A new palladium-based antiproliferative agent: synthesis, characterization, computational calculations, cytotoxicity, and DNA binding properties.

A new palladium(II) complex entitled [Pd(phendione)(8Q)]NO3, (PdPQ), where phendione is N,N-donor heterocyclic 1,10-phenanthroline-5,6-dion and 8Q is 8-hydroxyquinolinate, has been synthesized and then characterized by molar conductivity, CHN analysis and spectral data (UV-Vis, FT-IR, NMR). DFT/ TDDFT procedures were also performed to determine the electronic structure and the nature of the electronic transitions of PdPQ. Moreover, the affinity and binding properties of DNA to the desired complex have been studied in details using electronic absorption, fluorescence, circular dichroism spectroscopies, and viscosity measurement in combination with molecular docking technique. The obtained results exhibit relatively high DNA binding values with a static quenching mechanism, which suggest that an intercalative mode plays a peridominate role in interaction process concluded by experimental/theoretical measurements. As a result of drug exposure, in vitro cytotoxicity assay demonstrated the antiproliferative activity of the PdPQ against leukemia cancer cell line, K562.

A new approach to enhance the conventional two-phase anaerobic co-digestion of food waste and sewage sludge.

BACKGROUND: Two-phase anaerobic co-digestion (TAcoD) is a versatile technology for the simultaneous treatment of organic materials and biogas production. However, the produced digestate and supernatant of the system contain heavy metals and organic substances that need to be treated prior to discharge or land application. Therefore, in this study, an innovative TAcoD for organic fertilizer and high supernatant quality achievement was proposed. METHODS: In the conventional TAcoD, mixed sewage sludge (SS) and food waste (FW) were first hydrolyzed in the acidogenic reactor, and then the hydrolyzate substrate was subjected to the methanogenic reactor (TAcoD 1). In the modified TAcoD (TAcoD 2), only FW was fed into the acidogenic reactor, and the produced hydrolyzed solid was directly converted to the organic fertilizer, while the supernatant with high soluble chemical demand (SCOD) concentration was further co-digested with SS in the methanogenic reactor. RESULTS: Although TAcoD 1 produced bio-methane yield and potential energy of 56.18% and 1.6-fold higher than TAcoD 2, the economical valorization of TAcoD 2 was 9-fold of that from TAcoD 1. The supernatant quality of TAcoD 2 was far better than TAcoD 1, since the SCOD, total nitrogen (TN), and total phosphor (TP) removal in TAcoD 2 and TAcoD 1 were 94.3%, 79.4%, 90.7%, and 68.9%, 28%, 46%, respectively. In terms of solid waste management, the modified TAcoD converted FW to organic fertilizer and achieved a solid reduction of 43.62% higher than that of conventional TAcoD. CONCLUSIONS: This new modification in two-phase anaerobic co-digestion of food waste and sewage sludge provides a potentially feasible practice for simultaneous bio-methane, organic fertilizer, and high supernatant quality achievement. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s40201-020-00603-8.

Synthesis of new ultrasonic-assisted palladium oxide nanoparticles: an in vitro evaluation on cytotoxicity and DNA/BSA binding properties.

Better solubility and improved toxicity of palladium complexes compared with cisplatin were major reasons for synthesis of novel Pd(II) complex, [Pd(8Q)(bpy)]NO3 (8Q=8-hydroxyquinolinate, bpy=2,2'-bipyridine). Interaction between the [Pd(8Q)(bpy)]NO3 complex and calf thymus DNA in aqueous solution has been investigated by circular dichroism (CD), UV-Visible absorption and fluorescence spectroscopic techniques. These experiments showed that prepared Pd(II) complex can effectively intercalate into CT-DNA and weakly bind to BSA in which the bovine serum albumin molecule was unfolded slightly. The cytotoxicity of the prepared complex has been evaluated on the MCF-7 and DU145 cell lines by MTT and TUNEL assay. The MTT results were showed that in DU145, the CC50 values of [Pd(8Q)(bpy)]NO3 and cisplatin are very close together (10.4 and 8.3 µM, respectively), unlike MCF-7. Accordingly, TUNEL assay was performed on DU145 and apoptosis was clearly obvious by 43% DNA fragmentation in the treated cell lines. So, we can suggest the [Pd(8Q)(bpy)]NO3 as alternative drug for cisplatin in the future which has great potential in DNA denaturation and apoptosis specially on prostate cancer. PdO nanoparticles were successfully prepared without supported any surfactants via sonochemical approach. The synthesized PdONPs were characterized using UV-Vis and FTIR spectroscopy, X-ray diffraction (XRD), dynamic light scattering (DLS), energy-dispersive X-ray spectroscopy (EDX), scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Communicated by Ramaswamy H. Sarma.

