COMP-Ang1 Potentiates EPC Treatment of Ischemic Brain Injury by Enhancing Angiogenesis Through Activating AKT-mTOR Pathway and Promoting Vascular Migration Through Activating Tie2-FAK Pathway

Successful recovery from brain ischemia is limited due to poor vascularization surrounding the ischemic zone. Cell therapy with strong angiogenic factors could be an effective strategy to rescue the ischemic brain. We investigated whether cartilage oligomeric matrix protein (COMP)-Ang1, a soluble, stable and potent Ang1 variant, enhances the angiogenesis of human cord blood derived endothelial progenitor cells (hCB-EPCs) for rescuing brain from ischemic injury. COMP-Ang1 markedly improved the tube formation of capillaries by EPCs and incorporation of EPCs into tube formation with human umbilical vein endothelial cells (HUVECs) upon incubation on matrigel in vitro. COMP-Ang1 stimulated the migration of EPCs more than HUVECs in a scratch wound migration assay. The transplanted EPCs and COMP-Ang1 were incorporated into the blood vessels and decreased the infarct volume in the rat ischemic brain. Molecular studies revealed that COMP-Ang1 induced an interaction between Tie2 and FAK, but AKT was separated from the Tie2-FAK-AKT complex in the EPC plasma membrane. Tie2-FAK increased pp38, pSAPK/JNK, and pERK-mediated MAPK activation and interacted with integrins alphanubeta3, alpha4, beta1, finally leading to migration of EPCs. AKT recruited mTOR, SDF-1, and HIF-1alpha to induce angiogenesis. Taken together, it is concluded that COMP-Ang1 potentiates the angiogenesis of EPCs and enhances the vascular morphogenesis indicating that combination of EPCs with COMP-Ang1 may be a potentially effective regimen for ischemic brain injury salvage therapy.

Urinary MicroRNAs of Prostate Cancer: Virus-Encoded hsv1-miRH18 and hsv2-miR-H9-5p Could Be Valuable Diagnostic Markers / 대한배뇨장애요실금학회지

PURPOSE: MicroRNAs (miRNAs) in biological fluids are potential biomarkers for the diagnosis and assessment of urological diseases such as benign prostatic hyperplasia (BPH) and prostate cancer (PCa). The aim of the study was to identify and validate urinary cell-free miRNAs that can segregate patients with PCa from those with BPH. METHODS: In total, 1,052 urine, 150 serum, and 150 prostate tissue samples from patients with PCa or BPH were used in the study. A urine-based miRNA microarray analysis suggested the presence of differentially expressed urinary miRNAs in patients with PCa, and these were further validated in three independent PCa cohorts, using a quantitative reverse transcriptionpolymerase chain reaction analysis. RESULTS: The expression levels of hsa-miR-615-3p, hsv1-miR-H18, hsv2-miR-H9-5p, and hsa-miR-4316 were significantly higher in urine samples of patients with PCa than in those of BPH controls. In particular, herpes simplex virus (hsv)-derived hsv1-miR-H18 and hsv2-miR-H9-5p showed better diagnostic performance than did the serum prostate-specific antigen (PSA) test for patients in the PSA gray zone. Furthermore, a combination of urinary hsv2-miR-H9-5p with serum PSA showed high sensitivity and specificity, providing a potential clinical benefit by reducing unnecessary biopsies. CONCLUSIONS: Our findings showed that hsv-encoded hsv1-miR-H18 and hsv2-miR-H9-5p are significantly associated with PCa and can facilitate early diagnosis of PCa for patients within the serum PSA gray zone.

