Accelerated Vascular Aging in Chronic Kidney Disease: The Potential for Novel Therapies.

The pathophysiology of vascular disease is linked to accelerated biological aging and a combination of genetic, lifestyle, biological, and environmental risk factors. Within the scenario of uncontrolled artery wall aging processes, CKD (chronic kidney disease) stands out as a valid model for detailed structural, functional, and molecular studies of this process. The cardiorenal syndrome relates to the detrimental bidirectional interplay between the kidney and the cardiovascular system. In addition to established risk factors, this group of patients is subjected to a plethora of other emerging vascular risk factors, such as inflammation, oxidative stress, mitochondrial dysfunction, vitamin K deficiency, cellular senescence, somatic mutations, epigenetic modifications, and increased apoptosis. A better understanding of the molecular mechanisms through which the uremic milieu triggers and maintains early vascular aging processes, has provided important new clues on inflammatory pathways and emerging risk factors alike, and to the altered behavior of cells in the arterial wall. Advances in the understanding of the biology of uremic early vascular aging opens avenues to novel pharmacological and nutritional therapeutic interventions. Such strategies hold promise to improve future prevention and treatment of early vascular aging not only in CKD but also in the elderly general population.

Fermented food: Should patients with cardiometabolic diseases go back to an early neolithic diet?

Fermentation has been used since the Early Neolithic period to preserve foods. It has inherent organoleptic and nutritive properties that bestow health benefits, including reducing inflammation and oxidative stress, supporting the growth of salutogenic microbiota, enhancing intestinal mucosal protection and promoting beneficial immunometabolic health effects. The fermentation of food with specific microbiota increases the production salutogenic bioactive compounds that can activate Nrf2 mediated cytoprotective responses and mitigate the effects of the 'diseasome of aging' and its associated inflammageing, which presents as a prominent feature of obesity, type-2 diabetes, cardiovascular and chronic kidney disease. This review discusses the importance of fermented food in improving health span, with special reference to cardiometabolic diseases.

Short Report: Choline plasma levels are related to Nrf2 transcriptional expression in chronic kidney disease?

BACKGROUND & AIMS: Choline is an amine osmolyte with antioxidant potential. A limited number of studies have implied that choline modulates the nuclear factor-erythroid 2-related factor 2 (Nrf 2) pathway, a major cytoprotectant system. However, there are no data regarding such an interaction in patients with chronic kidney disease (CKD). This cross-sectional pilot study therefore aimed to evaluate the possible relationship between choline plasma levels and transcriptional expression of Nrf2 in patients with CKD on hemodialysis (HD). METHODS: This study was performed in 24 HD patients [54 ± 10 years, 14 men, BMI 26.4 ± 4.5 kg/m2]. Choline plasma levels were measured by LC-MS/MS. Nrf2 mRNA expression was measured by real-time quantitative polymerase chain reaction (rt-PCR) in isolated peripheral blood mononuclear cells (PBMC). RESULTS: We used Pearson's correlation (rho) to determine the correlations with Nrf 2 expression and observed a positive correlation between choline plasma levels and Nrf2 (rho = 0.56, P = 0.004). CONCLUSIONS: This finding suggests that choline may play a role in Nrf2 expression in CKD.

The role of the microbiota in sedentary lifestyle disorders and ageing: lessons from the animal kingdom.

A paradox of so-called developed countries is that, as the major historical causes of human mortality are eliminated or mitigated by medical progress, lifestyle-related diseases have become major killers. Furthermore, as lifespan is extended by the combined effects of modern medicine, health span is struggling to keep apace because of the burden of noncommunicable diseases linked to diet and sedentary lifestyle. The gut microbiome is now recognized as a plastic environmental risk factor for many of these diseases, the microbiome being defined as the complex community of co-evolved commensal microbes that breaks down components of a complex diet, modulates innate immunity, and produces signalling molecules and metabolites that can impact on diverse regulatory systems in mammals. Aspects of the so-called 'Western' lifestyle linked to disease risk such as energy dense diet and antibiotic treatment are known to affect the composition and function of the microbiome. Here, we review the detailed mechanisms whereby the gut microbiome may modulate risk of diseases linked to sedentary lifestyle and ageing-related health loss. We focus on the comparative value of natural animal models such as hibernation for studying metabolic regulation and the challenge of extrapolating from animal models to processes that occur in human ageing.

Has the Expansion in Extended Criteria Deceased Donors Led to a Different Type of Delayed Graft Function and Poorer Outcomes?

