TAK-875, a GPR40/FFAR1 agonist, in combination with metformin prevents progression of diabetes and ß-cell dysfunction in Zucker diabetic fatty rats.

BACKGROUND AND PURPOSE: TAK-875, a selective GPCR40/free fatty acid receptor 1 agonist, improves glycaemic control by increasing glucose-dependent insulin secretion. Metformin is a first-line drug for treatment of type 2 diabetes that improves peripheral insulin resistance. Based on complementary mechanism of action, combining these agents is expected to enhance glycaemic control. Here, we evaluated the chronic effects of TAK-875 monotherapy and combination therapy with metformin in diabetic rats. EXPERIMENTAL APPROACH: Long-term effects on glycaemic control and ß-cell function were evaluated using Zucker diabetic fatty (ZDF) rats, which develop diabetes with hyperlipidaemia and progressive ß-cell dysfunction. KEY RESULTS: Single doses of TAK-875 (3-10 mg·kg(-1) ) and metformin (50-150 mg·kg(-1) ) significantly improved both postprandial and fasting hyperglycaemia, and additive improvements were observed in their combination. Six-week treatment with TAK-875 (10 mg·kg(-1) , b.i.d.) significantly decreased glycosylated Hb (GHb) by 1.7%, and the effect was additively enhanced by combination with metformin (50 mg·kg(-1) , q.d.; GHb: -2.4%). This improvement in glycaemic control in the combination group was accompanied by significant 3.2-fold increase in fasting plasma insulin levels. Pancreatic insulin content was maintained at a level comparable to that in normal rats by combination treatment (vehicle: 26, combination: 67.1; normal lean: 69.1 ng·mg(-1) pancreas) without affecting pancreatic glucagon content. Immunohistochemical analyses revealed normal morphology, enhanced pancreas duodenum homeobox-1 expression and increased PCNA-positive cells in islets of the combination group. CONCLUSION AND IMPLICATIONS: Our results indicate that combination therapy with TAK-875 and metformin could be a valuable strategy for glycaemic control and ß-cell preservation in type 2 diabetes.

Long-term observation of avascular necrosis of the femoral head in systemic lupus erythematosus: an MRI study.

OBJECTIVES: To assess long-term prognosis of clinically silent, early-stage avascular necrosis of the femoral head (ANFH) in patients with systemic lupus erythematosus (SLE). METHODS: Twenty-four hips that showed ANFH by magnetic resonance imaging (MRI) in 13 patients with SLE were studied. All hips were radiographically normal and clinically asymptomatic. The percentage volume of necrotic bone was calculated at each study by dividing the sum of the necrotic areas by the sum of the femoral head areas from all MRI slices. Hips were also classified into three categories by the relation of the necrotic area to the weight bearing portion according to the system of the Japanese Investigation Committee for avascular necrosis of the femoral head, with modifications: Type A (medial lesions): 8 hips, Type B (central lesions): 4 hips, and Type C (lateral lesions): 12 hips. Patients were followed up with MRI for 12-95 (mean 51) months. RESULTS: Fifteen hips improved (more than 15% reduction in the volume of necrosis), 5 did not change and 4 worsened during the observation period. All hips with a volume of necrotic area less than 25% showed improvement. All but one Type A hip and one Type B hip improved, while the mean volume of necrosis did not change in Type C. The volume of the necrotic area was smaller in Type A & B than in Type C hips (p < 0.001). CONCLUSIONS: Long-term prognosis of early-stage ANFH was favorable in patients with SLE when the necrotic area was small (less than 25%).

Beraprost sodium regulates cell cycle in vascular smooth muscle cells through cAMP signaling by preventing down-regulation of p27(Kip1).

