steatosis and fibrosis compared with control wild-type mice. In the liver, the number of activated hepatic Kupffer cells recruited was significantly decreased in TNFRDKO mice after MCD dietary feeding. In addition, hepatic induction of TNF-alpha, vascular cell adhesion molecule 1, and intracellular adhesion molecule 1 was significantly suppressed in TNFRDKO mice. While in control animals MCD dietary feeding dramatically increased mRNA expression of tissue inhibitor of metalloproteinase 1 (TIMP-1) in both whole liver and hepatic stellate cells, concomitant with enhanced activation of hepatic stellate cells, both factors were significantly lower in TNFRDKO mice. In primary cultures, TNF-alpha administration enhanced TIMP-1 mRNA expression in activated hepatic stellate cells and suppressed apoptotic induction in activated hepatic stellate cells. Inhibition of TNF induced TIMP-1 upregulation by TIMP-1 specific siRNA reversed the apoptotic suppression seen in hepatic stellate cells. CONCLUSIONS: Enhancement of the TNF-alpha/TNFR mediated signalling pathway via activation of Kupffer cells in an autocrine or paracrine manner may be critically involved in the pathogenesis of liver fibrosis in this NASH animal model.

Stromal cells expressing ephrin-B2 promote the growth and sprouting of ephrin-B2(+) endothelial cells.

Ephrin-B2 is a transmembrane ligand that is specifically expressed on arterial endothelial cells (ECs) and surrounding cells and interacts with multiple EphB class receptors. Conversely, EphB4, a specific receptor for ephrin-B2, is expressed on venous ECs, and both ephrin-B2 and EphB4 play essential roles in vascular development. The bidirectional signals between EphB4 and ephrin-B2 are thought to be specific for the interaction between arteries and veins and to regulate cell mixing and the making of particular boundaries. However, the molecular mechanism during vasculogenesis and angiogenesis remains unclear. Manipulative functional studies were performed on these proteins in an endothelial cell system. Using in vitro stromal cells (OP9 cells) and a paraaortic splanchnopleura (P-Sp) coculture system, these studies found that the stromal cells expressing ephrin-B2 promoted vascular network formation and ephrin-B2(+) EC proliferation and that they also induced the recruitment and proliferation of alpha-smooth muscle actin (alpha-SMA)-positive cells. Stromal cells expressing EphB4 inhibited vascular network formation, ephrin-B2(+) EC proliferation, and alpha-SMA(+) cell recruitment and proliferation. Thus, these data suggest that ephrin-B2 and EphB4 mediate reciprocal interactions between arterial and venous ECs and surrounding cells to form each characteristic vessel. (Blood. 2001;98:1028-1037)

Mobilization of endothelial progenitor cells in patients with acute myocardial infarction.

BACKGROUND: Endothelial progenitor cells (EPCs) circulate in adult peripheral blood (PB) and contribute to neovascularization. However, little is known regarding whether EPCs and their putative precursor, CD34-positive mononuclear cells (MNC(CD34+)), are mobilized into PB in acute ischemic events in humans. METHODS AND RESULTS: Flow cytometry revealed that circulating MNC(CD34+) counts significantly increased in patients with acute myocardial infarction (n=16), peaking on day 7 after onset, whereas they were unchanged in control subjects (n=8) who had no evidence of cardiac ischemia. During culture, PB-MNCs formed multiple cell clusters, and EPC-like attaching cells with endothelial cell lineage markers (CD31, vascular endothelial cadherin, and kinase insert domain receptor) sprouted from clusters. In patients with acute myocardial infarction, more cell clusters and EPCs developed from cultured PB-MNCs obtained on day 7 than those on day 1. Plasma levels of vascular endothelial growth factor significantly increased, peaking on day 7, and they positively correlated with circulating MNC(CD34+) counts (r=0.35, P=0.01). CONCLUSIONS: This is the first clinical demonstration showing that lineage-committed EPCs and MNC(CD34+), their putative precursors, are mobilized during an acute ischemic event in humans.

VEGF-C signaling pathways through VEGFR-2 and VEGFR-3 in vasculoangiogenesis and hematopoiesis.