OBJECTIVES: There has been considerable change in the practice of deceased kidney transplantation in the past 15 years, with more extreme phenotypes implanted. The aim of this study was to determine whether increased use of expanded criteria donors (extended criteria donors and donors after circulatory death) affected clinical outcomes, including the incidence and pattern of delayed graft function. METHODS AND MATERIALS: A retrospective analysis of 1359 renal transplants was performed over 15 years. The first 10 years of data (group 1) were compared with the subsequent 5 years (group 2). Outcomes were analyzed at 6 months and 12 months in addition to serum creatinine and patterns of delayed graft function (posttransplant times: on hemodialysis, to peak creatinine, for creatinine to half, and for creatinine to fall within 10% of baseline). RESULTS: There was a significant increase in the percentage of expanded criteria donor allografts used in group 2 with a significant increase in the incidence of delayed graft function. Despite this, serum creatinine and the incidence of biopsy-proven acute rejection had both improved in group 2. Group 2 expanded criteria donor kidneys had a significantly lower incidence of type 1 delayed graft function and a significantly higher incidence of types 3 and 4 delayed graft function. Time for creatinine to half in both groups was the best predictor of a serum creatinine <180 µmol/L at 1 year. CONCLUSION: The increased use of expanded criteria donor kidneys has led to a higher incidence of delayed graft function, but the pattern has shown that the requirement for hemodialysis has significantly reduced.

Patients with gout have short telomeres compared with healthy participants: association of telomere length with flare frequency and cardiovascular disease in gout.

AIM AND BACKGROUND: Chronic inflammation associates with increased senescence, which is a strong predictor for cardiovascular disease. We hypothesised that inflammation accelerates senescence and thereby enhances the risk of cardiovascular disease in gout. METHODS: We assessed replicative senescence by quantifying telomere length (TL) in a discovery cohort of 145 Dutch patients with gout and 273 healthy individuals and validated our results in 474 patients with gout and 293 healthy participants from New Zealand. Subsequently, we investigated the effect of cardiovascular disease on TL of all participants. Also, we measured TL of CD4+ and CD8+ T lymphocytes, B lymphocytes, monocytes, natural killer cells and plasmacytoid dendritic cells. Additionally, we assessed the potential temporal difference in TL and telomerase activity. RESULTS: TL in PBMCs of healthy donors decreased over time, reflecting normal ageing. Patients with gout demonstrated shorter telomeres (p=0.001, R2=0.01873). In fact, the extent of telomere erosion in patients with gout was higher at any age compared with healthy counterparts at any age (p<0.0001, R2=0.02847). Patients with gout with cardiovascular disease had the shortest telomeres and TL was an independent risk factor for cardiovascular disease in patients with gout (p=0.001). TL was inversely associated with the number of gouty flares (p=0.005). CONCLUSIONS: Patients with gout have shorter telomeres than healthy participants, reflecting increased cellular senescence. Telomere shortening was associated with the number of flares and with cardiovascular disease in people with gout.

The in situ local immune response, tumour senescence and proliferation in colorectal cancer.

BACKGROUND: Immune cell infiltrates are important determinants of colorectal cancer (CRC) outcome. Their presence may be driven by tumour or host-specific factors. From previous studies in mice, senescence, a state of cell cycle arrest, may moderate tumour progression through upregulation of antitumour immune responses. The relationships between senescence and immune infiltrates have not previously been studied in humans. We explore whether a marker of senescence (p16(ink4a)) in combination with low level expression of a proliferation marker (ki-67) relate to T cell infiltrates in CRC, and whether p16(ink4a), Ki-67 and immune infiltrates have similar prognostic value. METHODS: Immunostaining of p16(inka) and Ki-67 was performed within a CRC tissue microarray. Nuclear p16(inka) and Ki-67 were categorised as high/low. T-cell markers, CD3, CD45RO, CD8 and FOXP3 were scored separately as high/low grade in three areas of the tumour: the invasive margin (IM), tumour stroma and cancer cell nests (CCNs). results: Two hundred and thirty stage I-III cancers were studied. High nuclear p16(ink4a) was expressed in 63% and high proliferation (Ki-67 >15%) in 61%. p16(ink4a) expression was associated with reduced CD45RO+ cells at the IM (P<0.05) and within the stroma (P<0.05) and reduced CD8+ cells at the IM (P<0.01). A low Ki-67 proliferative index was associated with reduced density of CD3+ cells in CCNs (P<0.01), reduced CD45RO+ cells at the IM (P<0.05) and within the CCNs (P<0.001), reduced FOXP3+ cells at the IM (P<0.001), within the stroma (P=0.001) and within CCNs (P<0.001) and reduced CD8+ cells at the IM (P<0.05) and within the CCNs (P<0.05). Tumours with both a low proliferative index and expression of p16(ink4a) demonstrated similar consistent relationships with reduced densities of T-cell infiltrates. On multivariate analysis, TNM stage (P<0.001), low CD3 cells at the IM (P=0.014), low CD8 cells at the IM (P=0.037), low proliferation (Ki-67; P=0.013) and low senescence (p16(ink4a); P=0.002) were independently associated with poorer cancer survival. CONCLUSION: Senescence, proliferation and immune cell infiltrates are independent prognostic factors in CRC. Although related to survival, p16(ink4a)-associated senescence is not associated with an upregulation of antitumour T-cell responses.

