Association between Achilles Tendon Softness and Atherosclerotic Cardiovascular Disease in Patients with Familial Hypercholesterolemia.

AIMS: Achilles tendon (AT) xanthomas are a specific physical finding of familial hypercholesterolemia (FH) and AT thickness has been used for its diagnosis and evaluation of its severity. Recently, we reported that the AT of FH patients was softer than that of non-FH patients and the combined use of a cut-off value for AT softness with that for AT thickness improved diagnostic accuracy. However, an association between AT softness and severity of atherosclerosis has not been reported. Accordingly, the present study aimed to investigate whether AT softness was associated with carotid atherosclerosis and presence of atherosclerotic cardiovascular disease (ASCVD) in FH. METHODS: The AT of 176 genetically diagnosed FH patients and 98 non-FH patients was examined to measure AT thickness and the elasticity index (EI) as an indicator for assessing AT softness using ultrasonography. RESULTS: Increased age was associated with AT softness, and overweight was negatively related to AT softness. There were significant inverse correlations between EI and maximum and mean intima-media thickness (IMT) within the common carotid artery only among FH patients. In multiple linear regression analysis, although the relationship between EI and mean IMT was attenuated, the association between EI and maximum IMT remained robust. In logistic regression analysis adjusted for age, sex and traditional cardiovascular risk factors (smoking history, presence of hypertension, presence of diabetes mellitus, overweight, LDL-cholesterol, HDL-cholesterol, and Log triglycerides), EI was associated with presence of ASCVD (Odds ratio per 1-SD increase, 0.37; 95% CI, 0.15 - 0.86; P=0.0252). CONCLUSION: The degree of lipid deposition in the AT of FH patients could be assessed by its thickness as well as its softness. AT softness is not only useful in diagnosing FH but is also associated with the severity of carotid atherosclerosis and presence of ASCVD. In addition, these findings suggest that AT softness would be helpful in risk assessment for FH patients.

Familial Hypercholesterolemia and Lipoprotein Apheresis.

Lipoprotein apheresis has been developed as the treatment for refractory familial hypercholesterolemia (FH) to remove low-density lipoprotein (LDL), which is the main pathogenic factor. Currently, three procedures are available in Japan, including the plasma exchange, double-membrane filtration, and selective LDL adsorption. Selective LDL adsorption, which was developed in Japan, has been one of the most common treatment methods in the world. Lipoprotein apheresis enabled the prevention of atherosclerosis progression even in homozygous FH (HoFH) patients. However, in our observational study, HoFH patients who started lipoprotein apheresis in adulthood had a poorer prognosis than those who started in childhood. Therefore, HoFH patients need to start lipoprotein apheresis as early as possible. Although the indication for lipoprotein apheresis in heterozygous FH (HeFH) patients has been decreasing with the advent of strong statins, our observational study showed that HeFH patients who discontinued lipoprotein apheresis had a poorer prognosis than patients who continued apheresis therapy. These results suggest that it is beneficial for very-high-risk HeFH patients to be treated by lipoprotein apheresis even if their LDL cholesterol is controlled well by lipid-lowering agents. Since launching a new class of lipid-lowering agents, proprotein convertase subtilisin/kexin type 9 (PCSK9) antibody and microsome triglyceride transfer protein inhibitors, the indication for lipoprotein apheresis in FH has been changing. However, despite the development of these drugs, lipoprotein apheresis is still an option with a high therapeutic effect for FH patients with severe atherosclerotic cardiovascular disease.

LDL apheresis in Japan.

