CRRL269.

RATIONALE: Acute kidney injury (AKI) has a high prevalence and mortality in critically ill patients. It is also a powerful risk factor for heart failure incidence driven by hemodynamic changes and neurohormonal activation. However, no drugs have been approved by the Food and Drug Administration. Endogenous pGC-A (particulate guanylyl cyclase A receptor) activators were reported to preserve renal function and improve mortality in AKI patients, although hypotension accompanied by pGC-A activators have limited their therapeutic potential. OBJECTIVE: We investigated the therapeutic potential of a nonhypotensive pGC-A activator/designer natriuretic peptide, CRRL269, in a short-term, large animal model of ischemia-induced AKI and also investigated the potential of uCNP (urinary C-type natriuretic peptide) as a biomarker for AKI. METHODS AND RESULTS: We first showed that CRRL269 stimulated cGMP generation, suppressed plasma angiotensin II, and reduced cardiac filling pressures without lowering blood pressure in the AKI canine model. We also demonstrated that CRRL269 preserved glomerular filtration rate, increased renal blood flow, and promoted diuresis and natriuresis. Further, CRRL269 reduced kidney injury and apoptosis as evidenced by ex vivo histology and tissue apoptosis analysis. We also showed, compared with native pGC-A activators, that CRRL269 is a more potent inhibitor of apoptosis in renal cells and induced less decreases in intracellular Ca2+ concentration in vascular smooth muscle cells. The renal antiapoptotic effects were at least mediated by cGMP/PKG pathway. Further, CRRL269 inhibited proapoptotic genes expression using a polymerase chain reaction gene array. Additionally, we demonstrated that AKI increased uCNP levels. CONCLUSIONS: Our study supports developing CRRL269 as a novel renocardiac protective agent for AKI treatment.

Inspiratory Muscle Weakness is Associated With Exercise Intolerance in Patients With Heart Failure With Preserved Ejection Fraction: A Preliminary Study.

BACKGROUND: The relationship between inspiratory muscle weakness (IMW) and exercise intolerance in patients with heart failure with preserved ejection fraction (HFpEF) remains unestablished. METHODS AND RESULTS: The present study enrolled 40 patients with HFpEF (EF ≥45%). IMW was defined as maximum inspiratory pressure <70% normal predicted values. The function of the diaphragm was assessed by means of ultrasound measurement of muscle thickening of the diaphragm. IMW was prevalent in 27.5% of patients. Patients with IMW had significantly lower vital capacity relative to normal predicted values (%VC), lower knee extensor muscle strength in relation to body weight (%KEMS), poorer nutritional status as assessed by means of the Geriatric Nutritional Risk Index, and shorter 6-minute walk distance (6MWD) compared with patients without IMW (all P < .05). Impaired diaphragm muscle thickening at end-inspiration (median value < 3.9 mm) was significantly associated with a high prevalence of IMW and reduced 6MWD (all P < .05). Subgroup analysis showed that IMW was accompanied by a further decrease in 6MWD in patients with restrictive pulmonary dysfunction (%VC <80%) or lower-limb muscle weakness (median %KEMS <30%; all P < .05). CONCLUSIONS: IMW is associated with exercise intolerance in patients with HFpEF.

Renal shear wave velocity by acoustic radiation force impulse did not reflect advanced renal impairment.

AIM: Acoustic radiation force impulse is a noninvasive method for evaluating tissue elasticity on ultrasound. Renal shear wave velocity measured by this technique has not been fully investigated in patients with renal disease. The aim of the present study was to compare renal shear wave velocity in end-stage renal disease patients and that in patients without chronic kidney disease and to investigate influencing factors. METHODS: Renal shear wave velocities were measured in 59 healthy young subjects (control group), 31 subjects without chronic kidney disease (non-CKD group), and 39 end-stage renal disease patients (ESRD group). Each measurement was performed 10 times at both kidneys, and the mean value of eight of 10 measurements, excluding the maximum and minimum values, was compared. RESULTS: Renal shear wave velocity could be measured in all subjects. Renal shear wave velocity in the control group was higher than in the non-CKD group and in the ESRD group, and no difference was found between the non-CKD group and the ESRD group. Age and depth were negatively correlated to the renal shear wave velocity. In multiple regression analysis, age and depth were independent factors for renal shear wave velocity, while renal impairment was not. There was no difference between the non-CKD group and the ESRD group, even when ages were matched and depth was adjusted. CONCLUSION: Renal shear wave velocity was not associated with advanced renal impairment. However, it reflected alteration of renal aging, and this technique may be useful to detect renal impairment in the earlier stages.