Below-target postoperative arterial blood pressure but not central venous pressure is associated with delayed graft function.

Delayed graft function (DGF) is a major issue in kidney transplantation and is associated with reduced graft and patient survival. The condition results from the summative effects of multiple injurious processes associated with transplantation with many underlying factors being nonmodifiable. Reducing cold ischemic time and machine perfusion have decreased the DGF incidence but peri-/postoperative injury resulting from suboptimal perfusion may also be critical to the development of DGF. We investigated the effect of perfusion parameters and other key variables on the incidence of DGF in 149 consecutive renal transplants. The occurrence of any recorded subtarget (70 mm Hg) mean arterial pressure (MAP) was significantly associated with DGF (perioperative P = .005; postoperative P = .002) while the occurrence of a subtarget (8 cm H(2)O) central venous pressure (CVP) among other variables was not. Routine continuous blood pressure monitoring is rare postoperatively and is shown to be more accurate than CVP in assessing renal perfusion and guiding management in the postoperative period.

Sirtuins, bioageing, and cancer.

The Sirtuins are a family of orthologues of yeast Sir2 found in a wide range of organisms from bacteria to man. They display a high degree of conservation between species, in both sequence and function, indicative of their key biochemical roles. Sirtuins are heavily implicated in cell cycle, cell division, transcription regulation, and metabolism, which places the various family members at critical junctures in cellular metabolism. Typically, Sirtuins have been implicated in the preservation of genomic stability and in the prolongation of lifespan though many of their target interactions remain unknown. Sirtuins play key roles in tumourigenesis, as some have tumour-suppressor functions and others influence tumours through their control of the metabolic state of the cell. Their links to ageing have also highlighted involvement in various age-related and degenerative diseases. Here, we discuss the current understanding of the role of Sirtuins in age-related diseases while taking a closer look at their roles and functions in maintaining genomic stability and their influence on telomerase and telomere function.

Breast cancer patients' clinical outcome measures are associated with Src kinase family member expression.

BACKGROUND: This study determined mRNA expression levels for Src kinase family (SFK) members in breast tissue specimens and assessed protein expression levels of prominent SFK members in invasive breast cancer to establish associations with clinical outcome. Ki67 was investigated to determine association between SFK members and proliferation. METHODS: The mRNA expression levels were assessed for eight SFK members by quantitative real-time PCR. Immunohistochemistry was performed for c-Src, Lyn, Lck and Ki67. RESULTS: mRNA expression was quantified in all tissue samples. SRC and LYN were the most highly expressed in malignant tissue. LCK was more highly expressed in oestrogen receptor (ER)-negative, compared with ER-positive tumours. High cytoplasmic Src kinase protein expression was significantly associated with decreased disease-specific survival. Lyn was not associated with survival at any cellular location. High membrane Lck expression was significantly associated with improved survival. Ki67 expression correlated with tumour grade and nuclear c-Src, but was not associated with survival. CONCLUSIONS: All eight SFK members were expressed in different breast tissues. Src kinase was highest expressed in breast cancer and had a negative impact on disease-specific survival. Membrane expression of Lck was associated with improved clinical outcome. High expression of Src kinase correlated with high proliferation.

Association of increased telomere lengths in limited scleroderma, with a lack of age-related telomere erosion.

OBJECTIVES: Telomere erosion, a feature of biological ageing, is implicated in a wide range of diseases. Its impact on autoimmune diseases remains unclear although autoantibodies against many telomere nucleoprotein components are prevalent in these diseases. We aimed to assess if telomere biology was abnormal in a cohort of patients with limited cutaneous systemic sclerosis (lcSSc). METHODS: Telomere lengths in peripheral blood leucocytes (PBL) were determined using Southern blotting methods in a cohort of lcSSc subjects (n=43; age range 37-80 years) and a control population (n=107; age range 21-65 years). RESULTS: Telomere lengths in lcSSc subjects were longer than controls (p<0.001), did not show age-related telomere erosion and differed significantly from age-matched controls only after 50 years of age (p<0.001). CONCLUSIONS: This is the first report of maintenance of telomere lengths in an autoimmune disease state. These data indicate aberrant telomere biology and irregular biological ageing from the fifth decade of life. These findings provide insight into compromised DNA damage repair in lcSSc. Whether these observations indicate a causal or consequential relationship requires further investigation. This in turn, may provide potential novel targets for therapeutic intervention.