Signaling by vascular endothelial growth factors (VEGFs) through VEGF receptors (VEGFRs) plays important roles in vascular development and hematopoiesis. The authors analyzed the function of VEGF-C signaling through both VEGFR-2 and VEGFR-3 in vasculoangiogenesis and hematopoiesis using a coculture of para-aortic splanchnopleural mesoderm (P-Sp) explants from mouse embryos with stromal cells (OP9). Vasculogenesis and angiogenesis were evaluated by the extent of vascular bed and network formation, respectively. Addition of VEGF-C to the P-Sp culture enhanced vascular bed formation and suppressed definitive hematopoiesis. Both vascular bed and network formations were completely suppressed by addition of soluble VEGFR-1-Fc competitor protein. Formation of vascular beds but not networks could be rescued by VEGF-C in the presence of the competitor, while both were rescued by VEGF-A. VEGFR-3-deficient embryos show the abnormal vasculature and severe anemia. Consistent with these in vivo findings, vascular bed formation in the P-Sp from the VEGFR-3-deficient embryos was enhanced to that in wild-type or heterozygous embryos, and hematopoiesis was severely suppressed. When VEGFR-3-Fc chimeric protein was added to trap endogenous VEGF-C in the P-Sp culture of the VEGFR-3-deficient embryos, vascular bed formation was suppressed and hematopoiesis was partially rescued. These results demonstrate that because VEGF-C signaling through VEGFR-2 works synergistically with VEGF-A, the binding of VEGF-C to VEGFR-3 consequently regulates VEGFR-2 signaling. In VEGFR-3-deficient embryos, an excess of VEGF-C signals through VEGFR-2 induced the disturbance of vasculogenesis and hematopoiesis during embryogenesis. This indicates that elaborated control through VEGFR-3 signaling is critical in vasculoangiogenesis and hematopoiesis. (Blood. 2000;96:3793-3800)

Hematopoiesis and angiogenesis.

Hematopoiesis is closely linked with angiogenesis, because they interact with each other and have common ancestors: hemangioblasts or hematogenic endothelial cells. The relationship is reasonable, because vascular and hematopoietic systems must develop together in order to establish the body's oxygen-delivery system during organogenesis. Hematopoietic stem cells have been shown to originate from the para-aortic splanchnopleural mesoderm region or aorta-gonads-mesonephros at successive stages. We discuss the molecular events in the differentiation of hematopoietic cells and endothelial cells. Transcription factors SCL/tal-1 and AML1; tyrosine kinase receptors Flk-1, Tie-2, Eph, and the sialomucins; CD34; thrombomucin; and AA4 play important biological roles in these lineages.

Mice homozygous for a truncated form of CREB-binding protein exhibit defects in hematopoiesis and vasculo-angiogenesis.

CREB-binding protein (CBP) and the closely related adenovirus E1A-associated 300-kD protein (p300) function as coactivators of transcription factors such as CREB, c-Fos, c-Jun, c-Myb, and several nuclear receptors. To study the roles of CBP in embryonic development, we generated CBP homozygous mutant mouse embryos that expressed a truncated form of CBP protein (1-1084 out of 2441 residues). The embryos died between embryonic days 9.5 (E9.5) and E10.5 and exhibited a defect in neural tube closure. They appeared pale and showed decreases in erythroid cells and colony-forming cells (CFCs) in the yolk sac, suggesting defects in primitive hematopoiesis. Immunohistochemistry with an anti-PECAM antibody showed a lack of vascular network formation. Organ culture of para-aortic splanchnopleural mesoderm (P-Sp) with stromal cells (OP9) showed an autonomous abnormality of putative endothelial precursors, which may induce the microenvironmental defect in hematopoiesis. In addition, these defects were partially rescued by the addition of VEGF to this culture. Our analyses demonstrate that CBP plays an essential role in hematopoiesis and vasculo-angiogenesis.

Truncated CBP protein leads to classical Rubinstein-Taybi syndrome phenotypes in mice: implications for a dominant-negative mechanism.

A mouse model of Rubinstein-Taybi syndrome (RTS) was generated by an insertional mutation into the cyclic AMP response element-binding protein (CREB)-binding protein (CBP) gene. Heterozygous CBP-deficient mice, which had truncated CBP protein (residues 1-1084) containing the CREB-binding domain (residues 462-661), showed clinical features of RTS, such as growth retardation (100%), retarded osseous maturation (100%), hypoplastic maxilla with narrow palate (100%), cardiac anomalies (15%) and skeletal abnormalities (7%). Truncated CBP is considered to have been acting during development as a dominant-negative inhibitor to lead to the phenotypes of RTS in mice. Our studies with step-through-type passive avoidance tests and with fear conditioning test showed that mice were deficient in long-term memory (LTM). In contrast, short-term memory (STM) appeared to be normal. These results implicate a crucial role for CBP in mammalian LTM. Our CBP +/- mice would be an excellent model for the study of the role of CBP in development and memory storage mechanisms.