Clinical scenario 1 is associated with winter onset of acute heart failure.

BACKGROUND: Several reports have evaluated the association between seasonal variation and acute heart failure (AHF) onset. Cold weather may induce AHF, but the clinical characteristics of patients susceptible to AHF during winter have not been established. Clinical Scenario (CS) is used in the early clinical management of AHF, so we investigated the relationship between CS classification and winter onset of AHF in Japan. METHODS AND RESULTS: We enrolled 582 patients hospitalized for AHF and compared the frequency of AHF among the 4 seasons in each CS group to clarify the clinical characteristics of the winter onset group. Significant increase of AHF during winter was seen in CS1 (systolic blood pressure [SBP] (>140 mmHg) (P=0.01) but not in CS2 (SBP ≥ 100 and ≤ 140 mmHg) or CS3 (SBP <100 mmHg). CS1 patients were divided into winter and other season admission groups. In multivariate analysis, only lack of loop diuretic use was associated with winter admission of CS1 patients (odds ratio 0.562, 95% confidence interval: 0.256-0.798, P=0.006). CONCLUSIONS: Winter predominance of AHF was seen only in CS1, and lack of loop diuretic use was a risk factor for winter onset. Future studies are necessary to confirm whether loop diuretics are useful in preventing AHF with CS1 in winter.

Depletion of Uric Acid Due to SLC22A12 (URAT1) Loss-of-Function Mutation Causes Endothelial Dysfunction in Hypouricemia.

BACKGROUND: Uric acid (UA) serves as an antioxidant in vascular endothelial cells. UA transporter 1 (URAT1) encoded by SLC22A12 is expressed in the kidney and vessels and its loss of function causes hypouricemia. The purpose of this study was to examine whether there is any endothelial dysfunction in patients with hypouricemia. METHODS AND RESULTS: Twenty-six patients with hypouricemia (<2.5 mg/dl) and 13 healthy control subjects were enrolled. Endothelial function was evaluated using flow-mediated dilation (FMD). mRNA of UA transporters expressed in cultured human umbilical endothelial cells (HUVEC) was detected on RT-PCR. There was a positive correlation between FMD and serum UA in the hypouricemia group. URAT1 loss-of-function mutations were found in the genome of 21 of 26 patients with hypouricemia, and not in the other 5. In the hypouricemia groups, serum UA in homozygous and compound heterozygous patients was significantly lower than in other groups, suggesting that severity of URAT1 dysfunction may influence the severity of hypouricemia. Thirteen of 16 hypouricemia subjects with homozygous and compound heterozygote mutations had SUA <0.8 mg/dl and their FMD was lower than in other groups. HUVEC do not express mRNA of URAT1, suggesting the null role of URAT1 in endothelial function. CONCLUSIONS: Depletion of UA due to SLC22A12/URAT1 loss-of-function mutations causes endothelial dysfunction in hypouricemia patients.

Multidisciplinary intensive education in the hospital improves outcomes for hospitalized heart failure patients in a Japanese rural setting.

BACKGROUND: Heart failure (HF) patients living in rural areas have a lack of HF knowledge and poor self-care because of limited medical care access. Multidisciplinary education to improve self-care behavior is indispensable for such patients. The present study evaluated whether intensive inpatient education improved outcomes of hospitalized HF patients in a Japanese rural setting. METHODS: An inpatient HF management program based on multidisciplinary team intervention was applied to hospitalized HF patients in a Japanese rural area. We defined patients treated within the program from May 2009 to April 2011 as the intervention group (n = 144), and those treated with the usual care from May 2006 to April 2009 as the usual care group (n = 133). The composite endpoints of HF hospitalization and all-cause mortality were compared between the two groups. RESULTS: Compared with patients in the usual care group, those in the intervention group more often received the optimal interventions such as discharge use of ß-blockers, cardiac rehabilitation, pre-discharge diagnostic tests, and multidisciplinary intensive education including nurse-led patient education, pharmacist's medication teaching, and dietitian's nutritional guidance (all P < 0.05). The incidence of the composite endpoints significantly decreased after introducing the program (P < 0.001). Among a number of interventions, multidisciplinary intensive education was the most effective intervention to improve the primary outcome (P < 0.001). CONCLUSIONS: Multidisciplinary intensive education is a key strategy for helping improve the outcome for Japanese HF patients in a rural setting. Our data may give a positive impact on the improvement of healthcare system in Japan.