COMP-Angiopoietin-1 Stimulates Synovial Proliferation but Suppresses Osteoclast by Enhancing Angiogenesis and Osteoblast Maturation in Collagen-Induced Arthritis

OBJECTIVE: Angiopoietin-1 (Ang1) is a potent angiogenic factor that can increase synovial angiogenesis and also enhance osteoblast maturation and bone formation. However, its role in rheumatoid arthritis (RA) has not been well documented. Thus, we investigated roles of Ang1 in collagen-induced arthritis (CIA). METHODS: A recombinant adenovirus carrying the gene that encodes either cartilage oligomeric matrix protein (AdCOMP)-Ang1 (a modified form of Ang1) or LacZ (AdLacZ) was injected intravenously into CIA mice. Clinical, radiological, histopathological, and immunofluorescent analyses were performed. Serum levels of receptor activators of nuclear factor kappaB ligand (RANKL) and osteoprotegerin (OPG) and expression of osteoblast maturation genes were analyzed. RESULTS: AdCOMP-Ang1-injected mice developed more severe inflammation than the AdLacZ-injected mice. However, there were no significant differences in cartilage damage and bone erosion. More PECAM-1-positive blood vessels were seen in the synovium of the AdCOMP-Ang1-injected mice than in those injected with AdLacZ. Interestingly, a lower number of TRAP-positive osteoclasts were observed in AdCOMP-Ang1-injected CIA mice than in the AdLacZ group when comparing sections obtained from joints showing similar synovial proliferation. The serum OPG/RANKL ratio and expression of osteoblast maturation genes, such as runt-related transcription factor 2, bone sialoprotein, type 1 collagen, osteopontin, and osterix, were significantly upregulated in the AdCOMP-Ang1 group. CONCLUSION: COMP-Ang1 facilitates arthritis onset and increases synovial inflammation, but enhances osteoblast maturation, which in turn inhibits osteoclastogenesis by increasing the OPG/RANKL ratio in CIA. Our results suggest that careful investigation is necessary to delineate the possible therapeutic use of COMP-Ang1 as an adjunctive agent, in combination with anti-inflammatory therapies, for the prevention of bone destruction in RA.

Angiopoietin-1 Gene Therapy Attenuates Hypertension and Target Organ Damage in Nitric Oxide Synthase Inhibited Spontaneously Hypertensive Rats

BACKGROUND AND OBJECTIVES: In our previous study, we found that the gene transfer of a potent derivative of cartilage oligomeric matrix protein Angiopoietin-1 (COMP-Ang-1) substantially prevented hypertension, microvascular rarefaction, and target organ damage in spontaneously hypertensive rats (SHRs). The purpose of the present study was to examine the role of nitric oxide (NO) in the therapeutic effects observed after COMP-Ang-1 gene transfer. MATERIALS AND METHODS: To exclude the NO-mediated effects in COMP-Ang-1 gene therapy, the SHRs were treated with an NO synthase (NOS) inhibitor, Nw-nitro-L-arginine methyl ester (L-NAME) before the electrophoretic gene transfer. RESULTS: The pretreatment with L-NAME induced a severe and sustained increase in systolic blood pressure (BP) in a LacZ plasmid transferred control SHR. However, the electrophoretic transfer of a COMP-Ang-1 plasmid instead of LacZ plasmid in L-NAME-pretreated SHRs substantially blocked the development of hypertension without any significant difference in comparison with L-NAME-untreated COMP-Ang-1 plasmid transferred groups. In addition, the COMP-Ang-1 plasmid transfer substantially attenuated microvascular rarefaction and arteriole remodeling in the heart and kidney, which might account for the mild histological alterations observed in the COMP-Ang-1 plasmid transferred group, in contrast to the severe fibrosis and necrosis seen in the LacZ plasmid controls. CONCLUSION: These therapeutic outcomes of COMP-Ang-1 gene transfer even in NOS inhibited SHRs suggested that the antihypertensive effect of COMP-Ang-1 was not merely secondary to NO-mediated vasorelaxation, but it may be associated with its ability to protect the vascular endothelium probably via an NO-independent mechanism which serves to attenuate microvascular rarefaction and target organ damage, and also to prevent hypertension by reducing peripheral vascular resistance.