The role of quadriceps muscle strength in the development of falls in the elderly people, a cross-sectional study.

Background: Falls are a major health issue in the elderly people and an important cause of bone fracture. The aim of this study was to determine the association between quadriceps muscle strength (QMS) and falls in the elderly subjects. Methods: All eligible participants of the Amirkola Cohort Study entered the study. Data regarding demographic characteristics, clinical and laboratory examinations were provided between 2011 to 2014. Occurrence of falls during previous year was determined by interview and review of the medical records. The study patients were divided into low, moderate and high muscle strength groups according to QMS values ≥ 30, 15-30, and < 15 kg respectively). Association between muscle strength and falls was determined by using multiple logistic regression analysis with calculation of odds ratio (OR). Results: A total 1028 participants (females, 44.3%) were analyzed and 178(17.3%) subjects experienced a fall. Individuals with falls had higher age (p = 0.001) and lower QMS value (p = 0.001). After adjustment for all clinical and demographic variables, occurrence of falls was negatively associated with QMS and positively associated with age > 70 years old. Compared to group with QMS ≥ 30 kg, the prevalence of falls in low and moderate QMS groups increased by OR = 3(95% CI, 1.78-5.05) and 2.18 (95% CI, 1.22-3.42) respectively. Conclusion: These findings indicate that older subjects with lower QMS are at greater risk of falls. These findings provide a rational for muscle strengthening exercise in older people.

2,2'- bipyridine coplanar with coordination square of Pd(II) nonyldithiocarbamato antitumor complex interacting with DNA in two distinct steps.

Cisplatin is one of the most effective chemotherapy drugs, and has been widely employed for more than four decades in the treatment of different forms of human tumors. In recent years, various examples of metal complex-based compounds have been used for medicinal purposes. In this context, the novel palladium(II) complex, [Pd(non-dtc)(bpy)]NO3, (non-dtc = nonyldithiocarbamate and bpy = 2,2'- bipyridine) has been synthesized and characterized by means of elemental analysis, conductivity measurements, FT-IR, 1H NMR, 13C NMR, and electronic spectroscopy studies. The 50% cytotoxic concentrations (Ic50) of this Pd(II) complex (0.53 mM) and cisplatin (154 mM) against human cell tumor line (K562) indicates its interaction with DNA of cancer cell at quite low concentration. Thus, binding characteristics of this compound to calf thymus DNA (CT-DNA) has been investigated by UV-vis absorption spectroscopy and fluorescence spectra. The exciting observation of this work in the UV-visible studies was that the Pd(II) complex exhibit two or more types of interaction with CT-DNA. Such properties have rarely been observed in the literature. This complex cooperatively binds with DNA and denatures it too. Fluorescence studies proved the intercalation mode of binding and the other modes seems to be hydrophobic and electrostatic interactions. Binding parameters and thermodynamics of the interaction with CT-DNA are also described. Finally, multifunctional interactions of [Pd(non-dtc)(bpy)]NO3 make it suitable to interact with DNA of cancer cell at quite low concentration and if it is used as anticancer agent, very low doses will be needed which may have fewer side effects.

Therapeutic efficacy of cardiosphere-derived cells in a transgenic mouse model of non-ischaemic dilated cardiomyopathy.