LDL apheresis has been developed as the treatment for refractory familial hypercholesterolemia (FH). Currently, plasma exchange, double membrane filtration, and selective LDL adsorption are available in Japan, and selective LDL adsorption is most common method. LDL apheresis can prevent atherosclerosis progression even in homozygous (HoFH). However, in our observational study, HoFH who started LDL apheresis from adulthood had poor prognosis compared with patients who started from childhood. Therefore, as far as possible, HoFH patients need to start LDL apheresis from childhood. Although indication of LDL apheresis in heterozygous FH (HeFH) has been decreasing with the advent of strong statin, our observational study showed that HeFH patients who were discontinued LDL apheresis therapy had poor prognosis compared with patients who were continued apheresis therapy. These results suggest that high risk HeFH need to be treated by LDL apheresis even if their LDLC is controlled by lipid-lowering agents. However, by launching new class of lipid lowering agents, that is, PCSK-9 antibody and MTP inhibitor, indication of LDL-apheresis in FH may be changed near the future. LDL-apheresis can provide symptom relief of peripheral artery disease (PAD). Therefore, PAD patients who have insufficient effect by other therapeutic approach including revascularization are also treated by LDL apheresis. Thus, LDL apheresis is still one of good therapeutic options for severe atherosclerotic diseases in Japan.

Lipoprotein apheresis is essential for managing pregnancies in patients with homozygous familial hypercholesterolemia: Seven case series and discussion.

BACKGROUND AND AIMS: For patients with homozygous familial hypercholesterolemia (HoFH), atherogenic lipoprotein changes and increased stress on cardiovascular system during pregnancy may pose substantial risk for both the mother and her fetus. Although lipoprotein apheresis (LA) is reported as the most effective therapy to control LDL-C levels during pregnancy in HoFH patients, only case reports have been published, and there is no guidance for management. METHODS: We report twelve pregnancies and ten deliveries in seven patients with HoFH, and compare the clinical outcomes between patients who received LA during pregnancy and those who did not. RESULTS: One patient who refused LA during pregnancy died from acute myocardial infarction after delivery. Another patient whose adherence to LA was poor also died of myocardial infarction during pregnancy. One patient who initiated LA at the age of 18 had to discontinue LA due to severe symptoms of angina pectoris during pregnancy. Another had symptoms of nausea, hypotension, and bradycardia with increased levels of serum bradykinin during a dextran sulfate cellulose absorption-based LA procedure. Although two of the other three patients had already had coronary artery disease by the time of pregnancy, early initiation of LA from childhood and good adherence to it during pregnancy resulted in the delivery of healthy infants without adverse effects. CONCLUSIONS: LA is essential for managing pregnancy safely in patients with HoFH. Increasing numbers of documented cases, including ours, will be helpful to guide future therapeutic decisions.

Proprotein convertase subtilisin/kexin 9 V4I variant with LDLR mutations modifies the phenotype of familial hypercholesterolemia.

BACKGROUND: Familial hypercholesterolemia (FH) is caused by mutations in the genes encoding low-density lipoprotein receptor (LDLR), apolipoprotein B, or proprotein convertase subtilisin/kexin 9 (PCSK9). However, FH shows variability of the clinical phenotype modified by other genetic variants or environmental factors. OBJECTIVE: Our objective was to determine the distribution of PCSK9 variants in Japanese FH heterozygotes and to clarify whether those variants and the combination of those variants and LDLR mutations modify the clinical phenotypes. METHODS: A direct sequence analysis was performed for all 18 exons of LDLR gene and 12 exons of PCSK9 gene in 269 clinically diagnosed FH heterozygotes. The serum lipid levels of the carriers of each variant were compared to those of noncarriers. We also assessed Achilles tendon xanthoma and the prevalence of coronary artery disease (CAD) in the patients aged ≥30 years. RESULTS: Eleven PCSK9 variants were detected. There were 4 frequent PCSK9 variants: L21_22insL, A53 V, V4I, and E32 K. The PCSK9 L21_22insL and A53 V were in linkage disequilibrium with each other. There were no significant differences in serum lipids levels and the prevalence of CAD at the age of ≥ 30 years between PCSK9 V4I, L21_22insL/A53 V, or E32 K variant carriers and noncarriers without LDLR mutations. In the patients carrying LDLR mutations and aged ≥ 30 years, the additional PCSK9 V4I variant was linked to a significantly increased prevalence of CAD in accord with the elevation of the LDL-cholesterol level. CONCLUSIONS: The addition of the PCSK9 V4I was suggested to modify the phenotype of patients carrying LDLR mutations by affecting their LDLR metabolism.