Activation of PPARgamma induces profound multilocularization of adipocytes in adult mouse white adipose tissues

We sought to determine the effects of activation of peroxisome proliferator-activated receptor-gamma (PPAR-gamma) on multilocularization of adipocytes in adult white adipose tissue (WAT). Male C57BL/6 normal, db/db, and ob/ob mice were treated with agonists of PPAR-gamma, PPAR-alpha, or beta3-adrenoceptor for 3 weeks. To distinguish multilocular adipocytes from unilocular adipocytes, whole-mounted adipose tissues were co-immunostained for perilipin and collagen IV. PPAR-gamma activation with rosiglitazone or pioglitazone induced a profound change of unilocular adipocytes into smaller, multilocular adipocytes in adult WAT in a time-dependent, dose-dependent, and reversible manner. PPAR-alpha activation with fenofibrate did not affect the number of locules or remodeling. db/db and ob/ob obese mice exhibited less multilocularization in response to PPAR-gamma activation compared to normal mice. Nevertheless, all adipocytes activated by PPAR-gamma contained a single nucleus regardless of locule number. Multilocular adipocytes induced by PPAR-gamma activation contained substantially increased mitochondrial content and enhanced expression of uncoupling protein-1, PPAR-gamma coactivator-1-alpha , and perilipin. Taken together, PPAR-gamma activation induces profound multilocularization and enhanced mitochondrial biogenesis in the adipocytes of adult WAT. These changes may affect the overall function of WAT.

Angiopoietin-1 Protects Endothelial Cells From Hypoxia-Induced Apoptosis via Inhibition of Phosphatase and Tensin Homologue Deleted From Chromosome Ten

BACKGROUND AND OBJECTIVES: Angiopoietin-1 (Ang1) is a regulator of blood vessel growth and maturation, and prevents radiation-induced or serum deprivation-induced apoptosis. Phosphatase and tensin homologue deleted from chromosome ten (PTEN), a well-known tumor suppressor, regulates cell cycle arrest and apoptosis. Hypoxia induces apoptosis by increasing the expression of PTEN. We hypothesized that Ang1 may regulate PTEN expression and, thus, reduce endothelial apoptosis under hypoxia in vitro and in vivo. Materials and METHODS: In vitro, human umbilical vein endothelial cells (HUVECs) were treated with Ang1, and signaling pathways were investigated. In vivo, eight-week-old C57BL/6 mice were used for a hind limb ischemia model. Ang1 or normal saline was intramusculary injected. Blood flow was evaluated by a laser Doppler perfusion analyzer and tissue histology. RESULTS: The expression of PTEN was markedly upregulated in HUVECs after hypoxic stimulation, whereas Ang1 suppressed PTEN expression. Tie2-Fc, a soluble form of Tie2 (sTie2) that blocks Ang1, reversed the Ang1 effect on PTEN reduction under hypoxia. Ang1 inhibited the nuclear translocation of nuclear transcription factor-kB (NF-kB), a binding factor for the PTEN promoter and Foxo1. Hypoxia-induced p27 expression and apoptosis were also suppressed by Ang1. In the mouse hind limb ischemia model, we observed a high capillary density, numerous proliferating cells and diminished cell death in skeletal muscle tissue in the Ang1 injected group. CONCLUSION: Ang1 enhanced endothelial cell survival by reducing apoptosis via PTEN down-regulation in HUVECs under hypoxia. Local injection of Ang1 significantly reduced apoptotic cells in vivo, and prevented limb loss for ischemic hind limb mice. Thus, Ang1 may be an effective therapeutic for protection from ischemic-endothelial cell injury.