Discharge use of carvedilol is associated with higher survival in Japanese elderly patients with heart failure regardless of left ventricular ejection fraction.

Previous clinical trials have proven beneficial effects of beta-blockers in patients with heart failure (HF) with reduced ejection fraction (EF). However, those studies excluded elderly patients from the subjects or included only a small number of them. We assessed whether beta-blocker treatment with carvedilol improves survival in elderly patients with HF regardless of left ventricular EF (LVEF). We retrospectively analyzed a total of 189 patients older than 75 years who were hospitalized with HF from January 2004 to December 2010. Of these, 84 patients (44%) had been treated with carvedilol at discharge. Patients treated with carvedilol were younger, were less likely to have chronic obstructive pulmonary disease, and had lower LVEF compared with those without carvedilol (all P < 0.05). During the median follow-up of 2.5 years after discharge, 92 patients died. Cox hazard analysis showed that, even after adjustment for covariates, carvedilol significantly decreased all-cause mortality in this cohort (P < 0.01). Furthermore, a beneficial effect on outcome was found in patients with reduced (LVEF ≤ 40%) and preserved (LVEF > 40%) EF (all P < 0.05). In conclusion, Beta-blockers may provide beneficial effects on Japanese elderly patients with HF regardless of LVEF.

Geriatric nutritional risk index predicts functional dependency and mortality in patients with heart failure with preserved ejection fraction.

BACKGROUND: The clinical significance of nutritional risk assessment in patients with heart failure with preserved ejection fraction (HFpEF) remains undefined. Geriatric nutritional risk index (GNRI) is a simple nutritional assessment tool for elderly subjects. Its predictive value was evaluated in patients with HFpEF, a common HF phenotype in the elderly population. METHODS AND RESULTS: The present study enrolled 152 consecutive patients (mean age, 77 ± 11 years; male, 53.9%) who were hospitalized with HFpEF at the authors' institution. GNRI on admission was calculated as follows: 14.89 × serum albumin (g/dl)+41.7×body mass index/22. Characteristics and mortality (median follow-up of 2.1 years) were compared between 2 groups: low GNRI (<92) with moderate or severe nutritional risk; and high GNRI (≥ 92) with no or low nutritional risk. Patients in the low-GNRI group were more often female, and had lower serum hemoglobin and sodium, but higher serum blood urea nitrogen (BUN), C-reactive protein, and B-type natriuretic peptide (BNP) compared to those in the high-GNRI group (P<0.05, respectively). Physical activity at discharge measured by Barthel index was significantly lower in the low-GNRI group than the high-GNRI group (P<0.05). On Cox hazard analysis, lower GNRI predicted increased mortality independent of age, gender, prior HF hospitalization, and higher BUN and BNP (P<0.01). CONCLUSIONS: GNRI may be useful for predicting functional dependency and mortality in patients with HFpEF.

Impact of Hyper- and Hypo-Uricemia on Kidney Function.

Uric acid (UA) forms monosodium urate (MSU) crystals to exert proinflammatory actions, thus causing gout arthritis, urolithiasis, kidney disease, and cardiovascular disease. UA is also one of the most potent antioxidants that suppresses oxidative stress. Hyper andhypouricemia are caused by genetic mutations or polymorphism. Hyperuricemia increases urinary UA concentration and is frequently associated with urolithiasis, which is augmented by low urinary pH. Renal hypouricemia (RHU) is associated with renal stones by increased level of urinary UA, which correlates with the impaired tubular reabsorption of UA. Hyperuricemia causes gout nephropathy, characterized by renal interstitium and tubular damage because MSU precipitates in the tubules. RHU is also frequently associated with tubular damage with elevated urinary beta2-microglobulin due to increased urinary UA concentration, which is related to impaired tubular UA reabsorption through URAT1. Hyperuricemia could induce renal arteriopathy and reduce renal blood flow, while increasing urinary albumin excretion, which is correlated with plasma xanthine oxidoreductase (XOR) activity. RHU is associated with exercise-induced kidney injury, since low levels of SUA could induce the vasoconstriction of the kidney and the enhanced urinary UA excretion could form intratubular precipitation. A U-shaped association of SUA with organ damage is observed in patients with kidney diseases related to impaired endothelial function. Under hyperuricemia, intracellular UA, MSU crystals, and XOR could reduce NO and activate several proinflammatory signals, impairing endothelial functions. Under hypouricemia, the genetic and pharmacological depletion of UA could impair the NO-dependent and independent endothelial functions, suggesting that RHU and secondary hypouricemia might be a risk factor for the loss of kidney functions. In order to protect kidney functions in hyperuricemic patients, the use of urate lowering agents could be recommended to target SUA below 6 mg/dL. In order to protect the kidney functions in RHU patients, hydration and urinary alkalization may be recommended, and in some cases an XOR inhibitor might be recommended in order to reduce oxidative stress.