Characterization of Autosomal Dominant Hypercholesterolemia Caused by PCSK9 Gain of Function Mutations and Its Specific Treatment With Alirocumab, a PCSK9 Monoclonal Antibody.

BACKGROUND: Patients with PCSK9 gene gain of function (GOF) mutations have a rare form of autosomal dominant hypercholesterolemia. However, data examining their clinical characteristics and geographic distribution are lacking. Furthermore, no randomized treatment study in this population has been reported. METHODS AND RESULTS: We compiled clinical characteristics of PCSK9 GOF mutation carriers in a multinational retrospective, cross-sectional, observational study. We then performed a randomized placebo-phase, double-blind study of alirocumab 150 mg administered subcutaneously every 2 weeks to 13 patients representing 4 different PCSK9 GOF mutations with low-density lipoprotein cholesterol (LDL-C) ≥70 mg/dL on their current lipid-lowering therapies at baseline. Observational study: among 164 patients, 16 different PCSK9 GOF mutations distributed throughout the gene were associated with varying severity of untreated LDL-C levels. Coronary artery disease was common (33%; average age of onset, 49.4 years), and untreated LDL-C concentrations were higher compared with matched carriers of mutations in the LDLR (n=2126) or apolipoprotein B (n=470) genes. Intervention study: in PCSK9 GOF mutation patients randomly assigned to receive alirocumab, mean percent reduction in LDL-C at 2 weeks was 62.5% (P<0.0001) from baseline, 53.7% compared with placebo-treated PCSK9 GOF mutation patients (P=0.0009; primary end point). After all subjects received 8 weeks of alirocumab treatment, LDL-C was reduced by 73% from baseline (P<0.0001). CONCLUSIONS: PCSK9 GOF mutation carriers have elevated LDL-C levels and are at high risk of premature cardiovascular disease. Alirocumab, a PCSK9 antibody, markedly lowers LDL-C levels and seems to be well tolerated in these patients. CLINICAL TRIAL REGISTRATION: URL: http://www.clinicaltrials.gov. Unique Identifier: NCT01604824.

Removal of plasma mature and furin-cleaved proprotein convertase subtilisin/kexin 9 by low-density lipoprotein-apheresis in familial hypercholesterolemia: development and application of a new assay for PCSK9.

CONTEXT: Proprotein convertase subtilisin/kexin 9 (PCSK9) is known to be a good target to decrease LDL cholesterol (LDL-C) and two forms of PCSK9, mature and furin-cleaved PCSK9, circulate in blood. However, it has not been clarified whether and how the levels of each PCSK9 are affected by LDL-apheresis (LDL-A) treatment, a standard therapy in patients with severe forms of familial hypercholesterolemia (FH). OBJECTIVE: Our objective was to investigate the differences in LDL-A-induced reduction of mature and furin-cleaved PCSK9 between homozygous and heterozygous FH, and between dextran sulfate (DS) cellulose adsorption and double membrane (DM) columns and to clarify the mechanism of their removal. DESIGN: A sandwich ELISA to measure two forms of PCSK9s using monoclonal antibodies was developed. Using the ELISA, PCSK9 levels were quantified before and after LDL-A with DS columns in 7 homozygous and 11 heterozygous FH patients. A crossover study between the two column types was performed. The profiles of PCSK9s were analyzed after fractionation by gel filtration chromatography. Immunoprecipitation of apolipoprotein B (apoB) in FH plasma was performed. RESULTS: Both mature and furin-cleaved PCSK9s were significantly decreased by 55-56% in FH homozygotes after a single LDL-A treatment with DS columns, and by 46-48% or 48-56% in FH heterozygotes after treatment with DS or DM columns. The reduction ratios of LDL-C were strongly correlated with that of PCSK9 in both FH homozygotes and heterozygotes. In addition, more than 80% of plasma PCSK9s were in the apoB-deficient fraction and a significant portion of mature PCSK9 was bound to apoB, as shown by immunoprecipitation. CONCLUSIONS: Both mature and furin-cleaved PCSK9s were removed by LDL-A in homozygous and heterozygous FH either by binding to apoB or by other mechanisms. The ELISA method to measure both forms of plasma PCSK9 would be useful for investigating physiological or pathological roles of PCSK9.