Angiopoietin-1 variant, COMP-Ang1 attenuates hydrogen peroxide-induced acute lung injury

Reactive oxygen species (ROS) play a crucial role in acute lung injury. Tissue inflammation, the increased vascular permeability, and plasma exudation are cardinal features of acute lung injury. Angiopoietin-1 (Ang1) has potential therapeutic applications in preventing vascular leakage and also has beneficial effects in several inflammatory disorders. Recently developed COMP-Ang1 is more potent than native Ang1 in phosphorylating tyrosine kinase with immunoglobulin and EGF homology domain 2 receptor in endothelial cells. However, there are no data on effects and related molecular mechanisms of COMP- Ang1 on ROS-induced acute lung injury. We used hydrogen peroxide (H2O2)-inhaled mice to evaluate the effect of COMP-Ang1 on pulmonary inflammation, bronchial hyper-responsiveness, and vascular leakage in acute lung injury. The results have revealed that VEGF expression, the levels of IL-4, TNF-alpha, IL-1 beta, intercellular adhesion molecule-1, and vascular cell adhesion molecule-1 in lungs, the levels of hypoxia-inducible factor-1alpha (HIF-1 alpha) and NF-kappa B in nuclear protein extracts, phosphorylation of Akt, and vascular permeability were increased after inhalation of H2O2 and that the administration of COMP-Ang1 markedly reduced airway hyper-responsiveness, pulmonary inflammation, plasma extravasation, and the increases of cytokines, adhesion molecules, and VEGF in lungs treated with H2O2. We have also found that the activation of HIF-1a and NF-kappa B and the increase of phosphoinositide 3-kinase activity in lung tissues after H2O2 inhalation were decreased by the administration of COMP-Ang1. These results suggest that COMP-Ang1 ameliorates ROS-induced acute lung injury through attenuating vascular leakage and modulating inflammatory mediators.

COMP-angiopoietin 1 Gene Transfer Enhances Cutaneous Wound Healing by Promoting Angiogenesis

BACKGROUND: Angiogenesis is crucial for wound healing and exogenous supplements of the angiogenic growth factors have been known to promote cutaneous wound healing. Angiopoietin (Ang) 1 is a recently discovered angiogenic factor and there have been few studies of its effect on cutaneous wound healing. OBJECTIVE: We examined the effect of Ang 1 on cutaneous wound healing. METHODS: Cartilage oligomeric matrix protein (COMP)-Ang 1 (Ade-COMP-Ang 1)- was intravenously injected to rats two days before surgery creating full-thickness wounds. The clinical wound healing rate and the number of vessels in the skin samples were evaluated on days 3, 7 and 14 post operation. RESULTS: At post-operation day 3, 7 and 14, the clinical wound healing rate was 38.3%, 59.4% and 92.1%, respectively, in the Ade-COMP-Ang 1-treated group, compared with 20.5%, 47.5% and 87.3%, respectively, in the Ade-LacZ-treated group. There were significant differences in the results of day 3 and day 7 between two groups (p<0.05). Histopathologically, the number of the vessels of the Ade-COMP-Ang 1-treated group was 73.7, 94.1 and 62.7 at day 3, 7 and 14, compared with that of the Ade-LacZ-treated group, 53.5, 83.9, and 56.9. The differences in the results of the two groups were statistically significant (p<0.05). CONCLUSION: These results indicate that Ade-COMP-Ang 1 therapy significantly accelerats wound healing by promoting angiogenesis. However, further study using Ade-COMP-Ang 1 gene therapy for chronic wounds in which the formation of new blood vessels is impaired is needed in the near future.

Blockade of airway inflammation and hyper-responsiveness by an angiopoietin-1 variant, COMP-Ang1