The Clock-Drawing Test as a Useful Screening Assessment of Preoperative Cognitive Impairment with Readmission After Transcatheter Aortic Valve Implantation.

Background: Transcatheter aortic valve implantation (TAVI) has recently become more common as a treatment for severe, symptomatic aortic stenosis (AS). Cognitive impairment (CI) is strongly associated with the prognosis of TAVI patients. However, some cognitive assessments currently in use are difficult to perform routinely in the clinical setting. To easier CI evaluation, we investigated whether CI using the clock-drawing test (CDT), one part of the Mini-Cog, affects the postoperative prognosis of TAVI patients with AS. Methods: The present study enrolled 52 patients (median age, 85 years; 28.8% male) who underwent TAVI and were discharged between 2019 and 2021. The outcome was readmission for all causes within one year of discharge and patients were grouped according to whether they were readmitted or not. Cognitive function was assessed using the Mini-Cog which combines verbal playback and CDT. Results: Of the 52, 11 patients (21.2%) comprised readmission group, including 4 (36.4%) each for fracture and infection, and 1 (9.1%) each for heart failure, subdural hematoma, and pneumothorax. Median Mini-Cog score was lower in the readmission group than in the non-readmission group (4 vs. 5; P < 0.05). The frequency of Mini-Cog score < 3 (indicative of CI) and CDT failure were significantly higher in the readmission group than in the non-readmission group, respectively (46% vs. 7%, P < 0.01) (46% vs. 12%, P < 0.05). Both of Mini-Cog score < 3 and CDT failure were independently associated with readmission. The areas under the curve showed CDT was an indicator of readmission with similar accuracy to the Mini-Cog score < 3. Kaplan-Meier curves showed significant differences in readmission after 1 year between the 2 Mini-Cog groups with scores of < 3 or ≥ 3 points and CDT failure and success. Conclusion: The CDT may be a very easy and simple screening assessment of preoperative CI with readmission within one year after TAVI.

Characterization of Urate Metabolism and Complications of Patients with Renal Hypouricemia.

Objective Both renal hypouricemia (RHU) and gout are associated with renal dysfunction and urolithiasis. The difference in renal complications associated with RHU and gout, however, has not been studied. We characterized the urate metabolism and complications of patients with RHU and compared them with patients with gout. Methods Eighteen patients with RHU who had a serum uric acid (SUA) level <2 mg/dL (10 men and 8 women), 44 patients with gout (44 men) and 16 normouricemic patients (4 men and 12 women) were included. The blood and urinary biochemical data were evaluated. A genetic analysis of uric acid transporter 1 (URAT1) was also conducted in 15 cases with RHU. Results The SUA level of RHU was 0.9±0.5/mg/dL, and the Uur/Ucr and Cur/Ccr were 0.56±0.14% and 45.7±18.0%, respectively. A genetic analysis of URAT1 in 15 RHU patients showed that 13 harbored a URAT1 gene mutation, whereas 2 harbored the wild-type gene. The SUA level was significantly lower in RHU patients (n=11) than in either gout patients (n=44) or normouricemic patients (n=16). This reduction was accompanied by the elevation of Cua/Ccr. Urinary beta 2-microglobulin levels were higher in RHU patients than in gout or normouricemia patients. Cua/Ccr correlated with normalized urinary beta 2-microglobulin levels. The prevalence of urolithiasis was 18.2% in RHU cases and 6.8% in gout cases. A homozygous URAT1 mutation was associated with urolithiasis. Conclusion Besides urolithiasis, RHU can be associated with tubular dysfunction, such as elevated urinary beta 2-microglobulin levels.

Xanthinuria Type 1 with a Novel Mutation in Xanthine Dehydrogenase and a Normal Endothelial Function.