OBJECTIVE: Beraprost sodium (BPS), a prostacyclin (PGI(2)) analogue, has been reported to exhibit beneficial effects on atherosclerosis in both human and animal models. To clarify the underlying mechanism, we investigated the effects of BPS on neointimal formation after balloon injury in the canine coronary artery. Furthermore, we determined its anti-atherosclerotic effects in cultured smooth muscle cells (SMCs). METHODS: Adult beagle dogs (10-12 kg) were fed on a high-cholesterol diet (10 g/day) and underwent balloon-denudation of the coronary artery. The dogs were divided into two groups: a BPS-treated group (20 microg/kg per day) and a control group. Twenty-eight days after injury, the dogs were killed and the coronary arteries were examined morphometrically. Three days after injury, the proliferative activity in the medial layer of the coronary artery was evaluated by 5-bromo-2'-deoxyuridine (BrdU) incorporation, and p27(Kip1), a cyclin-dependent kinase (cdk) inhibitor, expression was examined by immunohistochemistry. We also examined the effects of BPS on SMC proliferation based on BrdU incorporation and cell cycle analysis. In addition, p27(Kip1) regulation was evaluated in primary-cultured SMCs. RESULTS: BPS administration decreased the intima/media ratio (I/M) by 88% in the control group. Three days after injury, BPS attenuated the proliferation rate of the cells in the media of the coronary artery by 35%, and maintained p27(Kip1) expression, which declined in the control cells. In the cultured proliferating SMC, BPS prevented the down-regulation of p27(Kip1). The 8-bromo-cyclic adenosine monophosphate (8-br-cAMP), a cAMP analogue, had similar actions as BPS in the regulation of p27(Kip1). The proliferation of cultured SMC was inhibited in a dose-dependent manner, and cell cycle arrest in the G1 phase was induced by BPS. CONCLUSIONS: Our data suggest that BPS inhibits neointimal formation after balloon denudation in the coronary artery through its inhibitory effect on SMC proliferation by preventing p27(Kip1) down-regulation.

Psoriasis in a patient with juvenile rheumatoid arthritis.

A 20-year-old woman had polyarticular-onset type of juvenile rheumatoid arthritis with a chronic and destructive course since 6 years of age. She had developmental retardation, deformity and disability. Asymptomatic erythematous scaly plaques developed on the trunk. A skin biopsy specimen revealed psoriasis. This is the first report of psoriasis developing in a patient with juvenile rheumatoid arthritis.

Increased migration in late G(1) phase in cultured smooth muscle cells.

Migration and proliferation of smooth muscle cells (SMC) contribute to neointimal formation after arterial injury. However, the relation between migration and proliferation in these cells is obscure. To discriminate between migration and proliferation, we employed a migration assay of SMC at different phases of the cell cycle. Serum-deprived SMC were synchronized in different phases of the cell cycle by addition of serum for various periods of time. Migration induced by platelet-derived growth factor B-chain homodimer was maximal in SMC that were predominantly in the late G(1) (G(1b)) phase. In addition, in nonsynchronized SMC, 65-75% of SMC that had migrated were in the G(1b) phase. Phosphorylated myosin light chain was enriched around the cell periphery in SMC in the G(1b) phase compared with SMC in the other cell cycle phases. Interestingly, the Triton X-100-insoluble fraction of myosin was remarkably decreased in G(1b)-enriched SMC. These findings suggest that migratory activity of SMC may be coupled with the G(1b) phase. The phosphorylation and retention of myosin might explain some of the properties responsible for increased migration.

Angiographically demonstrated coronary collaterals predict residual viable myocardium in patients with chronic myocardial infarction: a regional metabolic study.

