Covalent Stem Cell-Combining Injectable Materials with Enhanced Stemness and Controlled Differentiation In Vivo.

Biohybrid materials, which are defined as engineered functional materials combining living components with nonliving synthetic materials, are considered promising bioactive materials for applications in in vivo tissue engineering. However, the rational design of biohybrid materials applicable to in vivo tissue engineering faces major challenges associated with techniques for combining living cells with nonliving synthetic materials and cell sources. Here, we report injectable covalent stem cell-combing biohybrid materials prepared via a bio-orthogonal click cross-linking reaction of azide-modified adipose-derived stem cells (N3[+]ADSCs), one of the most promising cell sources utilized clinically, with alkyne-modified biocompatible alginate polymers. The mechanical properties of the covalent stem cell-combining biohybrid materials can be adapted to the mechanical properties of the surrounding environment in which they are transplanted by alternating the number of N3[+]ADSCs, the concentration of alkyne-modified alginate, and the number of alkyne groups. Importantly, ADSCs in the covalent biohybrid materials expressed a high level of CD-105, a marker for undifferentiated mesenchymal stem cells, in the body in the absence of differentiation signals, whereas very little CD-105 was expressed in the control physical cell-loading materials, demonstrating that this covalent stem cell-combining approach results in enhanced retention of the material's "stemness" and controlled differentiation in the body. We assessed the potential utility of the covalent stem cell-combining biohybrid materials for in vivo tissue engineering using a murine severe skeletal muscle defect-healing model. Importantly, all of the tissues regenerated by the covalent biohybrid material treatment expressed MYH3, a myogenic marker protein, whereas no expression of MYH3 was detected in the tissues reconstructed by treatment with control physical stem cell-loading materials and Matrigel, indicating that this covalent stem cell-combining approach results in controlled differentiation in the body. Our data demonstrate the potential utility of covalent stem cell-combining biohybrid materials with host tissue-integrative and controlled differentiation capabilities available for in vivo tissue engineering.

Liddle's-like syndrome associated with nephrotic syndrome secondary to membranous nephropathy: the first case report.

BACKGROUND: Liddle's syndrome is a rare monogenic form of hypertension caused by truncating or missense mutations in the C termini of the epithelial sodium channel (ENaC) ß or γ subunits. Patients with this syndrome present with early onset of hypertension, hypokalemia, metabolic alkalosis, hyporeninemia and hypoaldosteronism, and a potassium-sparing diuretics (triamterene or amiloride) can drastically improves the disease condition. Although elderly patients having these characteristics were considered to have Liddle's syndrome or Liddle's-like syndrome, no previous report has indicated that Liddle's-like syndrome could be caused by nephrotic syndrome of primary glomerular disease, which is characterized by urinary excretion of > 3 g of protein/day plus edema and hypoalbuminemia, or has explained how the activity function of ENaC could be affected in the setting of high proteinuria. CASE PRESENTATION: A 65-year-old Japanese man presented with nephrotic syndrome. He had no remarkable family history, but had a medical history of hypertension and hyperlipidemia. On admission, hypertension, spironolactone-resistant hypokalemia (2.43 mEq/l), hyporeninemic hypoaldosteronism, and metabolic alkalosis, which suggested Liddle's syndrome, were observed. Treatment with triamterene together with a steroid for nephrotic syndrome resulted in rapid and remarkable effective on improvements of hypertension, hypokalemia, and edema of the lower extremities. Renal biopsy revealed membranous nephropathy (MN) as the cause of nephrotic syndrome, and advanced gastric cancer was identified on screening examination for cancers that could be associated with the development of MN. After total gastrectomy, triamterene was not required and proteinuria decreased. A mutation in the ß or γ subunits of the ENaC gene was not identified. CONCLUSION: We reported for the first time a case of Liddle's-like syndrome associated with nephrotic syndrome secondary to MN. Aberrant activation of ENaC was suggested transient during the period of high proteinuria, and the activation was reversible with a decrease in proteinuria.

Standardized Prevalence Ratios for Atrial Fibrillation in Adult Dialysis Patients in Japan.