Granulocyte/macrophage colony-stimulating factor and interleukin-3 correct osteopetrosis in mice with osteopetrosis mutation.

Although young mice homozygous for the osteopetrosis (op) mutation usually developed prominent osteopetrosis, its severity was markedly reduced in aged op/op mice. This age-associated reversal of osteopetrosis was accompanied by the expansion of bone marrow cavities and increased numbers of tartrate-resistant acid phosphatase (TRAP)-positive cells and of macrophages in the bone marrow. The TRAP-positive cells were mononuclear and developed ruffled borders and numerous vesicles, vacuoles, and granules. Enzyme-linked immunosorbent assay demonstrated a significant elevation of serum granulocyte/ macrophage colony-stimulating factor (GM-CSF) and interleukin (IL)-3 levels in the aged op/op mice. To examine whether GM-CSF and/or IL-3 could correct osteopetrosis in young op/op mice, 5 ng of recombinant murine (rm)GM-CSF and/or 100 ng of rmIL-3 were injected daily into young op/op mice. In these treated young op/op mice, the bone marrow cavities were expanded significantly at 2 weeks after administration, associated with significantly increased numbers of TRAP-positive cells and bone marrow macrophages. TRAP-positive cells increased in number with days after injection. These results suggest that GM-CSF and IL-3 induce the development of osteoclasts to correct osteopetrosis in the op/op mice with aging.

A psychobehavioral factor, alexithymia, is related to coronary spasm.

The aim of this study was to assess whether the psychobehavioral pattern alexithymia is related to coronary artery spasm. Alexithymia, deficient psychological awareness, was examined using the Minnesota Multiphasic Personality Inventory Alexithymia Scale in 100 patients with angina pectoris in whom coronary spasm, defined as > or = 99% coronary narrowing, was documented upon ergonovine provocation, and in 109 patients with chest pain syndrome who were shown to have almost normal coronaries without inducible coronary spasm on coronary angiogram (control group). Alexithymia was approximately twice as prevalent in the coronary spasm group (31%) as in the control group (14%) (p<0.01). Among various conventional risk factors including hyperlipidemia, obesity, diabetes mellitus, hypertension, hyperuricemia, or family history of ischemic heart disease, only male sex and smoking were more prevalent in the coronary spasm group than in the control group (p<0.001). The odds ratios of coronary spasm adjusted for all the other risk parameters including sex and age were 4.14 [95% confidence interval (CI) 1.81-9.47] for alexithymia and 2.38 (95, CI 1.18-4.82) for smoking. A psychobehavioral pattern, alexithymia, relates to coronary spasm. This relationship is independent of the conventional coronary risk factors.

The effects of radix Salviae miltiorrhizae on lipid accumulation of peroxidized low density lipoprotein in mouse peritoneal macrophages--lipid analysis and morphological studies.

Mouse peritoneal macrophages were incubated in DMEM with pox-LDL and Rradix Salviae Miltiorrhizae (RSM) to investigate the effects of RSM on the internalization of peroxidized low density lipoprotein (pox-LDL) by using lipid analysis and electron microscopy. Lipid peroxide (LPO) concentrations were increased slightly in the medium after incubation of macrophages with normal LDL (n-LDL), while decreased significantly in the media after incubation of macrophages with pox-LDL. In the three groups with pox-LDL, it could be found that there was a dose-dependent decrease of concentrations of LPO and total cholesterol (TCH) in the two RSM groups, and the decrease in the two RSM groups was much greater than in the group without RSM. RSM accelerated a more decrease of LPO than cholesterol contents in the media containing pox-LDL. The ultrastructural studies also showed that RSM induced the accumulation of lipid droplets in the cytoplasm of mouse peritoneal macrophages. The results suggested that RSM could accelerate the phagocytosis and degradation of pox-LDL by macrophages.

Hyperventilation as a specific test for diagnosis of coronary artery spasm.