Angiographical demonstration of coronary collateral circulation may suggest the presence of residual viable myocardium. The development of coronary collaterals was judged according to Rentrop's classification in 37 patients with old anteroseptal myocardial infarction and 13 control patients with chest pain syndrome. The subjects with myocardial infarction were divided into 2 groups: 17 patients with the main branch of the left coronary artery clearly identified by collateral blood flow from the contralateral coronary artery [Coll(+)group, male/female 10/7, mean age 56.6 years]and 20 patients with obscure coronary trunk [Coll(-)group, male/female 16/4, mean age 54.9 years]. Thallium-201 myocardial scintigraphy and examination of local myocardial metabolism were carried out by measuring the flux of lactic acid under dipyridamole infusion load. Coronary stenosis of 99% or total occlusion was found in only 5 of 20 patients (25%)in the Coll(-)group but in 16 of 17 patients(94%)in the Coll(+)group(p < 0.001). Redistribution of myocardial scintigraphy was found in 11 of 15 patients(73%)in the Coll(+)group, but only 3 of 18 patients (17%)in the Coll(-)group(p < 0.01). The myocardial lactic acid extraction rate was--13.2 +/- 17.0% in the Coll(+)group, but 9.1 +/- 13.2% in the Coll(-)group(p < 0.001). These results suggest that coronary collateral may contribute to minimizing the infarct area and to prediction of the presence of viable myocardium.

Scleroderma-like reaction induced by uracil-tegafur (UFT), a second-generation anticancer agent.

UFT, a combination of uracil and tegafur, is a second-generation anticancer agent. UFT has been used in Japan, other Asian countries, South America, and Russia. Recently, UFT has been extensively studied for colorectal, pancreatic, and various types of cancer in North America and Europe, especially with leukovorin. We report a case of a scleroderma-like reaction induced by long-term administration of UFT. This is the first report of UFT-induced scleroderma-like reaction.

The kinase inhibitor fasudil (HA-1077) reduces intimal hyperplasia through inhibiting migration and enhancing cell loss of vascular smooth muscle cells.

Smooth muscle cell (SMC) migration plays an important role in restenosis after angioplasty. Myosin phosphorylation is necessary for cell migration. Fasudil is an inhibitor of protein kinases, including myosin light chain kinase and Rho associated kinase, thereby inhibiting myosin phosphorylation, and it has been clinically used to prevent vasospasm following subarachnoid hemorrage. Based on these findings, we examined the anti-migrative action of fasudil. In SMC (SM-3), fasudil (1-100 microM) inhibited SMC migration in a dose-dependent manner (p < 0.001). Fasudil suppressed actin stress fiber formation dose dependently. In rabbit carotid artery, fasudil (10 mg/kg/day) markedly reduced intimal hyperplasia 14 days following balloon injury. Cell kinetic study showed that fasudil did not affect proliferation but enhanced cell loss in the media after injury. We concluded that fasudil reduced neointimal formation after balloon injury through both inhibiting migration and enhancing cell loss of medial SMC.

Inhibition of smooth muscle cell migration by the p21 cyclin-dependent kinase inhibitor (Cip1).

In vascular smooth muscle cells (SMCs), proliferation and migration contribute to lesion formation after arterial injury. In the cell cycle, several cyclin-dependent kinases (cdks) inhibitors are implicated in the regulating of cyclin-cdk activity such as p21Cip1, p16Ink4 and p27Kip1. Although Cip1 inhibits SMC proliferation, its effects on SMC migration are unknown. To test the hypothesis that Cip1 inhibits SMCs migration and proliferation, we transfected the Cip1 gene into a strain of rabbit aortic SMCs (SM3 cells). Both the spreading and the attachment of Cip1-transfected SM3 cells to extracellular matrices (ECMs) were inhibited compared to that of vector-transfected cells. In the modified Boyden's chamber assay the effect of fibronectin on the migratory activity of Cip1-transfected SM3 cells was significantly less than that of vector transfected cells in response to PDGF-BB. These data suggested that Cip1 inhibited both the migration and proliferation of SMC.

Anti-angiogenic compound (TNP-470) inhibits mesangial cell proliferation in vitro and in vivo.