AIM: Cardiosphere-derived cells (CDCs) produce regenerative effects in the post-infarct setting. However, it is unclear whether CDCs are beneficial in non-ischaemic dilated cardiomyopathy (DCM). We tested the effects of CDC transplantation in mice with cardiac-specific Gαq overexpression, which predictably develop progressive cardiac dilation and failure, with accelerated mortality. METHODS AND RESULTS: Wild-type mouse CDCs (10(5) cells) or vehicle only were injected intramyocardially in 6-, 8-, and 11-week-old Gαq mice. Cardiac function deteriorated in vehicle-treated mice over 3 months of follow-up, accompanied by oxidative stress, inflammation and adverse ventricular remodelling. In contrast, CDCs preserved cardiac function and volumes, improved survival, and promoted cardiomyogenesis while blunting Gαq-induced oxidative stress and inflammation in the heart. The mechanism of benefit is indirect, as long-term engraftment of transplanted cells is vanishingly low. CONCLUSIONS: Cardiosphere-derived cells reverse fundamental abnormalities in cell signalling, prevent adverse remodelling, and improve survival in a mouse model of DCM. The ability to impact favourably on disease progression in non-ischaemic heart failure heralds new potential therapeutic applications of CDCs.

Leptin enhances endothelial cell differentiation and angiogenesis in murine embryonic stem cells.

The metabolic regulation of leptin and its angiogenic effects have been well characterized in adult mammals. However, the role of leptin in the differentiation of embryonic stem cells (ESCs) to endothelial cells (ECs) has not been characterized. We hypothesized that leptin enhances the generation of ECs derived from ESCs and, in this way, promotes angiogenesis in embryonic vessels. To address this hypothesis, we utilized an in vitro model consisting of murine ESCs-derived embryoid bodies (EBs). Vascular density, EC and angiogenesis markers as well as phosphorylation levels of signal transducer and activator of transcription 3 (pSTAT3) were investigated in leptin-treated EBs and in untreated EBs as controls. ESC-derived ECs were isolated by magnetic sorting based on the expression of platelet endothelial cell adhesion molecule (PECAM-1/CD31). Significant upregulation of EC and angiogenic markers as well as higher vessel density were found in leptin-treated EBs compared to controls. CD31 positive enriched cells derived from leptin-treated EBs had improved proliferation and survival rate and showed higher levels of pSTAT3. These results suggested that leptin promotes EC differentiation and angiogenesis in mouse EBs and that janus tyrosine kinase (JAK)/STAT pathway can play a role in this biological process. Leptin-mediated EC differentiation and angiogenesis in ESCs can be a useful application towards regenerative medicine and tissue engineering.

Human cardiosphere-derived cells from advanced heart failure patients exhibit augmented functional potency in myocardial repair.

OBJECTIVES: This study sought to compare the regenerative potency of cells derived from healthy and diseased human hearts. BACKGROUND: Results from pre-clinical studies and the CADUCEUS (CArdiosphere-Derived aUtologous stem CElls to reverse ventricUlar dySfunction) trial support the notion that cardiosphere-derived cells (CDCs) from normal and recently infarcted hearts are capable of regenerating healthy heart tissue after myocardial infarction (MI). It is unknown whether CDCs derived from advanced heart failure (HF) patients retain the same regenerative potency. METHODS: In a mouse model of acute MI, we compared the regenerative potential and functional benefits of CDCs derived from 3 groups: 1) non-failing (NF) donor: healthy donor hearts post-transplantation; 2) MI: patients who had an MI 9 to 35 days before biopsy; and 3) HF: advanced cardiomyopathy tissue explanted at cardiac transplantation. RESULTS: Cell growth and phenotype were identical in all 3 groups. Injection of HF CDCs led to the greatest therapeutic benefit in mice, with the highest left ventricular ejection fraction, thickest infarct wall, most viable tissue, and least scar 3 weeks after treatment. In vitro assays revealed that HF CDCs secreted higher levels of stromal cell-derived factor (SDF)-1, which may contribute to the cells' augmented resistance to oxidative stress, enhanced angiogenesis, and improved myocyte survival. Histological analysis indicated that HF CDCs engrafted better, recruited more endogenous stem cells, and induced greater angiogenesis and cardiomyocyte cell-cycle re-entry. CDC-secreted SDF-1 levels correlated with decreases in scar mass over time in CADUCEUS patients treated with autologous CDCs. CONCLUSIONS: CDCs from advanced HF patients exhibit augmented potency in ameliorating ventricular dysfunction post-MI, possibly through SDF-1­mediated mechanisms.