The hyperventilation test has been used as a clinical tool to induce coronary spasm. However, its diagnostic and prognostic values have not been fully elucidated. This study was designed to establish the sensitivity and specificity of the hyperventilation test and to clarify the characteristics of hyperventilation test-positive patients. We examined 206 patients in whom coronary spasm was documented by angiography (spasm group), and 183 patients without angina at rest in whom acetylcholine failed to induce spasm (nonspasm group). All patients performed vigorous hyperventilation for 6 minutes in the early morning. Of the spasm group patients, 127 showed positive responses to the test, including ST elevation (n = 111), ST depression (n = 15) and negative U wave (n = 1). None in the nonspasm group showed any ischemic electrocardiographic change. Thus, the sensitivity and specificity of this test for diagnosis of coronary spasm were 62% and 100%, respectively. In the spasm group, there were no significant differences between hyperventilation test-positive and test-negative patients in age, sex, the prevalence of hypertension, diabetes mellitus, obesity, smoking, and the number of diseased vessels. When clinical characteristics were compared, the proportions of the patients with high disease activity (> or =5 attacks a week), with severe arrhythmias (second- or third-degree atrioventricular block and/or ventricular tachycardia) during attacks, and with multivessel spasm were significantly higher in the hyperventilation test-positive patients than in the negative patients (69% vs 20%, p <0.0001; 31% vs 11%, p <0.005; and 58% vs 34%, p <0.01, respectively). These findings imply that hyperventilation is a highly specific test for the diagnosis of coronary artery spasm, and that hyperventilation test-positive patients are likely to have life-threatening arrhythmias during attacks and multivessel spasm.

Efficiency of recombination by Cre transient expression in embryonic stem cells: comparison of various promoters.

The Cre-loxP recombination system of bacteriophage P1 is frequently utilized in genetic manipulation in embryonic stem (ES) cells. The level of Cre expression is critical to induce loxP site-specific recombination in ES cells. To compare the efficiency of recombination, we constructed four Cre expression vectors driven by different promoters: cytomegarovirus/chicken beta-actin (CAG) promoter, human polypeptide chain elongation factor 1alpha (hEF-1alpha) promoter, mouse phosphoglycerate kinase-1 (mPGK) promoter, and polyoma enhancer/herpes simplex virus thymidine kinase (MC1) promoter. We introduced these Cre expression vectors by electroporation into three ES cell lines carrying a single copy of CAG-loxP-chloramphenicol acetyltransferase (CAT) gene-loxP-beta-galactosidase (beta-gal) gene construct. Since the Cre-mediated recombination leads to excision of the CAT gene, the efficiency of recombination can be monitored as beta-gal expression. No selection system was used in the experiments. The maximum recombination frequency was obtained when the CAG promoter was used, followed by the hEF-1alpha promoter, the mPGK promoter and the MC1 promoter in order. These results indicate that the efficiency of recombination in transient expression system correlates with the promoter activity of Cre expression vector. Thus, it is important to choose the promoter for effective recombination by Cre.

Sodium ionophore converts growth manner of vascular smooth muscle cells from spontaneously hypertensive rats.

OBJECTIVES: Vasoconstrictor peptides such as endothelin (ET) cause hypertrophy of vascular smooth muscle cells (VSMC) in Wistar Kyoto rats (WKY) and hyperplasia in spontaneously hypertensive rats (SHR). They also induce an increase in Na+ concentration ([Na+]i) and activate protein kinase C (PKC) independently. Therefore, we tested the hypothesis that the increase in [Na+]i may be involved in the conversion of growth manner under activated PKC in SHR VSMC. METHODS AND RESULTS: 10(-7) M phorbol ester (TPA) increased the diameter and protein content of VSMC from both strains under 18% serum conditions. Further addition of 10(-6) M gramicidin (Na+ ionophore) converted TPA-induced hypertrophy to hyperplasia, which was due to the quick transition from S to G2/M phase, only in SHR VSMC. Western blot analysis showed that serum- and TPA-induced tyrosine phosphorylation of mitogen-activated protein (MAP) kinase was potentiated by 10(-6) M gramicidin in SHR. [Na+]i, which was measured by sodium-binding benzofuran isophthalate (SBFI), was increased about 35 mM by 10(-6) M gramicidin in both strains, but TPA did not affect basal [Na+]i and the gramicidin-induced increase in [Na+]i. CONCLUSIONS: We conclude that sodium ionophore may convert hypertrophy to hyperplasia synergistically with activated PKC in SHR VSMC, possibly by MAP kinase phosphorylation.

Batroxobin accelerates lipid accumulation of peroxidized low density lipoprotein in mouse peritoneal macrophages.

Mouse peritoneal macrophages were incubated in DMEM medium with batroxobin (DF-521) to determine the effect of batroxobin on the internalization of peroxidized low-density lipoprotein (pox-LDL) by transmission electron microscopy. Although the morphology of the mouse peritoneal macrophages after incubation with DMEM, normal LDL (n-LDL) and n-LDL plus batroxobin was similar to that of the cells before incubation, they exhibited numerous cytoplasmic lipid droplets after incubation with pox-LDL for 4 h. After addition of batroxobin to the medium containing pox-LDL, the production of lipid droplets in the mouse peritoneal macrophages was tremendously accelerated. Batroxobin accelerates the phagocytosis and degradation of pox-LDL by macrophages.