Growth factors, especially basic fibroblast growth factor (bFGF), platelet-derived growth factor (PDGF), and transforming growth factor-beta (TGF-beta) are known to play key roles in the pathogenesis of mesangial proliferative glomerulonephritis. TNP-470 (AGM-1470), a potent anti-angiogenic compound, has anti-growth factor properties and inhibits the activation of cyclin-dependent kinase (cdk) 2 and phosphorylation of RB protein. We investigated whether TNP-470 could suppress growth factor induced mesangial cell proliferation in vitro and experimental model of mesangial proliferative glomerulonephritis in vivo. TNP-470 inhibited potently PDGF- and bFGF-stimulated proliferation of rat mesangial cells in vitro (IC50 = 50 pg/ml). In anti-Thy 1.1 glomerulonephritis, high dose use of TNP-470 (20 mg/kg/day) markedly suppressed mesangial cell proliferation and mesangial matrix expansion on day 6; however, mesangiolysis remained. Low dose use of TNP-470 (10 mg/kg/day) moderately inhibited mesangial cell proliferation and mesangial matrix synthesis, and induced appropriate glomerular healing on day 14 in anti-Thy 1.1 glomerulonephritis. Thus, TNP-470 potently inhibits growth factor-induced proliferation of mesangial cells in vitro, and mesangial cell proliferation and extracellular matrix expansion in anti-Thy 1.1 glomerulonephritis in vivo. These results suggest a novel therapeutic potential of TNP-470 in mesangial proliferative glomerulonephritis.

Enhanced gene expression of scavenger receptor in peripheral blood monocytes from patients on cuprophane haemodialysis.

BACKGROUND: Macrophage scavenger receptor (SR) is implicated in playing a key role in macrophage-derived foam cell formation by taking up a large amount of modified low-density lipoproteins (LDL). It has also been postulated that alpha 2-macroglobulin receptor/LDL receptor-related protein (alpha 2MG/LRP) is involved in the development of foam cells by taking up apo E-enriched chylomicrons and VLDL remnants, and lipoprotein lipase-triglyceride-rich lipoprotein complexes. Accumulation of these lipid-loaded monocyte/ macrophages in the subendothelial space is considered to be an early event of atherogenesis. Since atherogenesis is considered to be accelerated in dialysis patients, we attempted to investigate whether gene expression of SR and alpha 2MG/LRP are altered in peripheral blood monocytes from patients on haemodialysis with a cuprophane (Cu) or polymethylmethacrylate (PMMA) membranes. METHODS: Peripheral blood monocytes (PBM) were prepared from patients undergoing haemodialysis with a Cu membrane (n = 9), patients undergoing haemodialysis with a PMMA membrane (n = 9), and healthy controls (n = 7). In a separate experiment we examined SR gene expression in uraemic patients (n = 12) and healthy controls (n = 9). SR and alpha 2MG/LRP mRNA were semiquantitated using reverse-transcription polymerase chain reaction (RT-PCR) assay followed by Southern blotting. RESULTS: SR mRNA expression in PBM from patients on chronic haemodialysis with a Cu membrane was about twofold higher than that in PBM from patients on chronic haemodialysis with a PMMA membrane or the controls (P < 0.05). alpha 2MG/LRP mRNA expression in PBM showed no difference among these, three groups. SR gene expression in monocytes from uraemic patients was not increased compared with that in the controls. CONCLUSION: PBM from patients under Cu membrane dialysis showed higher gene expression of SR than patients under PMMA membrane dialysis, uraemic patients, or healthy controls. This increased gene expression of SR in monocytes may be associated with the pathogenesis of accelerated atherosclerosis in patients on dialysis with a Cu membrane.

Lovastatin inhibits gene expression of type-I scavenger receptor in THP-1 human macrophages.