Whether or not extremely low levels of serum uric acid (SUA) in xanthinuria are associated with impairment of the endothelial function and exercise-induced acute kidney injury (EIAKI) is unclear. A 59-year-old woman without EIAKI or urolithiasis had undetectable levels of UA in serum and urine and elevated levels of hypoxanthine and xanthine in urine. A genetic analysis revealed homozygous mutations in the XDH gene [c.1585 C>T (p. Gln529*)]. Flow-mediated dilation was within the normal range. This is the first report of a case with extremely low levels of SUA, xanthinuria with novel mutations of xanthine dehydrogenase (XDH) and a normal endothelial function.

Addition of losartan to angiotensin-converting enzyme inhibitors improves insulin resistance in patients with chronic heart failure treated without ß-blockers.

BACKGROUND: Angiotensin II and insulin resistance (IR) have clinical implications in the pathophysiology of chronic heart failure (CHF). However, it is still unclear whether the combination of an angiotensin-receptor blocker and angiotensin-converting enzyme inhibitor (ACEI) improves IR in CHF patients who do not receive ß-blockers. Thus, the aim of the present study was to evaluate the effects of losartan on glucose metabolism and inflammatory cytokines in CHF patients treated with ACEI but not ß-blockers. METHODS AND RESULTS: The effect of losartan treatment for 16 weeks on IR was analyzed in 16 CHF patients in a randomized crossover trial. Insulin level and homeostasis model IR index (HOMA-IR) decreased significantly (P<0.05), but fasting plasma glucose did not change significantly. Serum tumor necrosis factor (TNF)-α, interleukin (IL)-6, and monocyte chemoattractant protein (MCP)-1 levels were significantly decreased with losartan (P<0.05). Furthermore, the changes in IL-6 and MCP-1 levels were significantly correlated with the reduction in HOMA-IR (P<0.05), but the change in TNF-α levels was not significantly correlated. CONCLUSIONS: The addition of losartan to ACEI therapy improved IR and decreased inflammatory cytokines in CHF patients who did not receive ß-blockers.

A simple risk score to predict in-hospital death of elderly patients with acute decompensated heart failure--hypoalbuminemia as an additional prognostic factor.

BACKGROUND: Risk stratification for elderly patients with acute decompensated heart failure (ADHF) may help clinicians to select the appropriate therapy and raise the quality of care. METHODS AND RESULTS: The present study enrolled 349 patients aged over 65 years who were hospitalized with ADHF from January 2004 to October 2008. Five independent prognostic factors were identified by multivariate logistic regression analysis, and each factor was assigned a number of points proportional to its regression coefficient: prior heart failure hospitalization (2 points), sodium or=35 mg/dl (2 points), albumin or=980 pg/ml (2 points); in particular, hypoalbuminemia was identified as the strongest prognostic factor. The patients were stratified into 3 groups: low risk (0-4 points), moderate risk (5-7 points), and high risk (8-11 points). The respective in-hospital mortality rates were 1.6%, 15.8%, and 42.1% (P<0.05). CONCLUSIONS: In addition to known prognostic factors, hypoalbuminemia may provide important information for elderly patients with ADHF. A simple risk score may help to stratify the risk of in-hospital mortality and contribute to better clinical management of these elderly patients.

Ultrasonographic assessment of organs other than the heart in patients with heart failure.

The number of patients with heart failure has been dramatically increasing in Japan in association with aging of the society. This phenomenon is referred to as a heart failure pandemic. The fundamental origin of heart failure is cardiac dysfunction. Echocardiography is widely used to assess cardiac function, as well as to diagnose heart diseases that cause cardiac dysfunction. However, the severity of heart failure is not necessarily correlated with that of cardiac dysfunction. This is partly explained by the fact that heart failure induces dysfunction of organs other than the heart through hemodynamic deterioration and neurohumoral changes. In addition, one of the characteristics of patients with heart failure, particularly elderly patients, is the presence of numerous comorbidities. Symptoms of heart failure are not specific, and assessment of cardiac function, particularly left ventricular diastolic function, has not been established. Thus, ultrasonographic assessment of organs other than the heart helps the diagnosis of heart failure, assessment of the severity of heart failure, and development of our understanding of the pathophysiology in each patient. This review summarizes current knowledge about the usefulness of ultrasonographic assessment of organs other than the heart in heart failure.