The synthetic triterpenoid RTA dh404 (CDDO-dhTFEA) restores Nrf2 activity and attenuates oxidative stress, inflammation, and fibrosis in rats with chronic kidney disease.

1. Chronic oxidative stress and inflammation are major mediators of chronic kidney disease (CKD) and result in impaired activation of the cytoprotective transcription factor Nrf2. Given the role of oxidative stress and inflammation in CKD pathogenesis, strategies aimed at restoring Nrf2 activity may attenuate CKD progression. 2. The present study investigated whether the synthetic triterpenoid RTA dh404 (2-cyano-3,12-dioxooleana-1,9-dien-28-oic acid-9,11-dihydro-trifluoroethyl amide or CDDO-dhTFEA) would afford renal protection in a 5/6 nephrectomized rat model of CKD. RTA dh404 (2 mg/kg/day) was orally administered once daily for 12 weeks after 5/6 nephrectomy surgery. 3. The remnant kidneys from the vehicle-treated CKD rats showed activation of nuclear factor kappaB (NF-κB), upregulation of NAD(P)H oxidase, glomerulosclerosis, interstitial fibrosis and inflammation, as well as marked reductions in Nrf2 and its target gene products (i.e. catalase, heme oxygenase-1, thioredoxin 1, thioredoxin reductase 1 and peroxiredoxin 1). The functional and structural deficits in the kidney were associated with increased (∼30%) mean arterial pressure (MAP). Treatment with RTA dh404 restored MAP, increased Nrf2 and expression of its target genes, attenuated activation of NF-κB and transforming growth factor-ß pathways, and reduced glomerulosclerosis, interstitial fibrosis and inflammation in the CKD rats. 4. Thus, chronic treatment with RTA dh404 was effective in restoring Nrf2 activity and slowing CKD progression in rats following 5/6 nephrectomy.

The synthetic triterpenoid RTA dh404 (CDDO-dhTFEA) restores endothelial function impaired by reduced Nrf2 activity in chronic kidney disease.

Chronic kidney disease (CKD) is associated with endothelial dysfunction and accelerated cardiovascular disease, which are largely driven by systemic oxidative stress and inflammation. Oxidative stress and inflammation in CKD are associated with and, in part, due to impaired activity of the cytoprotective transcription factor Nrf2. RTA dh404 is a synthetic oleanane triterpenoid compound which potently activates Nrf2 and inhibits the pro-inflammatory transcription factor NF-κB. This study was designed to test the effects of RTA dh404 on endothelial function, inflammation, and the Nrf2-mediated antioxidative system in the aorta of rats with CKD induced by 5/6 nephrectomy. Sham-operated rats served as controls. Subgroups of CKD rats were treated orally with RTA dh404 (2 mg/kg/day) or vehicle for 12 weeks. The aortic rings from untreated CKD rats exhibited a significant reduction in the acetylcholine-induced relaxation response which was restored by RTA dh404 administration. Impaired endothelial function in the untreated CKD rats was accompanied by significant reduction of Nrf2 activity (nuclear translocation) and expression of its cytoprotective target genes, as well as accumulation of nitrotyrosine and upregulation of NAD(P)H oxidases, 12-lipoxygenase, MCP-1, and angiotensin II receptors in the aorta. These abnormalities were ameliorated by RTA dh404 administration, as demonstrated by the full or partial restoration of the expression of all the above analytes to sham control levels. Collectively, the data demonstrate that endothelial dysfunction in rats with CKD induced by 5/6 nephrectomy is associated with impaired Nrf2 activity in arterial tissue, which can be reversed with long term administration of RTA dh404.

Role of impaired Nrf2 activation in the pathogenesis of oxidative stress and inflammation in chronic tubulo-interstitial nephropathy.