Renal insufficiency without albuminuria is associated with peripheral artery atherosclerosis and lipid metabolism disorders in patients with type 2 diabetes.

AIMS: A high prevalence of a low glomerular filtration rate (GFR) has recently been reported in patients with diabetes without albuminuria. We aimed to clarify the clinical characteristics of such patients, including the associations between these characteristics and atherosclerosis. METHODS: We investigated the correlations between the estimated GFR (eGFR) and lipid profiles, the ankle-brachial index (ABI) and the intima-media thickness (IMT) in 450 patients with type 2 diabetes without macroalbuminuria. RESULTS: The prevalence of renal insufficiency (RI) (GFR ＜60 mL/min/1.73 m(2)) in the patients without albuminuria was 19.1%. The ABI values of the patients with RI were significantly lower than those of the patients without RI, regardless of the presence of microalbuminuria, while there were no significant differences in IMT between the patients with and without RI. In a multivariate analysis, a low ABI was found to be significantly associated with a low eGFR, independent of age, sex, smoking, history of hypertension and/or dyslipidemia and duration of diabetes (ß=0.134, p=0.013), whereas no significant associations were observed between the ABI and the urinary albumin excretion rate (UAER). The ApoB/LDL-C ratios and levels of ApoC3 were significantly higher in the patients with RI than those observed in the patients without RI, regardless of the presence of albuminuria. CONCLUSIONS: RI without albuminuria is closely associated with atherosclerosis of the peripheral arteries in diabetic patients. Furthermore, alterations in lipid metabolism may underlie this association.

Defining patients at extremely high risk for coronary artery disease in heterozygous familial hypercholesterolemia.

AIM: Heterozygous patients with familial hypercholesterolemia (FH) are known to be associated with a high risk of coronary artery disease (CAD), which is a major determinant of their clinical outcome. The prognosis of heterozygous FH patients substantially varies, being dependent on the level of their CAD risk, and their therapeutic regimen should be individualized. We assessed critical levels of LDL-cholesterol (LDL-C) and Achilles tendon thickness (ATT) to identify heterozygous FH patients at "very high" risk for CAD. METHODS: One hundred and nine heterozygous FH patients who had no history of CAD and had had their plasma lipid profile and ATT assessed before treatment were followed up until their first CAD event or 31 December 2010. Multivariable logistic regression models were used to analyze the correlation of LDL-C and/or ATT levels with the risk of developing CAD. RESULTS: During the follow-up period, 21 of the 109 patients had a CAD event, diagnosed by coronary angiogram. Individuals in the highest tertile of LDL-C had a CAD risk 8.29-fold higher than those in the lowest tertile. Individuals in the highest tertile of the ATT group had a 7.82-fold higher CAD risk than those in the lowest tertile. Those who had either LDL-C ≥ 260 mg/dL or ATT ≥ 14.5 had a 23.94-fold higher CAD risk than those with LDL-C < 260 mg/dL and ATT <14.5 mm. CONCLUSIONS: In heterozygous FH patients, LDL-C 260 mg/dL or higher and/or ATT 14.5 mm or thicker are useful markers for extracting patients at "very high" risk for CAD.

Impact of statin treatment on the clinical fate of heterozygous familial hypercholesterolemia.