Uricosuric action of losartan via the inhibition of urate transporter 1 (URAT 1) in hypertensive patients.

BACKGROUND: The angiotensin receptor blocker losartan inhibited urate transporter 1 (URAT1) according to in vitro experiments. However, it is still unknown whether the inhibitory effect of losartan on URAT1 contributes to its uricosuric action in humans. METHODS: Thirty-two patients with hypertension and nine patients with idiopathic renal hypouricemia (five with and four without hypertension) were enrolled for this study. Hypertensive patients were prescribed oral losartan (50 mg/day, n = 16) or candesartan (8 mg/day, n = 16). Before and after 1-month treatment, the serum concentration of urate (Sur) and creatinine (Scr), and the clearance value of urate (Cur) and creatinine (Ccr) were determined. Clearance studies using the URAT1 inhibitor benzbromarone (100 mg/day) or losartan (50 mg/day) loading test were also performed in these patients. RESULTS: Blood pressure (BP) significantly decreased in the patients treated with either losartan or candesartan. Losartan significantly reduced Sur, which was associated with a concomitant increase in the Cur/Ccr ratio, whereas candesartan did not alter these parameters. In hypertensive patients with loss-of-function mutation of URAT1, losartan did not alter either Sur or Cur/Ccr, nor did benzbromarone. The lack of effect of URAT1 inhibitors on renal excretion of urate was independent of the renal function of hypouricemic patients. On the other hand, both losartan and benzbromarone increased Cur/Ccr ratio in hypertensive patients harboring the wild URAT1 gene, regardless of the presence of hypouricemia. CONCLUSIONS: These findings suggested that losartan inhibited URAT1 and thereby it lowered Sur levels in hypertensive patients.

Sex differences in surrogate decision-maker preferences for life-sustaining treatments of Japanese patients with heart failure.

AIMS: Patients with end-stage heart failure (HF) often require surrogate decision making for end-of-life care owing to a lack of decision-making capacity. However, the clinical characteristics of surrogate decision making for life-sustaining treatments in Japan remain to be investigated. METHODS AND RESULTS: Among 934 patients admitted to our hospital for HF from January 2004 to December 2015, we retrospectively reviewed the medical records of consecutive 106 patients who died in hospital (mean age 73 ± 13 years; male, 52.6%). During hospitalization, attending physicians conducted an average of 2.1 ± 1.4 end-of-life conversations with patients and/or their families. Only 4.7% of patients participated in the conversations and declared their preferences; surrogates made medical care decisions in 95.3% of cases. Most decisions by surrogates (98.1%) were made without the patient's advance directive. During initial end-of-life conversations, 49.4% of surrogates requested cardiopulmonary resuscitation (CPR). However, 72.0% of CPR preferences were changed to do not attempt resuscitation (DNAR) orders in the final conversation. Female surrogates were more likely to change the preference from CPR to DNAR than were male surrogates (47.1% vs. 25.0%, P = 0.023). CONCLUSIONS: Compared with male surrogates, female surrogates wavered more often in their decisions regarding life-sustaining treatments of Japanese patients with end-stage HF.

Age-related BM-MNC dysfunction hampers neovascularization.

Although ischemia-induced neovascularization is reportedly impaired with aging, the effect of aged-bone marrow mononuclear cells (BM-MNCs) on neovascularization has not been investigated. The neovascularization capacity of BM-MNCs obtained from 8-week-old mice (young) was compared to those obtained from 18-month-old mice (old), both in vivo and in vitro. Neovascularization in ischemic limbs was significantly impaired in old mice. Whereas transplantation of young BM-MNCs significantly improved blood perfusion, tissue capillary density, and vascular endothelial growth factor (VEGF) production in transplanted ischemic limbs, no such effects were observed with old BM-MNCs. Old BM-MNCs also showed a significant impairment of in vitro VEGF production and migratory capacity in response to VEGF. The number of Dil/lectin-positive cells was significantly lower in old mice, but there was no difference in the number of AC133(+)/CD34(+) and CD34(+)/VEGF-R2(+) positive cells between young and old BM-MNCs. Transplantation of young BM-MNCs improved neovascularization and VEGF production in the ischemic limbs of old recipients, with results that were similar to those obtained in young recipients. These results indicate that the neovascularization capacity of transplanted BM-MNCs is impaired with aging. However, aging does not hamper the revitalization of neovascularization in the murine host in response to transplantation of young BM-MNCs.