BACKGROUND: Tubulo-interstitial nephropathy (TIN) is a common cause of chronic kidney disease (CKD). Consumption of an adenine-containing diet causes the accumulation of 2,8-dihydroxyadenine in the renal tubules triggering intense chronic TIN and progressive CKD in rats. CKD in this model is associated with, and largely driven by, oxidative stress and inflammation. Oxidative stress and inflammation in rats with spontaneous focal segmental glomerulosclerosis and rats with CKD induced by 5/6 nephrectomy are associated with an impaired activation of nuclear factor-erythroid-2-related factor 2 (Nrf2) which is the master regulator of genes encoding many antioxidant and detoxifying enzymes. The effect of TIN on the Nrf2 pathway and its key target genes is unknown and was investigated here. METHODS: Sprague-Dawley rats were randomized to control and adenine-treated (rat chow-containing 0.7% adenine for 2 weeks) groups and followed up for 4 weeks. RESULTS: The adenine-treated animals exhibited marked azotemia, impaired urinary concentrating capacity, intense tubular and glomerular damage, interstitial inflammation and fibrosis. This was associated with an increased expression of NAD(P)H oxidase, cyclooxygenase-2 and 12-lipoxygenase, and activation of NF-κB, the master regulator of pro-inflammatory cytokines and chemokines. Oxidative stress and inflammation in the kidneys of adenine-treated animals was accompanied by an impaired activation of Nrf2 and down-regulation of its target gene products including, catalase, heme oxygenase-1 and glutamate-cysteine ligase. CONCLUSIONS: Chronic TIN is associated with impaired Nrf2 activity which contributes to the pathogenesis of oxidative stress and inflammation and amplifies their damaging effects on the kidney.

Cardiomyocyte proliferation and progenitor cell recruitment underlie therapeutic regeneration after myocardial infarction in the adult mouse heart.

Cardiosphere-derived cells (CDCs) have been shown to regenerate infarcted myocardium in patients after myocardial infarction (MI). However, whether the cells of the newly formed myocardium originate from the proliferation of adult cardiomyocytes or from the differentiation of endogenous stem cells remains unknown. Using genetic fate mapping to mark resident myocytes in combination with long-term BrdU pulsing, we investigated the origins of postnatal cardiomyogenesis in the normal, infarcted and cell-treated adult mammalian heart. In the normal mouse heart, cardiomyocyte turnover occurs predominantly through proliferation of resident cardiomyocytes at a rate of ∼1.3-4%/year. After MI, new cardiomyocytes arise from both progenitors as well as pre-existing cardiomyocytes. Transplantation of CDCs upregulates host cardiomyocyte cycling and recruitment of endogenous progenitors, while boosting heart function and increasing viable myocardium. The observed phenomena cannot be explained by cardiomyocyte polyploidization, bi/multinucleation, cell fusion or DNA repair. Thus, CDCs induce myocardial regeneration by differentially upregulating two mechanisms of endogenous cell proliferation.

Participation of endoplasmic reticulum stress in the pathogenesis of spontaneous glomerulosclerosis--role of intra-renal angiotensin system.

Endoplasmic reticulum (ER) is the site of synthesis, folding, assembly, and degradation of proteins. Disruption of ER function leads to ER stress, which is marked by accumulation of unfolded proteins in the ER lumen. Detection of unfolded proteins by the ER membrane receptors triggers the "unfolded protein response (UPR)" designed to restore ER function via activation of the adaptive/cytoprotective responses. Failure of UPR or persistent stress triggers activation of ER stress-mediated apoptotic pathway. Several in vivo and in vitro studies have demonstrated the association of ER stress with glomerular diseases. Imai rats develop progressive glomerulosclerosis (GS), which is associated with oxidative stress, inflammation and activation of intra-renal angiotensin system, and can be prevented by AT-1 receptor blockade (ARB). Since persistent oxidative and inflammatory stresses trigger ER stress-induced apoptosis and tissue injury, we hypothesized that kidneys in the Imai rats may exhibit failure of the adaptive and activation of the apoptotic ER stress responses, which could be prevented by ARB. To this end 10-week old Imai rats were randomized to untreated and ARB-treated groups and observed for 24 weeks. At age 34 weeks, untreated rats showed heavy proteinuria, azotemia, advanced GS, impaired ER stress adaptive/cytoprotective responses (depletion of UPR-mediating proteins), and activation of ER stress apoptotic responses. ARB treatment attenuated GS, suppressed intra-renal oxidative stress, restored ER-associated adaptive/cytoprotective system, and prevented the ER stress mediated apoptotic response in this model. Thus, progressive GS in Imai rats is accompanied by activation of ER stress-associated apoptosis, which can be prevented by ARB.