Telomere attrition is associated with inflammation, low fetuin-A levels and high mortality in prevalent haemodialysis patients.

INTRODUCTION: Chronic kidney disease (CKD) predisposes to a 10- to 20-fold increased cardiovascular risk. Patients undergo accelerated atherogenesis and vascular ageing. We investigated whether telomere attrition, a marker of cell senescence, contributes to this increased mortality risk. METHODS: This is a cross-sectional study in prevalent haemodialysis patients [n = 175; 98 Males; median (range) age: 66 (23-86) years]. Biochemical markers of oxidative stress and inflammatory status were measured in relation to the patient's leucocyte telomere length. Overall mortality was assessed after a median of 31 (range 2-42) months. RESULTS: Telomere length was shorter in CKD men, despite women being older (average +/- SD 6.41 +/- 1.23 vs. 6.96 +/- 1.48 kb, P = 0.002). Telomere length was associated with age (rho = -0.18, P = 0.01), fetuin-A (rho = 0.26, P = 0.0004), high-sensitivity C-reactive protein (rho = -0.21, P = 0.005) and IL-6 (rho = -0.17, P = 0.02). In a multivariate logistic regression (pseudo r(2) = 0.14), telomere length was associated with age >65 years (odds ratio: 2.11; 95% CI: 1.10, 4.06), sex (2.01; 1.05, 3.86), fetuin-A (1.85; 0.97, 3.50) and white blood cell count (2.04; 1.02, 4.09). Receiver operating characteristic curves identified a telomere length < 6.28 kb as a fair predictor of mortality. Finally, reduced telomere length was associated with increased mortality, independently of age, gender and inflammation (likelihood ratio 41.6, P < 0.0001), but dependently on fetuin-A levels. CONCLUSION: Age and male gender seem to be important contributors to reduced telomere length in CKD patients, possibly via persistent inflammation. Reduced telomere length also contributes to the mortality risk of these patients through pathways that could involve circulating levels of fetuin-A.

Breath ethane peaks during a single haemodialysis session and is associated with time on dialysis.

There is growing evidence that oxidative stress is increased in haemodialysis patients and that dialysis per se is a contributory factor. The elevated oxidant stress, a result of increased production of reactive oxidant species (ROS), may be due to increased pro-inflammatory activity and reduced antioxidant mechanisms. ROS are transitory molecules and therefore surrogate markers of oxidant damage are required. Identification of potential causes of oxidative damage such as dialyser membranes or dialysate has been proposed and therefore assessment of oxidative damage during a single dialysis session would be of interest. We have used breath ethane, a widely accepted marker of oxidative stress, to investigate the cause and extent of the resulting oxidative damage during single dialysis sessions. Our study involved assessment of breath ethane levels during haemodialysis in an end-stage renal failure haemodialysis population (n = 24). Breath samples were collected using discrete sampling techniques and were subsequently analysed using laser spectroscopy. Each patient adopted the role of longitudinal control in this study and his or her breath ethane level was monitored regularly during the dialysis session. Significant breath ethane elevation was observed at the beginning (within the first 10 min) of each dialysis session. This paper provides an in-depth statistical analysis and clinical discussion of the recent findings. A regression analysis of the collected breath ethane data showed a trend towards increased ethane levels for patients on dialysis for a shorter duration of time (r = 0.656, R-Sq = 43.3%, p = 0.001). Multiple linear regression was undertaken to further assess these associations and revealed that peak ethane levels were significantly and independently associated with time period on dialysis (p < 0.000), vascular access (p = 0.013) and male sex (p = 0.005). However, whilst diabetes status had demonstrated a correlation with peak ethane levels (0.525, p = 0.008) this was not independent of vascular access status. This multivariate linear model was significantly associated with Ln peak ethane levels (S = 0.744, R-Sq = 80.8%). The observed rapid rise in oxidative stress during the first few minutes after commencement of dialysis gives new insight into the dynamics of the oxidative damage resulting from dialysis treatment.

Altered sirtuin expression is associated with node-positive breast cancer.