BACKGROUND: While it is assumed that dialysis patients in Japan have a higher prevalence of atrial fibrillation (AF) than the general population, the magnitude of this difference is not known. METHODS: Standardized prevalence ratios (SPRs) for AF in dialysis patients (n = 1510) were calculated compared to data from the general population (n = 26 454) living in the same area. RESULTS: The prevalences of AF were 3.8% and 1.6% in dialysis patients and the general population, respectively. In male subjects, these respective values were 4.9% and 3.3%, and in female subjects they were 1.6% and 0.6%. The SPRs for AF were 2.53 (95% confidence interval [CI], 1.88-3.19) in all dialysis patients, 1.80 (95% CI, 1.30-2.29) in male dialysis patients, and 2.13 (95% CI, 0.66-3.61) in female dialysis patients. CONCLUSIONS: The prevalence of AF in dialysis patients was twice that in the population-based controls. Since AF strongly contributes to a higher risk of cardiovascular mortality and morbidity in the general population, further longitudinal studies should be conducted regarding the risk of several outcomes attributable to AF among Japanese dialysis patients.

Associations of marital status with mortality from all causes and mortality from cardiovascular disease in Japanese haemodialysis patients.

BACKGROUND: Marital status is an important social factor associated with increased mortality from cardiovascular disease (CVD) and all causes. However, there has been no study on the association of marital status with mortality in haemodialysis patients. METHODS: We analysed data from a 5-year prospective cohort study of 1064 Japanese haemodialysis patients aged 30 years or older. Marital status was classified into three groups: married, single and divorced/widowed. Cox's regression was used to estimate multivariate hazard ratios (HRs) [95% confidence intervals (CIs)] for all-cause mortality and CVD mortality according to marital status after adjusting for age, sex, duration of haemodialysis, cause of renal failure, body mass index, systolic blood pressure, total cholesterol, high density lipoprotein-cholesterol, albumin, high-sensitivity C-reactive protein, co-morbid conditions, smoking, alcohol consumption, education levels and job status. RESULTS: Single patients had higher risks than married patients for mortality from all causes (HR = 1.51, 95% CI: 1.06-2.16) and mortality from CVD (HR = 1.68, 95% CI: 1.03-2.76), and divorced/widowed patients had a higher risk than married patients for mortality from CVD (HR = 1.73, 95% CI: 1.15-2.60). After stratification by age, single patients aged 30-59 years had significantly higher risks for all-cause mortality and CVD mortality. CONCLUSIONS: The findings suggest that single status is a significant predictor for all-cause mortality and CVD mortality and that divorced/widowed status is a significant predictor for CVD mortality in haemodialysis patients.

Seropositivity for anti-HCV core antigen is independently associated with increased all-cause, cardiovascular, and liver disease-related mortality in hemodialysis patients.

BACKGROUND: It is not known whether chronic or past hepatitis C virus (HCV) infection contributes to the high mortality rate in hemodialysis patients. METHODS: This prospective study of 1077 adult hemodialysis patients without hepatitis B virus infection used Poisson regression analysis to estimate crude and sex- and age-adjusted rates (per 1000 patient-years) of all-cause, cardiovascular, infectious disease-related and liver disease-related mortality in patients negative for HCV antibody (group A), patients positive for HCV antibody and negative for anti-HCV core antigen (group B), and patients positive for anti-HCV core antigen (group C). The relative risks (RRs) for each cause of death in group B vs group C as compared with those in group A were also estimated by Poisson regression analysis after multivariate adjustment. RESULTS: A total of 407 patients died during the 5-year observation period. The sex- and age-adjusted mortality rate was 71.9 in group A, 80.4 in group B, and 156 in group C. The RRs (95% CI) for death in group B vs group C were 1.23 (0.72 to 2.12) vs 1.60 (1.13 to 2.28) for all-cause death, 0.75 (0.28 to 2.02) vs 1.64 (0.98 to 2.73) for cardiovascular death, 1.64 (0.65 to 4.15) vs 1.58 (0.81 to 3.07) for infectious disease-related death, and 15.3 (1.26 to 186) vs 28.8 (3.75 to 221) for liver disease-related death, respectively. CONCLUSIONS: Anti-HCV core antigen seropositivity independently contributes to elevated risks of all-cause and cause-specific death. Chronic HCV infection, but not past HCV infection, is a risk for death among hemodialysis patients.

The blood compatibilities of blood purification membranes and other materials developed in Japan.