Inflammation of the asthmatic airway is usually accompanied by increased vascular permeability and plasma exudation. Angiopoietin-1 (Ang1) has potential therapeutic applications in preventing vascular leakage. Recently, we developed a soluble, stable, and potent Ang1 variant, COMP-Ang1. COMP-Ang1 is more potent than native Ang1 in phosphorylating the tyrosine kinase with immunoglobulin and epidermal growth factor homology domain 2 receptor in lung endothelial cells. We have used a mouse model for allergic airway disease to determine effects of COMP-Ang1 on allergen-induced bronchial inflammation and airway hyper-responsiveness. These mice develop the following typical pathophysiological features of allergic airway disease in the lungs: increased numbers of inflammatory cells of the airways, airway hyper-responsiveness, increased levels of Th2 cell cytokines (IL-4, IL-5, and IL-13), adhesion molecules (intercellular adhesion molecule-1 and vascular cell adhesion molecule-1), and chemokines (eotaxin and RANTES), and increased vascular permeability. Intravenous administration of COMP-Ang1 reduced bronchial inflammation and airway hyper-responsiveness. In addition, the increased plasma extravasation in allergic airway disease was significantly reduced by the administration of COMP-Ang1. These results suggest that COMP-Ang1 attenuates airway inflammation and hyper-responsiveness, prevents vascular leakage, and may be used as a therapeutic agent in allergic airway disease.

Relationship of Angiopoietin-2 Expression to Atherosclerosis in Middle Cerebral and Coronary Arteries / 대한해부학회지

Atherosclerosis is a systemic and multifactorial disease, its incidence is raised recently. Cerebral and coronary atherosclerosis have some similar pathogenesis, but their relationship and mechanisms are still remain unclear. Intimal neovascularization in the atherosclerotic plaque was focused with respect to its pathological roles, intimal thickening and atherosclerotic progression. Ang-2, which is an angiogenesis regulating factor, provides a destabilizing signal for endothelial cells, leading to vessel regression or sprouting. However the role and distribution of Ang-2 in atherosclerotic coronary and cerebral arteries are still not well known. Thus, we analyzed 1) atherosclerotic lesion progression 2) relationship of atherosclerosis to Ang-2 expression in human middle cerebral and coronary artery. Paraffin sections from 25 human coronary (COA) and 36 middle cerebral arteries (MCA) were characterized according to AHA classification. In the same person, the score of atherosclerosis progression in COA was higher than that of MCA. In the two kinds of arteries having same atherosclerotic progression, the degree of intimal proliferation and luminal stenosis in COA was higher than that of MCA. Expression of Ang-2 was not shown in normal artery but localized in lumen-lining endothelium, macrophage in preatheroma, atheroma and complicated lesion. Ang-2 expression and infiltration of macrophages were rich in COA than MCA. Our result indicated that cerebral atherosclerosis has some different pathogenic mechanisms with coronary atherosclerosis according to difference of progression and angiogenic factor Ang-2 expression. Thus this is a fundamental study for understanding the progression of atherosclerosis in different vascular beds.

Pathophysiological Role of Angiopoietin-2 in Human Coronary Atherosclerosis / 대한해부학회지

Neovascularization is well known to occur in human atherosclerotic plaques; however, its pathophysiological roles, mechanism, and stimuli still remain unclear. Angiopoietin-1 and -2 belong to another vascular-specific growth factor family and regulate angiogenesis. Angiopoietin-2 (Ang-2) provides a destabilizing signal for endothelial cells, leading to vessel regression or sprouting depending on the presence of other angiogenic factor. But role and distribution of Ang-2 in atherosclerosis are not well known. Thus, we studied 1) the distribution and amount of Ang-2 2) the relationship between Ang-2 expression and vascular morphometrical change 3) the relationship between Ang-2 expression and neovascularization in atherosclerotic lesions. Paraffin sections from 36 human coronary arterial segments were characterized as normal, preatheroma, atheroma, fibroatheroma and complicated lesion according to American heart association classification. Expression of Ang-2 and related factors were examined using immunohistochemistry and western blotting with antibodies against Ang-2, CD31 (endothelial cells), alpha-actin (vascular smooth muscle cells), CD36 (monocyte & macrophage), Tie-2 and VEGF. Expression of Ang-2 was not shown in normal arterial segment. Ang-2 were localized in lumen-lining endothelium, macrophage, some SMCs of atheromatous plaque in advanced lesion. Amount of Ang-2 was increased according to progression of atherosclerosis. Intraplaque microvessels had Ang-2 and VEGF positive endothelial cells and number of those in plaque increased according to progression of disease. Intimal neovascularization is correlated with intimal thickening in atherosclerotic lesion (R2 = 0.7424). Therefore, they suggest that Ang-2 has an important role in the progression of human coronary atherosclerosis, as well as in neovascularization. This study implicates Ang-2 as an important potential therapeutic target in vascular disease