Downregulation of the renal and hepatic hydrogen sulfide (H2S)-producing enzymes and capacity in chronic kidney disease.

BACKGROUND: Oxidative stress and inflammation are constant features and major mediators of progression and cardiovascular complications of chronic kidney disease (CKD). Hydrogen sulfide (H(2)S) is an endogenous signaling gas, which possesses potent anti-oxidant, anti-inflammatory, anti-hypertensive and other regulatory functions. H(2)S is produced by cystathionine ß-synthase (CBS), cystathionine γ-lyase (CSE) and 3-mercaptopyruvate sulphurtransferase (MST). Plasma H(2)S is reduced in humans with hypertension, atherosclerosis and end-stage renal disease (ESRD). Atherosclerosis, hypertension and ischemia/reperfusion-induced acute kidney injury are associated with and, in part, mediated by diminished tissue H(2)S in experimental animals. Expression of the H(2)S-producing enzymes is reduced in the circulating leukocytes of patients with ESRD. However, the effect of CKD on expression of H(2)S-producing enzymes in the diseased kidney and other tissues is unknown and was studied here. METHODS: Subgroups of rats were subjected to 5/6 nephrectomy or sham operation and observed for 6-12 weeks. Expression of H(2)S-producing enzymes and H(2)S-producing capacity was measured in kidney, liver and brain tissues. RESULTS: The CKD group exhibited oxidative stress and significant reduction of plasma H(2)S concentration. This was associated with marked reduction of H(2)S-producing capacity of the kidney and liver, marked downregulation of CBS, CSE and MST in the kidney and of CBS and CSE expression in the liver. However, expression of H(2)S-producing enzymes in the brain was not significantly altered in CKD rats. CONCLUSIONS: CKD is associated with significant reduction in plasma H(2)S concentration, diminished remnant kidney and liver tissue H(2)S-producing capacity and downregulation of the H(2)S-producing enzymes. Given the potent anti-oxidant, anti-inflammatory and cytoprotective properties of H(2)S, its deficiency may contribute to progression of CKD and the associated complications.

Effect of nephrotic syndrome on homocysteine metabolism.

BACKGROUND: Proteinuria and hyperhomocysteinaemia are independently associated with increased risk of atherosclerosis and cardiovascular disease. The available data on plasma homocysteine (Hcy) level in patients with nephrotic syndrome (NS) are contradictory with increased, decreased and unchanged values reported by different investigators. The majority of Hcy in the plasma is bound to albumin whose urinary losses and diminished plasma concentration are the defining features of NS. The present study was designed to explore the effect of NS on plasma concentration and urinary excretion of Hcy and hepatic expression of methylenetetrahydrofolate reductase (MTHFR) and cystathionine-ß-synthase (CBS), the key enzymes in re-methylation and trans-sulphuration of Hcy, respectively. METHODS: Sprague-Dawley rats were rendered nephrotic by IP injection of puromycin aminonucleoside. Urine and plasma were used for measurement of Hcy, and the liver was processed for assessment of MTHFR and CBS protein expression. RESULTS: Compared with the controls, nephrotic rats showed heavy proteinuria, hypoalbuminaemia, hypercholesterolaemia, normal plasma creatinine and creatinine clearance, reduced plasma Hcy, increased urinary Hcy, and downregulation of CBS but not MTHFR expression. Plasma Hcy correlated directly with plasma albumin and inversely with urinary protein excretion. The urinary Hcy excretion correlated directly with urine protein excretion. CONCLUSIONS: NS results in significant reduction in plasma total Hcy concentration which is due to the reduction in albumin-bound Hcy as opposed to the free Hcy fraction. This is coupled with increased urinary excretion of albumin-bound Hcy. In addition, NS results in down-regulation of CBS which can curtail conversion of Hcy to cysteine and reduce production of H(2)S which is an important endogenous signalling molecule.

Human uraemic plasma stimulates release of leptin and uptake of tumour necrosis factor-alpha in visceral adipocytes.