Lovastatin, an inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, inhibits the synthesis of mevalonic acid and is widely used as an anti-atherosclerotic drug. The macrophage scavenger receptor (SCR), a trimeric membrane glycoprotein, is postulated to play a key role in atheroma macrophage foam cell formation. HMG-CoA reductase is involved in the control of the synthesis of glycoproteins and farnesylated proteins, including ras proteins, which are involved in the transcriptional regulation of SCR gene expression. Accordingly, we examined whether lovastatin alters the gene expression of SCRs in THP-1 cell derived human macrophages. Lovastatin (5-15 microM) caused a significant dose-related reduction in steady state levels of type-I SCR mRNA in phorbol 12-myristate 13-acetate (PMA)-treated THP-1 cells. The addition of exogenous mevalonate (1 mM) completely restored the lovastatin-induced decrease of type-I SCR mRNA levels. While the addition of the isoprenoid end-product, isopentenyl adenine (50 microM), had little effect on the type-I SCR mRNA levels in lovastatin treated cells, the addition of isoprenoid farnesol (5 microM) largely restored the lovastatin-induced decrease of type-I SCR mRNA levels. Actinomycin D treatment showed that degradation rates of type-I SCR mRNA did not differ between the THP-1 derived cells with and without lovastatin treatment. Nuclear run-on assays showed that lovastatin markedly decreased the transcription of SCR gene in the cells. These results suggest that lovastatin inhibits the transcription of type-I SCR gene by affecting mevalonate metabolism, possibly through the farnesyl-pyrophosphate related end-product(s) in the THP-1-derived macrophages.

Coronary artery to pulmonary artery fistula with two giant saccular aneurysms in an elderly patient determined noninvasively.

An 87-year-old woman was admitted to our hospital on an emergency basis with atypical chest pain and dyspnea. She had a continuous precordial murmur. Electrocardiogram showed no evidence of myocardial ischemia, but chest X-ray showed marked enlargement of the cardiac silhouette and an abnormal calcified vascular structure. Computed tomography of the chest revealed large abnormal masses next to the heart. Two-dimensional echocardiography showed enlargement of the main trunk of the left coronary artery and 2 giant saccular aneurysms. Abnormal diastolic inflow to the main pulmonary trunk was also observed by color flow imaging. These findings were supported by data obtained using magnetic resonance imaging and transesophageal echocardiography. Based on the above findings, we diagnosed this case as a coronary artery fistula originating from the proximal left anterior descending artery associated with 2 giant saccular aneurysms draining into the pulmonary artery. To our knowledge, this is the oldest patient ever reported with such an anomaly. This case emphasizes that a good prognosis is possible even with a very pronounced visible structural abnormality.

Gene expression of endothelin receptors in aortic cells from cyclosporine-induced hypertensive rats.

1. We examined preproendothelin-1, ETA and ETB receptor mRNA levels in aortic endothelial and smooth muscle cells from cyclosporine (CyA)-induced hypertensive rats using the reverse transcription polymerase chain reaction method. 2. Aortic endothelial preproendothelin-1 mRNA expression was about 1.5-fold higher, while that of ETB receptor mRNA was markedly decreased in CyA-treated rats compared with those in controls. 3. The expression of ETA receptor mRNA in smooth muscle cells from CyA-induced hypertensive rats was increased about two-fold over that in cells from control animals. 4. Thus, increased endothelial preproendothelin-1, and ETA receptor mRNA levels in smooth muscle cells, which are concomitant with the decrease in ETB receptor mRNA levels in endothelium, may contribute to CyA-induced hypertension in rats.

Blood pressure regulates platelet-derived growth factor A-chain gene expression in vascular smooth muscle cells in vivo. An autocrine mechanism promoting hypertensive vascular hypertrophy.