Biomedical significance of endothelial cell specific growth factor, angiopoietin

Until recently, vascular endothelial growth factor (VEGF) was the only growth factor proven to be specific and critical for blood vessel formation. Other long-known factors, such as the fibroblast growth factors (FGFs), platelet-derived growth factor, or transforming growth factor-beta, had profound effects in endothelial cells. But such factors were nonspecific, in that they could act on many other cells, and it seemed unlikely that these growth factors would be effective targets for treatment of endothelial cell diseases. A recently discovered endothelial cell specific growth factor, angiopoietin, has greatly contributed to our understanding of the development, physiology, and pathology of endothelial cells (Davis et al., 1996; Yancopoulos et al., 2000). The recent studies that identified and characterized the physiological and pathological roles of angiopoietin have allowed us to widen and deepen our knowledge about blood vessel formation and vascular endothelial function. Therefore, in this review, we describe the biomedical significance of these endothelial cell growth factors, the angiopoietins, in the vascular system under normal and pathological states.

The Antiapoptotic Effect of Vascular Endothelial Growth Factor and Angiopoietin-1 on Glomerular Endothelial Cells in Adriamycin or Cisplatin-induced Apoptosis / 대한신장학회잡지

BACKGROUND: Because glomerular endothelium play a pivotal role in the renal diseases, damage of glomerular endothelial cells lead to progression of glomerular sclerosis and decrement of renal function. Apoptotic damage of cells is an important mechanism in renal disease. Therefore, several growth factors that have antiapoptotic effect may have a protective role in maintaining a renal function in apoptotic cell injury. METHODS: The present study evaluated whether cisplatin or adriamycin induce apoptosis in glomerular endothelial cells. We also evaluated the antiapoptotic effect of angiopoietin-1 and VEGF in cisplatin or adriamycin- induced apoptosis. RESULTS: Cisplatin or adriamycin induced apoptosis in glomerualr endothelial cell in dose dependent manner. Angiopoietin-1 and VEGF produced antiapoptotic effect in cisplatin or adriamycin-induced apoptosis in a dose dependent manner. The antiapoptotic effect of angiopoietin-1 was more potent than that of VEGF in glomerualr endothelial cells. Wortmannin, a phosphatidylinositol 3'-kinase inhibitor decrease the angiopoietin-1 or VEGF-induced antiapoptotic effect. CONCLUSION: These results suggest that angiopoietin-1 and VEGF may be a strong survival factor for the glomerular endothelial cells in the cisplatin or adriamycin-induced apoptosis through phosphatidylinositol 3'-kinase/Akt. Therefore, pretreatment of angiopoietin-1 and VEGF could play a beneficial role for maintaining normal glomerular endothelial cell integrity before and during systemic cisplatin or adriamycin therapy.

Angiogenic Factors and Their Receptors / 대한내분비학회지

No abstract available.