BACKGROUND: End-stage renal disease (ESRD) is commonly associated with anorexia, malnutrition and inflammation. In addition to serving as the primary reservoir for energy storage, adipocytes produce numerous pro- and anti-inflammatory mediators and regulate food intake by releasing the appetite-suppressing (leptin) and appetite-stimulating (adiponectin) hormones. Under normal conditions, release of leptin is stimulated by feeding to prevent excess intake, and release of adiponectin is stimulated by fasting to induce feeding. However, under certain pathological conditions such as inflammation, maladaptive release of these hormones leads to anorexia, wasting and malnutrition and simultaneously intensifies inflammation. Anorexia, malnutrition and inflammation in ESRD are frequently accompanied by hyper-leptinaemia. This study was designed to test the hypothesis that uraemic plasma may stimulate leptin release and suppress adiponectin release in normal adipocytes. METHODS: Visceral adipose tissue was harvested from normal rats, and adipocytes were isolated and incubated for 2-4 h in media containing 90% plasma from 12 ESRD patients (before and after haemodialysis) and 12 normal control subjects. RESULTS: The ESRD group had a marked elevation of plasma TNF-alpha, IL-6, IL-8 and leptin concentrations before and after haemodialysis. Incubation in media containing plasma from the ESRD group elicited a much greater leptin release by adipocytes than that containing normal plasma. Post-dialysis plasma evoked an equally intense leptin release. The rise in leptin release was coupled with a parallel fall in TNF-alpha concentration in the incubation media. In contrast to leptin, adiponectin release in the presence of uraemic plasma was similar to that found with the control plasma. CONCLUSIONS: Exposure to uraemic plasma induces exuberant release of leptin that is coupled with avid uptake of TNF-alpha by visceral adipocytes. These observations confirm the role of TNF-alpha, formerly known as cachexin, in the over-production and release of leptin in patients with ESRD.

Hyperhomocysteinemia induces insulin resistance in male Sprague-Dawley rats.

Association between elevated plasma homocysteine levels and insulin resistance has been reported, however, whether hyperhomocysteinemia induces insulin resistance or it is actually hyperinsulinemia that causes elevated plasma homocysteine levels, the direction of causality in this association is not still clear. In this study, we examined the hypothesis that hyperhomocysteinemia may cause hyperinsulinemia leading to insulin resistance in rats. Plasma glucose, insulin and total homocysteine concentrations were determined in two groups of male Sprague-Dawley rats, a test group that administered with homocysteine and a control group with no homocysteine in daily drinking water before and after 50 days. Oral glucose tolerance tests were also performed in control and test groups before and after 50 days. Mean fasting plasma insulin level was significantly higher (42.5+/-20.4 mU/L versus 23.2+/-5.9 mU/L, p=0.01), whereas mean glucose: insulin ratio was significantly lower in test rats than in control rats (0.12+/-0.07 versus 0.17+/-0.05, p=0.04) after 50 days. In addition, mean homeostasis assessment insulin resistance index was significantly higher in test rats than in control rats (7.5+/-3.5 versus 4.0+/-1.6, p=0.02) after 50 days. The mean plasma glucose level was not significantly different (4.1+/-1.1 mmol/L versus 3.9+/-0.8 mmol/L, p=0.57) between controls and test rats, however, the results from oral glucose tolerance tests showed the development of insulin resistance in test rats after 50 days administration of homocysteine. Results from this in vivo study suggest that homocysteine can cause insulin resistance and this relationship may need to be considered when evaluating the role of plasma homocysteine as a risk factor in patients with obesity and type II diabetes.

Association of methylenetetrahydrofolate reductase (C677T) polymorphism with hyperuricemia.