AIM: Familial hypercholesterolemia (FH) patients are at particular risk for premature coronary artery disease (CAD) caused by high levels of low density lipoprotein (LDL). Administration of statins enabled us to reduce LDL-C levels in heterozygous FH patients. To evaluate the impact of statins on the clinical fate of heterozygous FH, a retrospective study was performed. METHODS: We analyzed the clinical influence of statins on age at the first clinical onset of CAD in 329 consecutive FH patients referred to the lipid clinic of the National Cardiovascular Center. Among 329 heterozygous FH patients, the onset of CAD was identified in 101. RESULTS: The age at onset of CAD was 58.8+/-12.5 years in the 25 patients on statins at onset, significantly higher than that in the 76 patients not on statins (47.6+/-10.5 years) (p <0.001). The average age at CAD onset was significantly higher after widespread use of statins (54.2+/-13.2 years in 48 patients, Group 1) compared to before October 1989 when statins were approved in Japan (46.9+/-9.6 years in 53 patients; Group 2, p=0.002). A significant difference was seen between Groups 1 and 2 in the variables, including sex, prevalence of smoking habit, LDL-C, and the use of statins, aspirin and probucol. After adjusting for these variables, only statin use was independently associated with the difference in age at CAD onset by multivariable analysis. CONCLUSION: Statins have improved the clinical course of patients with heterozygous FH.

Altered gene expression related to glomerulogenesis and podocyte structure in early diabetic nephropathy of db/db mice and its restoration by pioglitazone.

Glomerular injury plays a pivotal role in the development of diabetic nephropathy. To elucidate molecular mechanisms underlying diabetic glomerulopathy, we compared glomerular gene expression profiles of db/db mice with those of db/m control mice at a normoalbuminuric stage characterized by hyperglycemia and at an early stage of diabetic nephropathy with elevated albuminuria, using cDNA microarray. In db/db mice at the normoalbuminuric stage, hypoxia-inducible factor-1alpha (HIF-1alpha), ephrin B2, glomerular epithelial protein 1, and Pod-1, which play key roles in glomerulogenesis, were already upregulated in parallel with an alteration of genes related to glucose metabolism, lipid metabolism, and oxidative stress. Podocyte structure-related genes, actinin 4alpha and dystroglycan 1 (DG1), were also significantly upregulated at an early stage. The alteration in the expression of these genes was confirmed by quantitative RT-PCR. Through pioglitazone treatment, gene expression of ephrin B2, Pod-1, actinin 4alpha, and DG1, as well as that of oxidative stress and lipid metabolism, was restored concomitant with attenuation of albuminuria. In addition, HIF-1alpha protein expression was partially attenuated by pioglitazone. These results suggest that not only metabolic alteration and oxidative stress, but also the alteration of gene expression related to glomerulogenesis and podocyte structure, may be involved in the pathogenesis of early diabetic glomerulopathy in type 2 diabetes.

Role of prostaglandin E receptor EP1 subtype in the development of renal injury in genetically hypertensive rats.

One of the major causes of end-stage renal diseases is hypertensive renal disease, in which enhanced renal prostaglandin (PG) E2 production has been shown. PGE2, a major arachidonic acid metabolite produced in the kidney, acts on 4 receptor subtypes, EP1 through EP4, but the pathophysiological importance of the PGE2/EP subtypes in the development of hypertensive renal injury remains to be elucidated. In this study, we investigated whether an orally active EP1-selective antagonist (EP1A) prevents the progression of renal damage in stroke-prone spontaneously hypertensive rats (SHRSP), a model of human malignant hypertension. Ten-week-old SHRSP, with established hypertension but with minimal renal damage, were given EP1A or vehicle for 5 weeks. After the treatment period, vehicle-treated SHRSP showed prominent proliferative lesions in arterioles, characterized by decreased alpha-smooth muscle actin expression in multilayered vascular smooth muscle cells. Upregulation of transforming growth factor-beta expression and tubulointerstitial fibrosis were also observed in vehicle-treated SHRSP. All these changes were dramatically attenuated in EP1A-treated SHRSP. Moreover, EP1A treatment significantly inhibited both increase in urinary protein excretion and decrease in creatinine clearance but had little effect on systemic blood pressure. These findings indicate that the PGE2/EP1 signaling pathway plays a crucial role in the development of renal injury in SHRSP. This study opens a novel therapeutic potential of selective blockade of EP1 for the treatment of hypertensive renal disease.