Angiotensin converting enzyme as a genetic risk factor for coronary artery spasm. Implication in the pathogenesis of myocardial infarction.

It has been reported that individuals with the D allele of an insertion/deletion (I/D) polymorphism of the angiotensin converting enzyme (ACE) gene are at greater risk for myocardial infarction (MI), especially among subjects normally considered to be at low risk. However, little is known about the mechanism by which the ACE polymorphism affects the risk of MI. Coronary artery spasm (CAS) is considered to be one possible mechanism for developing MI. We therefore examined the ACE polymorphism relation to CAS to determine if this was the mechanism by which the DD genotype influences MI. We studied 150 angiographically assessed Japanese males, all more than 60 yr old. CASs were detected using intracoronary injection of ergonovine maleate. Subjects were divided into three groups: those with CAS (group 1), those without CAS, but with fixed organic stenosis (group 2); and those without CAS and no organic stenosis (group 3). DD subjects were significantly represented in group 1 when compared with groups 2 (P = 0.002) and 3 (P = 0.026). These results suggest that the DD genotype relates to the greater risk for MI in the patients with CAS.

Protective effects of soy protein on the peroxidizability of lipoproteins in cerebrovascular diseases.

To study the mechanism of dyslipoproteinemia, lipoproteins [very low density lipoprotein (VLDL), low density lipoprotein (LDL) and high density lipoprotein (HDL)] were isolated from stroke patients and healthy persons by ultracentrifugation. Lipoproteins were dialyzed into copper dichloride solution to study the effects of soycreme administration on lipoprotein peroxidation. Blood was drawn from 15 patients with cerebral thrombosis who were not administered soycreme, 10 patients with cerebral thrombosis who were administered soycreme and 11 healthy persons. The lipoproteins were dialyzed into 5 mol/l copper dichloride solution for various lengths of time, and then lipid constituents in the lipoproteins were measured by thin-layer chromatography. After the dialysis, percentages of cholesteryl ester and triglyceride in various lipoproteins decreased significantly (P < 0.05 or 0.01) in both patient groups and in healthy persons. Spot X1 was found between triglyceride and free fatty acid on the thin-layer chromatography, and spot X2 was located between free fatty acid and free cholesterol after dialysis. Spots X1 and X2 reflect lipoprotein peroxidation. Percentages of these spots were higher in VLDL, LDL and HDL in the patient groups than in the healthy subjects. Soycreme administration suppressed the appearance of spots X1 and X2. Furthermore, blood cholesterol concentrations were reduced by the administration of soy protein. Thus, soy may be useful in the prevention and/or treatment of atherosclerosis.

Transformation of cultured human monocytes by peroxidized low-density lipoprotein.

To study the roles of peroxidized low-density lipoprotein (pox-LDL) during the formation of foam cells from human monocytes, monocytes isolated from normal human blood were incubated with RPMI alone, normal low-density lipoprotein (n-LDL) and copper pox-LDL. The concentrations of total cholesterol (TCH) and lipid peroxide in the human monocyte medium did not change significantly after incubation with RPMI medium alone or n-LDL, but those of TCH decreased slightly after incubation with pox-LDL for 24 h. Triglyceride (TG) concentrations in the culture medium of human monocytes decreased after incubation with all above substances. Ultrastructural studies showed that the monocytes changed to macrophages after incubation with RPMI alone or n-LDL and to foam cells after incubation with pox-LDL for 48 h. We conclude that TGs may be metabolized for ATP production by human monocytes. The energy may play a role in the transformation of monocytes to macrophages, and pox-LDL can induce transformation of monocyte-derived macrophages to foam cells.

The effect of radix Salviae Miltiorrhizae Composita on peroxidation of low density lipoprotein due to copper dichloride.

It is well known that plasma lipoprotein, particularly oxidized LDL, plays an important role in the pathogenesis of atherosclerosis and atherosclerotic diseases. We used oxidized LDL generated by incubating LDL from healthy persons with copper dichloride as a model to investigate the antioxidate property of Radix Salviae Miltiorrhizae Composita (RSMC). On photos, the spot X1 and the spot X2 were clearly found in the control group after the dialysis into copper dichloride for 24 and 48 hours, but they could not found in the RSMC group. The analysis of the constituents of lipids in LDL (by charring method) showed that after dialysis the percentages of the spot X1 and the spot X2 in the RSMC group were significantly lower than those in the control group (P < 0.01). The results suggest that RSMC plays a potential role in antioxidation of lipids or LDL.