To clarify the role of PDGF A-chain in hypertensive vascular hypertrophy of spontaneously hypertensive rats (SHRs), we studied levels of PDGF A-chain gene expression and transcription factors related to the gene in vascular smooth muscle cells (VSMCs) of SHRs in vivo. RNase protection assay and in situ hybridization showed that PDGF A-chain mRNA levels in VSMCs of SHRs were twofold higher than in those of normotensive Wistar-Kyoto rats. Gel retardation assays showed that levels of Sp1 and AP-2 in VSMCs of SHRs were twofold more abundant than in those of Wistar-Kyoto rats. Treatment with four pharmacologically different species of antihypertensive drugs for 2 wk decreased the levels of both PDGF A-chain mRNA and Sp1, but not AP-2 level in VSMCs of SHRs with regression of aortic hypertrophy, indicating that increases in levels of both PDGF A-chain mRNA and Sp1 in VSMCs of SHRs were associated with high blood pressure. These results suggest that high blood pressure is a stimulus which upregulates PDGF A-chain gene expression in VSMCs of SHRs, resulting in an autocrine enhancement in hypertensive vascular hypertrophy, and that the activation of the gene may be mediated through increases in Sp1 in these cells.

Anti-Ro/SS-A antibody-positive interstitial pneumonitis in a non-lupus patient.

We describe a 42-year-old anti-Ro/SS-A antibody positive non-lupus patient who developed interstitial pneumonitis in combination with several common clinical features with previously reported lupus pneumonitis patients whose anti-Ro/SS-A antibodies were positive, while whose antinuclear antibodies were negative. We consider that these patients may belong to a same new clinical entity. A variety of characteristic manifestations related to anti-Ro/SS-A antibodies has been reported in patients with systemic lupus erythematosus (SLE) and other connective tissue diseases [1] [2], and an association between these antibodies with pulmonary parenchymal involvement has been reported by Hedgpeth and Boulware in patients with SLE [3] [4]. We report here a 42-year-old non-lupus male patient with pulmonary fibrosis who presented with anti-Ro/SS-A antibodies.

Modulation of protein kinase C in aorta of spontaneously hypertensive rats with enalapril treatment.

We measured protein kinase C (PKC) activity, levels of PKC alpha enzyme and PKC alpha mRNA in aortic media of spontaneously hypertensive rats (SHR), normotensive Wistar Kyoto rats (WKY) and enalapril treated SHR (enal-SHR) to examine whether hypotensive treatment of enalapril modulates PKC in aortic media of SHR. The cytosolic PKC activity in crude samples of aortic media of SHR was higher than in those of WKY or enal-SHR (p < 0.01) and was closely associated with blood pressure (r = 0.84, p < 0.001). The membrane PKC activity was detected in samples of SHR, but virtually no activity was detected in samples of WKY or enal-SHR. The cytosolic PKC activity in DEAE column purified samples of SHR was also higher than in those of WKY or enal-SHR (p < 0.01). The PKC alpha enzyme levels (74-kDa and 77-kDa protein) detected by immunoblot were higher in SHR than in WKY or enal-SHR (p < 0.01). The mRNA levels of PKC alpha were higher in SHR than in WKY (p < 0.01) and were much decreased in enal-SHR (p < 0.01). Thus, PKC activity, PKC alpha and its mRNA levels were higher in aortic media of SHR than those in WKY and these increased levels were reversed with enalapril treatment. Considering the pivotal roles of PKC in the mechanism of cellular proliferation and the pathogenesis of hypertension, these results provide clues in understanding the pathogenesis of hypertension, mechanisms of vascular hypertrophy in hypertension and the beneficial effects of angiotensin converting enzyme inhibitor in the treatment of hypertension.

[Surgical treatment of coronary artery aneurysm as a complication developed after PTCA].

A 56-year-old man was admitted to our medical center because of acute anterior myocardial infarction. Emergent percutaneous transluminal coronary angioplasty (PTCA) resulted in successful dilatation at the stenotic lesion of the left anterior descending artery (LAD). Three months later, coronary angiography showed not only a restenosis but also an aneurysm formation at the same portion. This lesion was too risky to redo PTCA. We successfully performed coronary bypass grafting with the internal thoracic artery.