Roles of connective tissue growth factor and prostanoids in early streptozotocin-induced diabetic rat kidney: the effect of aspirin treatment.

BACKGROUND: Connective tissue growth factor (CTGF) is a cysteine-rich growth factor induced by transforming growth factor-beta (TGF-beta) and is thought to play a critical role in TGF-beta-stimulated extracellular matrix accumulation. To explore its involvement in early diabetic nephropathy, we investigated the time course of CTGF gene expression and its regulation in streptozotocin (STZ)-induced diabetic rat kidney. METHODS: Northern blot analysis for CTGF, TGF-beta, and fibronectin expression was performed in the glomeruli of STZ-induced diabetic rats from 3 days to 12 weeks after the induction of diabetes, together with histological examination. To investigate the role of prostanoids in this process, aspirin was administered in one group of diabetic rats. Furthermore, CTGF expression was analyzed in rat mesangial cells cultured under high-glucose conditions. RESULTS: Glomerular expression of CTGF and TGF-beta1 mRNA was coordinately upregulated as early as day 3, followed by fibronectin induction and mesangial matrix accumulation. Chronic aspirin treatment in diabetic rats significantly attenuated mesangial expansion, and effectively suppressed CTGF induction, as well as inhibiting the upregulation of TGF-beta1 and fibronectin expression. In cultured mesangial cells, aspirin treatment abolished high glucose-stimulated CTGF upregulation. CONCLUSIONS: These results indicate that CTGF expressed in the glomeruli is upregulated in the early stage of STZ-induced diabetic nephropathy in rats, and could be a critical mediator of the development of diabetic glomerulosclerosis. In addition, the modulatory effects of aspirin during this process suggest a role of the cyclooxygenase pathway in the progression of diabetic nephropathy.

Long-term effect of low-density lipoprotein apheresis in patients with homozygous familial hypercholesterolemia.

Patients that are homozygous for familial hypercholesterolemia (FH) exhibit severe hypercholesterolemia, cutaneous and tendon xanthomas and premature atherosclerosis beginning in childhood. They are resistant to drug therapy and low-density lipoprotein (LDL) apheresis is the practical treatment. Here we review the technique of LDL apheresis treatment, the long-term effects of LDL apheresis, the effect of apheresis on pregnancy, and the drugs that have proven beneficial in patients with homozygous FH. We also record our experiences of treating eight homozygous FH patients using the LDL apheresis treatment. Among the eight patients, one has been free from cardiovascular disease and two patients have each regressed once. In two patients, aortic valve stenosis developed and the other two patients died for acute myocardial infarction. Furthermore, two patients delivered healthy babies in spite of coronary artery disease. Thus, LDL apheresis therapy has the possibility of preventing the progression of atherosclerosis, but the prognosis assessed by long-term observation is still not satisfactory. A recent clinical trial showed some efficacy of the combination therapy of LDL apheresis and atorvastatin for reducing serum cholesterol levels in homozygous FH, suggesting that this combination therapy may be useful for prevention of atherosclerosis in patients homozygous for FH.

Angiogenic protein Cyr61 is expressed by podocytes in anti-Thy-1 glomerulonephritis.

Dynamic recovery of glomerular structure occurs after severe glomerular damage in anti-Thy-1 glomerulonephritis (Thy-1 GN), but its mechanism remains to be investigated. To identify candidate genes possibly involved in glomerular reconstruction, screening was performed for genes that are specifically expressed by podocytes and are upregulated in glomeruli of Thy-1 GN. Among them, cysteine-rich protein 61 (Cyr61 or CCN1), a soluble angiogenic protein belonging to the CCN family, was identified. By Northern blot analysis, Cyr61 mRNA was markedly upregulated in glomeruli of Thy-1 GN from day 3 through day 7, when mesangial cell migration was most prominent. By in situ hybridization and immunohistochemistry, Cyr61 mRNA and protein were expressed by proximal straight tubules and afferent and efferent arterioles in normal rat kidneys and were intensely upregulated at podocytes in Thy-1 GN. Platelet-derived growth factor-BB (PDGF-BB) and transforming growth factor-beta1 (TGF-beta1), of which the gene expression in the glomeruli of Thy-1 GN was upregulated in similar time course as Cyr61, induced Cyr61 mRNA expression in cultured podocytes. Furthermore, supernatant of Cyr61-overexpressing cells inhibited PDGF-induced mesangial cell migration. In conclusion, it is shown that Cyr61 is strongly upregulated at podocytes in Thy-1 GN possibly by PDGF and TGF-beta. Cyr61 may be involved in glomerular remodeling as a factor secreted from podocytes to inhibit mesangial cell migration.