The biocompatibilities in blood purification therapy are defined as "a concept to stipulate safety of blood purification therapy by an index based on interaction in the body arising from blood purification therapy itself." The biocompatibilities are associated with not only materials to be used but also many factors such as sterilization method and eluted substance. It is often evaluated based on impacts on cellular pathways and on humoral pathways. Since the biocompatibilities of blood purification therapy in particular hemodialysis are not just a prognostic factor for dialysis patients but a contributory factor for long-term complications, it should be considered with adequate attention. It is important that blood purification therapy should be performed by consistently evaluating not only risks associated with these biocompatibilities but also the other advantages obtained from treatments. In this paper, the biocompatibilities of membrane and adsorption material based on Japanese original which are used for blood purification therapy are described.

Serum selenium levels are inversely associated with death risk among hemodialysis patients.

BACKGROUND: Previous studies have indicated that serum selenium levels are decreased in hemodialysis patients. Selenium deficiency may contribute to an increased risk for death among hemodialysis patients. METHODS: A population-based prospective cohort study in adult hemodialysis patients was conducted. A total of 1041 patients were enrolled. Patients were divided into quartile groups according to serum selenium levels. Mortality rates between the groups were compared by the log-rank test. Associations between serum selenium levels and cause-specific mortality risks in hemodialysis patients were examined by Cox's regression model. RESULTS: A total of 382 patients died during the 5-year follow-up period (median follow-up period, 4.9 years). Crude mortality rates in quartile groups according to serum selenium levels were 134.5, 99.9, 85.9 and 55.2 (per 1000 patient-years), respectively. The lowest quartile group had significantly higher mortality rates from all-cause and infectious disease-related death than the rates in the other three groups (P < 0.001, by log-rank test). Mortality rates from cardiovascular and malignant disease-related death were similar between the groups. A strong inverse relationship between selenium levels and infectious disease-related death was observed even after multivariate adjustment (trend P = 0.024). CONCLUSIONS: Serum selenium levels were inversely associated with death risk, especially death risk due to infectious disease, among hemodialysis patients. Decreased serum selenium level may contribute to immunity dysfunction and may increase the risk of death from infectious disease in hemodialysis patients.

Serum selenium levels in hemodialysis patients are significantly lower than those in healthy controls.

Serum selenium levels have been thought to be decreased in hemodialysis patients; however, results of previous studies have been inconsistent. Population-based hemodialysis patients (n = 1,041) and randomly recruited healthy controls (n = 384) were enrolled. Serum selenium levels were determined by inductively coupled plasma mass spectrometry and compared in hemodialysis patients and controls using analysis of covariance after adjustment for confounding factors with p < 0.1 as the result of the multiple regression analysis. Age, serum albumin levels, hsCRP levels, LDLC levels, HDLC levels, regular drinking habit and hemodialysis treatment were significantly associated with serum selenium levels in multiple regression analysis. Multivariate-adjusted means (95% CIs) of serum selenium levels were 103 µg/l (101-105) in hemodialysis patients and 117 µg/l (114-121) in controls. Selenium levels in hemodialysis patients were decreased. Whether decreased serum selenium levels contribute to increased risks for morbidity and mortality in hemodialysis patients should be examined.

Standardized prevalence ratios for chronic hepatitis C virus infection among adult Japanese hemodialysis patients.

BACKGROUND: Many studies have estimated the prevalence of anti-hepatitis C virus (HCV) antibody among hemodialysis (HD) patients; however, the prevalence of HCV core antigen-which indicates the presence of chronic HCV infection-is not known. METHODS: Standardized prevalence ratios (SPRs) for anti-HCV antibody and HCV core antigen among HD patients (n = 1214) were calculated on the basis of data from the general population (n = 22 472) living in the same area. RESULTS: The prevalences of anti-HCV antibody and HCV core antigen were 12.5% and 7.8%, respectively, in male hemodialysis patients, and 8.5% and 4.1% in female hemodialysis patients. The SPRs (95% confidence interval) for anti-HCV antibody and HCV core antigen were 8.39 (6.72-10.1) and 12.9 (9.66-16.1), respectively, in males, and 5.42 (3.67-7.17) and 8.77 (4.72-12.8) in females. CONCLUSIONS: The prevalences of chronic HCV infection among male and female HD patients were 13-fold and 9-fold, respectively, those of the population-based controls. Further studies should therefore be conducted to determine the extent of chronic HCV infection among HD patients in other populations and to determine whether chronic HCV infection contributes to increased mortality in HD patients.