Sirtuins are genes implicated in cellular and organismal ageing. Consequently, they are speculated to be involved in diseases of ageing including cancer. Various cancers with widely differing prognosis have been shown to have differing and characteristic expression of these genes; however, the relationship between sirtuin expression and cancer progression is unclear. In order to correlate cancer progression and sirtuin expression, we have assessed sirtuin expression as a function of primary cell ageing and compared sirtuin expression in normal, 'nonmalignant' breast biopsies to breast cancer biopsies using real-time polymerase chain reaction (PCR). Levels of SIRT7 expression were significantly increased in breast cancer (P<0.0001). Increased levels of SIRT3 and SIRT7 transcription were also associated with node-positive breast cancer (P<0.05 and P<0.0001, respectively). This study has demonstrated differential sirtuin expression between nonmalignant and malignant breast tissue, with consequent diagnostic and therapeutic implications.

The use of telomere biology to identify and develop superior nitrone based anti-oxidants.

We have employed a biological chemistry approach to dissect the mechanisms underpinning cellular responses to oxidant stress and to develop biologically relevant anti-oxidants. We have used telomere biology to define cellular stress responses and have observed telomere independent, p21- and p16-dependent stasis following oxidative insult in human fibroblasts. This was accompanied by a [corrected] reduction in XRCC5 expression and a reduction in [corrected] SIRT 1 expression. Using these markers in conjunction with senescence-associated beta-galactosidase expression, we have developed and screened novel nitrone based anti-oxidant compounds. We have identified functional compounds that are unsuitable for use in primary human cells. This has allowed subsequent identification of suitably structured compounds that act as superior biological anti-oxidants, which have potential for use in clinical interventions.

Postinjury vascular intimal hyperplasia in mice is completely inhibited by CD34+ bone marrow-derived progenitor cells expressing membrane-tethered anticoagulant fusion proteins.

BACKGROUND: Coagulation proteins promote neointimal hyperplasia and vascular remodelling after vessel injury, but the precise mechanisms by which they act in vivo remain undetermined. OBJECTIVES: This study, using an injury model in which the neointima is derived from bone marrow (BM)-derived cells, compared inhibition of tissue factor or thrombin on either BM-derived or existing vascular smooth muscle cells. METHODS: Two transgenic (Tg) mouse strains expressing membrane-tethered tissue factor pathway inhibitor (TFPI) or hirudin (Hir) fusion proteins driven by an alpha smooth muscle actin (SMA) promoter were generated (alpha-TFPI-Tg and alpha-Hir-Tg) and the phenotype after wire-induced endovascular injury was compared with that in wild-type (WT) controls. RESULTS: WT mice developed progressive neointimal expansion, whereas injury in either Tg was followed by repair back to a preinjured state. This was also seen when WT mice were reconstituted with BM from Tg mice but not when Tgs were reconstituted with WT BM, in which injury was followed by slowly progressive neointimal expansion. Injection of CD34+ cells from Tg mice into injured WT mice resulted in the accumulation of fusion protein-expressing cells from day 3 onwards and an absence of neointimal hyperplasia in those areas. CONCLUSIONS: Neointimal development after wire-induced endovascular injury in mice was completely inhibited when BM-derived cells infiltrating the damaged artery expressed membrane tethered anticoagulant fusion proteins under an alpha-SMA promoter. These findings enhance our understanding of the pathological role that coagulation proteins play in vascular inflammation.

Deficits in the mid-brain raphe nuclei and striatum of the AS/AGU rat, a protein kinase C-gamma mutant.

The AS/AGU rat carries a recessive mutation (agu) in the gene coding for the gamma isoform of protein kinase C. The rat is characterized by disordered locomotion and progressive dysfunction of the nigrostriatal dopaminergic (DA) system. This dysfunction begins with a failure to release DA within the striatum and culminates in cell loss within the substantia nigra pars compacta. The present study examines another midbrain aminergic system with input to the basal ganglia, the serotonergic (5-HT) raphe-striatal system originating in the dorsal raphe nucleus. By 3 months after birth, there is a very substantial reduction in the extracellular levels of 5-HT in the dorsal caudate-putamen of the mutants compared with controls (c. 70%). This is accompanied by a proportional increase in the levels of the 5-HT metabolite 5-hydroxyindole acetic acid (5-HIAA). At a later age, there are reductions in whole tissue 5-HT (and increases in 5-HIAA) in both the striatum and the region containing the dorsal raphe nucleus, as well as numbers of 5-HT-immunoreactive cells in the dorsal raphe nucleus. The median raphe appears to be unaffected. The results are seen in terms of an initial dysfunction in transmitter release leading to cell death, perhaps through the formation of free radicals or neurotoxins.