Role of connective tissue growth factor in fibronectin expression and tubulointerstitial fibrosis.

Connective tissue growth factor (CTGF) is one of the candidate factors mediating downstream events of transforming growth factor-beta (TGF-beta), but its role in fibrogenic properties of TGF-beta and in tubulointerstitial fibrosis has not yet been clarified. Using unilateral ureteral obstruction (UUO) in rats, we analyzed gene expression of TGF-beta1, CTGF, and fibronectin. We further investigated the effect of blockade of endogenous CTGF on TGF-beta-induced fibronectin expression in cultured rat renal fibroblasts by antisense oligodeoxynucleotide (ODN) treatment. After UUO, CTGF mRNA expression in the obstructed kidney was significantly upregulated subsequent to TGF-beta1, followed by marked induction of fibronectin mRNA. By in situ hybridization, CTGF mRNA was detected mainly in the interstitial fibrotic areas and tubular epithelial cells as well as in parietal glomerular epithelial cells in the obstructed kidney. The interstitial cells expressing CTGF mRNA were also positive for alpha-smooth muscle actin. CTGF antisense ODN transfected into cultured renal fibroblasts significantly attenuated TGF-beta-stimulated upregulation of fibronectin mRNA and protein compared with control ODN transfection, together with inhibited synthesis of type I collagen. With the use of a reporter assay, rat fibronectin promoter activity was increased by 2.5-fold with stimulation by TGF-beta1, and this increase was abolished with antisense CTGF treatment. Thus CTGF plays a crucial role in fibronectin synthesis induced by TGF-beta, suggesting that CTGF blockade could be a possible therapeutic target against tubulointerstitial fibrosis.

Overexpression of brain natriuretic peptide in mice ameliorates immune-mediated renal injury.

One of major causes of end-stage renal disease is glomerulonephritis, the treatment of which remains difficult clinically. It has already been shown that transgenic mice that overexpress brain natriuretic peptide (BNP), with a potent vasorelaxing and natriuretic property, have ameliorated glomerular injury after subtotal nephrectomy. However, the role of natriuretic peptides in immune-mediated renal injury still remains unknown. Therefore, the effects of chronic excess of BNP on anti-glomerular basement membrane nephritis induced in BNP-transgenic mice (BNP-Tg) were investigated and the mechanisms how natriuretic peptides act on mesangial cells in vitro were explored. After induction of nephritis, severe albuminuria (approximately 21-fold above baseline), tissue damage, including mesangial expansion and cell proliferation, and functional deterioration developed in nontransgenic littermates. In contrast, BNP-Tg exhibited much milder albuminuria (approximately fourfold above baseline), observed only at the initial phase, and with markedly ameliorated histologic and functional changes. Up-regulation of transforming growth factor-beta (TGF-beta) and monocyte chemoattractant protein-1 (MCP-1), as well as increased phosphorylation of extracellular signal-regulated kinase (ERK), were also significantly inhibited in the kidney of BNP-Tg. In cultured mesangial cells, natriuretic peptides counteracted the effects of angiotensin II with regard to ERK phosphorylation and fibrotic action. Because angiotensin II has been shown to play a pivotal role in the progression of nephritis through induction of TGF-beta and MCP-1 that may be ERK-dependent, the protective effects of BNP are likely to be exerted, at least partly, by antagonizing the renin-angiotensin system locally. The present study opens a possibility of a novel therapeutic potential of natriuretic peptides for treating immune-mediated renal injury.