Vitamin D receptor gene polymorphisms are associated with increased risk and progression of renal cell carcinoma in a Japanese population.

AIM: Biological and epidemiologic data suggest that 1 alpha, 25 dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) levels may influence development of renal cell carcinoma. The vitamin D receptor (VDR) is a crucial mediator for the cellular effects of 1,25(OH)(2)D(3) and additionally interacts with other cell signaling pathways that influence cancer progression. VDR gene polymorphisms may play an important role in risk of incidence for various malignant tumors. This study investigated whether VDR gene polymorphisms were associated with increased risk and prognosis of renal cell carcinoma (RCC) in a Japanese population. METHODS: To analyze risk of RCC depending on VDR polymorphism, a case-control association study was performed. The VDR gene polymorphisms at three locations, BsmI, ApaI and TaqI, were genotyped in 135 RCC patients and 150 controls in a Japanese population. Logistic regression models were used to assess the genetic effects on prognosis. RESULTS: Significant differences in the ApaI genotype were observed between RCC patients and controls (chi(2) = 6.90, P = 0.032). No statistical significant difference was found in the BsmI and TaqI polymorphisms. The frequency of the AA genotype in the ApaI polymorphism was significantly higher in the RCC patients than in the controls (odds ratio, 2.59; 95% confidence intervals, 1.21-5.55; P = 0.012). Multivariate regression analysis showed that the AA genotype was an independent prognostic factor for cause-specific survival (relative risk 3.3; P = 0.038). CONCLUSION: The AA genotype at the ApaI site of the VDR gene may be a risk of incidence and poor prognosis factor for RCC in the Japanese population. Additional studies with a large sample size and investigation of the functional significance of the ApaI polymorphism in RCC cells are warranted.

Mortality risk attributable to atrial fibrillation in middle-aged and elderly people in the Japanese general population: nineteen-year follow-up in NIPPON DATA80.

BACKGROUND: The extent to which atrial fibrillation (AF) contributes to mortality in the Japanese general population has not been clarified. METHODS AND RESULTS: A randomly sampled general population from all over Japan (4,154 men, 5,329 women; age > or =30 years) was enrolled. Single electrocardiogram recordings were taken in the baseline survey. Stroke death, cardiovascular deaths and all-cause deaths during the subsequent 19 years were analyzed by the presence of AF at baseline. Cox's regression analysis was carried out to estimate the hazard ratios (HRs) of each cause of death attributable to AF after adjusting for other risk factors. Prevalence of AF was 0.64% in the study. The observed person-years were 162,980 among persons without AF and 699 among persons with AF. There were 1,919 deaths. Multivariate adjusted HRs for stroke death, cardiovascular death and all-cause death were 2.69, 2.76 and 1.88, respectively (p<0.05). These HRs were 14.7, 9.63 and 4.00 among persons aged 64 years or younger (p<0.05). CONCLUSION: AF affects stroke mortality, cardiovascular mortality and all-cause mortality in the Japanese general population. Careful attention should be paid to persons with AF in order to prevent future cardiovascular events.

Eight year experience with Studer ileal neobladder.

BACKGROUND: We reviewed our experience with orthotopic continent urinary reconstruction after radical cystectomy to assess the feasibility of Studer ileal neobladder for patients who are relatively advanced in age. METHODS: Between June 1997 and January 2005, 31 consecutive male patients (mean age: 64 years) underwent lower urinary tract reconstruction after radical cystoprostatectomy. Perioperative and late complications, functional outcome of the neobladder, urinary continence, upper urinary tract status and renal function with the metabolic balance were evaluated in all patients. RESULTS: There was no perioperative death, and perioperative and late complication rates were 22.8% and 3.3%, respectively. All 31 patients were able to void urine. Although the mean maximal functional capacity of the neobladder was 122 ml at 1 month after surgery, the mean capacities were increased to 247 ml at 6 months and 321 ml at 1 year after the operation. Urodynamic results at 3 years showed unchanged characteristics as to micturition pattern and volume of residual urine and neobladder pressure remained low. Of 31 patients, 29 (93.5%) showed excellent or good continent status during the daytime and 9 (29%) were completely dry at night in 6 months after surgery. Even at 3 years after the operation, only 1 patient out of 21 evaluated required single pad during nighttime. In a subgroup of five patients (24%) older than 70 years, the status of continence was satisfactory at 3 years after the reconstruction, and only one patient required a pad during the night at that point. Renal function levels and metabolic status were comparable before surgery and 3 years after surgery. Moreover, pyelography revealed normal condition of the upper urinary tract 1 month postoperatively in almost all cases. CONCLUSIONS: These data provide evidence that Studer ileal neobladder is a satisfactory surgical technique for selected patients at our institute. Even for patients older than 70 years, this urinary diversion procedure is safe in terms of morbidity and efficacious as indicated by functional outcome.