Gene Ploymorphisms of Angiotensin Converting Enzyme and Endothelial Nitric Oxide Synthase in Patients with IgA Nephropathy / 대한신장학회잡지

To evaluate the association between ACE gene I/D polymorphism and ecNOS gene a/b polymorphism in IgA nephropathy, 158 IgA nephropathy patients and 121 control subjects were examined. In genotype distribution of the ACE gene I/D polymorphism, there was no significant difference in genotype distribution between controls and IgA nephropathy patients. We also examined the association between ACE genotype and clinical characteristics in the patients with IgA nephropathy. The incidence of hypertension in patients with DD genotype was higher than that of other genotypes. There were no significant association between I/D polymorphism distribution and chronic renal failure, nephrotic range proteinuria, and glomerular sclerosis in IgA nephropathy. In genotype distribution of ecNOS gene a/b polymorphism, there was no significant difference between IgA nephropathy patients and controls. There was no significant difference in frequency of chronic renal failure, hypertension, nephrotic range proteinuria and glomerular sclerosis among ecNOS genotypes. In addition, we failed to detect any significant association between the ACE and ecNOS gene-polymorphis ms and the decline of renal function in IgA nephropathy. A further study with larger number of patient population would be necessary.

Alteration of Insulin-like Growth Factor ( IGF ) -I and IGP-Binding Proteins in Renal Development and Regeneration

The effect of Roentgen rays on carcinoma of the cervix has long been of great interest to both radiologists and gynecologists. Since most cervical carcinomas are treated by irradiation, any additional knowledge either concerning the radiosensitivity of cervical tumors or their ultimate prognosis would be of value. The vaginal smear is considered to be one of convenient and rapid methods to study the effects of radiation on cervical malignancy. We observed morphologic changes in 297 cytologic preparations obtained from 60 patients who had underwent irradiation for cancer of the cervix. With the morphologic parameters such as cytoplasmic vacuolization, cytoplasmic basophilia, multinucleated giant cell formation, polymorphonuclear leucocytes (PMNL) sticking and postradiation dysplasia, we analyzed the findings in relation to the follow up time interval. The most common effect was the cytoplasmic vacuolization with basophilia of basaloid cells, which were noted in more than 90% of followed patients. The multinucleated giant cell formation and PMNL stickering were noted in 38 cases(63%) and 48 cases(80% ) respectively. The differential diagnosis of postradiation dysplasia from recurrent or persistent carcinoma, reparative atypical cells, and regressing tumor cells was difficult and further study seems to be needed to clarify the more accurate morphologic features and biologic behavior.

The Synthesis and Distribution of TGF-beta1 in Cardiac Valves

BACKGROUND AND OBJECTIVES: Transforming growth factor-beta1 (TGF-beta1) plays an important role on cardiac muscle differentiation, cardiac septa and valve formation during heart development. However, the role of TGF-beta1 in cardiac valves of adult animals is largely unknown. Cardiac valves are target portion from repetitive, periodic and continuous physical loading in the body. Therefore, we examined the mRNA, protein levels, and protein distribution of TGF-beta1 in cardiac valves of adult animals to clarify the biological importance of TGF-beta1. MATERIALS AND METHODS: Adult mice, rats and pigs were used. Cardiac valves of pig were frozen and were pulverized with liquid nitrogen. To measure the mRNA levels of TGF-beta1 in cardiac valves, total RNA was extracted using Tri-reagent and performed Northern blot analysis. To measure the protein levels of TGF-beta1 in cardiac valves, total protein was extracted and performed Western blot analysis. To examine the TGF-beta1 distribution, immuno-histochemistry with anti-CC-1-30 antibody was performed. RESULTS: The mRNA level of TGF-beta1 in pulmonary valve was higher than those in the other valves. However, the protein levels of TGF-beta1 were similar among valves. The mRNA and protein levels of TGF-beta1 in cardiac valves were higher than those in atria or ventricles. The TGF-beta1 protein was located mainly in cellular interstitium in cardiac valves. The distribution of TGF-beta1 protein in surface area was higher than in the mid-portion of valves. CONCLUSION: These results suggest that synthesis and distribution of TGF-beta1 in cardiac interstitum is essential for maintaining of normal structure and function on various physical loading.