Cardiovascular risk factors in hemodialysis patients: results from baseline data of kaleidoscopic approaches to patients with end-stage renal disease study.

BACKGROUND: The prevalence of cardiovascular risk factors and the prevalence of comorbidities in adult hemodialysis patients in Japan are not fully understood. METHODS: In "Kaleidoscopic Approaches to Patients with End-stage Renal Disease Study" (The KAREN Study, 2003), trained research stuff examined 1,214 adult hemodialysis patients (mean age, 61.2 years; 779 males and 435 females) of 1,506 patients in northern areas of Iwate Prefecture. Cardiovascular risk factors and the prevalence of comorbidities in hemodialysis patients were compared with those in the general population using direct age-adjustment methodology and standardized morbidity ratios (SMRs). RESULTS: In hemodialysis patients, common causes of end-stage renal disease were chronic glomerulonephritis (29.8%), diabetic nephropathy (24.5%), and other diseases. Prevalence and SMR of myocardial infarction were 5% and 9.6, respectively, and those of stroke were 13% and 5.7. The prevalences of hypertension and diabetes mellitus were 87% and 29%, respectively. Mean systolic blood pressure and mean diastolic blood pressure were 155 mmHg and 85 mmHg, respectively. Mean levels of total serum cholesterol, high-density lipoprotein cholesterol, and albumin in patients with end-stage renal disease were lower than those of the general population (160.6 vs. 203.3 mg/dL, 48.5 vs. 59.7 mg/dL, and 3.7 vs. 4.4 g/dL, respectively). Mean levels of C-reactive protein were higher than those of the general population (3.80 vs. 1.16 mg/L). CONCLUSION: Hemodialysis patients have a high prevalence of cardiovascular risk factors and comorbidities. Levels of nutrition-related markers were lower, and C-reactive protein levels were higher, in hemodialysis patients than in the general population.

CRP levels are elevated in smokers but unrelated to the number of cigarettes and are decreased by long-term smoking cessation in male smokers.

BACKGROUND: It is not clear whether there is a dose-response relationship between the number of cigarettes smoked per day and CRP level and whether there is a relationship between the length of smoking cessation and CRP level. METHODS: Geometric mean levels of CRP were compared in smoking status groups for 1926 men aged 40 to 69 years using analysis of covariance. RESULTS: After adjusting for several confounding factors, geometric mean levels of CRP (mg/L) were significantly different among the three smoking status groups (0.41 in non-smokers, 0.57 in current smokers, 0.48 in past smokers, P < 0.05). A linear trend was not found in the relationship between CRP level and number of cigarettes smoked per day. The mean CRP level in the long cessation (> or =5 years) group was significantly lower than that in the short cessation (<5 years) group (0.45 vs. 0.58, P < 0.05) and similar to that in the non-smokers group (0.45 vs. 0.41, NS). CONCLUSIONS: CRP levels in current smokers are elevated but unrelated to the number of cigarettes smoked per day. In past smokers, long-term smoking cessation may contribute to the reduction in risk of development of cardiovascular diseases through inflammatory mechanisms.

[Central and peripheral nerve disorder induced by interferon alpha therapy for renal cell carcinoma].

Interferon alpha (IFN-alpha) therapy was conducted for a male patient aged 70 years old, who underwent a two-stage radical nephrectomy for bilateral renal cell carcinoma with multiple pulmonary metastasis. He was hospitalized due to leg weakness and disorientation 45 days after this treatment was started. We discontinued INF-alpha therapy immediately after neurologists indicated the disorder of the central and the peripheral nervous systems induced by the administration of this cytokine. Steroid pulse therapy was effective to resolve the patient's neurological symptoms. To our knowledge, this is the first case of the side effects on both central and peripheral nervous systems by IFN-alpha therapy for renal cell carcinoma.