Angiotensin Converting Enzyme Gene Polymorphism in Patients with Minimal Change Nephrotic Syndrome and Focal Segmental Glomerulosclerosis / 대한신장학회잡지

Angiotensin converting enzyme gene insertion/ deletion polymorphism has been shown to be associated with cardiovascular disease including cardiomyopathy, myocardial infarction, essential hypertension, progression of IgA nephropathy and diabetic nephropathy. Since glomerulosclerosis has similarities to atherosclerosis, angiotensin converting enzyme gene polymorphism may be associated with glomerulsclerosis. Therefore, we tested whether genotype distribution of the insertion/deletion polymorphism in angiotensin converting enzyme gene is different in patients with minimal change nephrotic syndrome and focal segmental glomerulosclerosis. In genotype distribution of the angiotensin converting enzyme gene I/D polymorphism, control subjects were II type 44.3%, ID type 40.9%, DD type 14.8% and patients with minimal change nephrotic syndrome were II type 38.2%, ID type 45.5%, DD type 16.3% and patients with focal segmental glomerulosclerosis were II type 13.3%, ID type 46.7%, DD type 40.0%. This result suggests that DD genotype was more frequent in patients with focal segmental glomerulosclerosis than minimal change nephrotic syndrome and control subjects. We also examined the association between ACE genotype and clinical characteristics in the patients with minimal change nephrotic syndrome and focal segmental glomerulosclerosis. There were no significant association between I/D polymorphism distribution and hypertension, chronic renal failure, response to steroid in patients with minimal change nephrotic syndrome. The incidence of chronic renal failure in patients with focal segmental glomerulosclerosis DD genotype was higher than that of other genotypes. The response rate to steroid in patients with focal segmental glomerulosclerosis DD genotype was lower than that of other genotypes.

Cell cycle regulators during human atrial development

OBJECTIVES: The molecular mechanisms that regulate cardiomyocyte cell cycle and terminal differentiation in humans remain largely unknown. To determine which cyclins, cyclin dependent kinases (CDKs) and cyclin kinase inhibitors (CKIs) are important for cardiomyocyte proliferation, we have examined protein levels of cyclins, CDKs and CKIs during normal atrial development in humans. METHODS: Atrial tissues were obtained in the fetus from inevitable abortion and in the adult during surgery. Cyclin and CDK proteins were determined by Western blot analysis. CDK activities were determined by phosphorylation amount using specific substrate. RESULTS: Most cyclins and CDKs were high during the fetal period and their levels decreased at different rates during the adult period. While the protein levels of cyclin D1, cyclin D3, CDK4, CDK6 and CDK2 were still detectable in adult atria, the protein levels of cyclin E, cyclin A, cyclin B, cdc2 and PCNA were not detectable. Interestingly, p27KIP1 protein increased markedly in the adult period, while p21CIP1 protein in atria was detectable only in the fetal period. While the activities of CDK6, CDK2 and cdc2 decreased markedly, the activity of CDK4 did not change from the fetal period to the adult period. CONCLUSION: These findings indicate that marked reduction of protein levels and activities of cyclins and CDKs, and marked induction of p27KIP1 in atria, are associated with the withdrawal of cardiac cell cycle in adult humans.

Plasma Angiotensin / 대한내분비학회지

The purpose of the present study was to define the normal range of plasma concentration of angiotensin I-converting enzyme(ACE), N-acetyl-b-D-glucosaminidase(NAG), inactive and active renin, and atrial natriuretic peptide(ANP) in normal Korean adult male in terms of aging. Both plasma ACE activity and NAG concentration were measured by spectrofluorometry, and the plasma renin activity and ANP concentration were measured using radioimmunoassay. The ACE was 67.7+-3.6 nM His-Leu/min/ml and did not change in terms of age. The plasma NAG activity tended to decrease. Both plasma active and inactive renin activities were 2.1+-0.2 and 3.0+-0.3 ngAI/ml/h and tended to decrease in terms of aging. The percentage of inactive renin to total renin was 57.2+-2.9% at age 21-30 and also tended to decrease in terms of aging. Plasma ANP concentration at age 22 was 59.6+-2.9 pg/ml.