BACKGROUND AND AIMS: Homozygosity for the thermolabile variant of 5,10-methylene tetrahydrofolate reductase (C677T) has been suggested to be positively associated with the risk of vascular disease and neural tube defects. In addition, recent studies have suggested that elevated serum uric acid predicts ischemic heart disease, and epidemiological data on ethnic groups have suggested that genetic factors are determinants of serum uric acid levels. In this study, we tested the hypothesis that 5,10-methylenetetrahydrofolate reductase (C677T) polymorphism may be associated with hyperuricemia. METHODS AND RESULTS: Samples from 518 healthy individuals (268 men and 250 women) were analyzed for MTHFR genotyping and serum uric acid. The participants were categorized to homozygous wild type (CC), heterozygous for wild type and thermolabile (CT), or homozygous for the thermolabile (TT) variant. Serum uric acid was significantly higher in males and females with TT genotype than those with either CC or CT genotype (p=0.0001, ANOVA). Univariate and multivariate analysis showed that 5,10-methylenetetrahydrofolate reductase (C677T) polymorphism was a strong correlate and predictor of uric acid in males (r=0.28, p=0.0001, beta=0.673, p=<0.001) and in females (r=0.27, p=0.0001, beta=0.599, p=<0.001). Odds ratio analysis has also shown that the risk of hyperuricemia was greater in males (OR 3.1, CI 1.8-5.2, p=0.001) and females (OR 3.3, CI 1.9-5.7, p=<0.001) with CT genotypes and in males (OR 3.7, CI 1.3-10.7, p=0.014) and females (OR 3.2, CI 1.1-9.7, p=0.032) with TT genotypes than in those with CC genotypes. CONCLUSION: Results from this study suggest that mutation of 5-MTHFR C677T contributes to the higher uric acid levels in both males and females and may be a risk factor for hyperuricemia.

Association of low red blood cell folate concentrations with coronary artery disease in Iranians: a matched case-control study.

BACKGROUND: It is not fully established whether the increasing risk of coronary artery disease (CAD) is associated with high plasma homocysteine levels or components of the homocysteine remethylation pathway, e.g. vitamin B(12) or 5-methyltetrahydrofolate (5-MTHF) in plasma and red blood cells (RBC). In this study, we tested the hypothesis that 5-MTHF in RBC, which represents the long-term folate status of individuals, may be a more reliable marker of homocysteine remethylation pathway disturbances, and its deficiency may be associated with CAD in Iranians. METHODS: Plasma total homocysteine (tHcy), vitamin B(12), and plasma and RBC 5-MTHF were measured in 200 angiographically documented patients and 200 controls matched for sex and age. RESULTS: In the plasma, tHcy levels were significantly higher in cases compared to controls (geometric mean 12.9 +/- 6.5 vs. 10.6 +/- 5.6 micromol/l, p = 0.04). However, RBC 5-MTHF (527.2 +/- 185.9 vs. 461.3 +/- 117.9 nmol/l, p = 0.007) and vitamin B(12) (254.2 +/- 132.8 vs. 182.2 +/- 110.4 pmol/l, p = 0.04) were significantly higher in controls than patients. RBC 5-MTHF was a strong and independent predictor of plasma tHcy (beta = -0.01, p = 0.003, r(2) = 0.19). Subjects in the lowest quartile of red-cell 5-MTHF had a 2.5-fold increased prevalence of CAD compared to subjects in the highest quartile. The association of CAD in the first quartile with red-cell 5-MTHF remained significant when adjusted for plasma tHcy, vitamin B(12), hypertension and hypercholesterolemia (odds ratio, OR 2.3, confidence interval: 1.1-3.9, p = 0.01). However, the association between CAD in the highest quartile and plasma tHcy decreased and became insignificant when adjusted for red-cell 5-MTHF, vitamin B(12), hypertension and hypercholesterolemia (OR 1.27, confidence interval: 0.96-1.69, p = 0.11). CONCLUSION: In this study, the association between CAD and low RBC 5-MTHF was stronger than with plasma 5-MTHF and plasma tHcy levels, indicating that RBC 5-MTHF may be a more stable parameter to study disturbances in the homocysteine remethylation pathway in Iranians.

Association of plasma folate, plasma total homocysteine, but not methylenetetrahydrofolate reductase C667T polymorphism, with bone mineral density in postmenopausal Iranian women: a cross-sectional study.

Polymorphisms of methylenetetrahydrofolate reductase (MTHFR) have been well documented to cause hyperhomocysteinemia, and recent studies suggest an association of C677T mutation of methylenetetrahydrofolate reductase with low bone mineral density (BMD). In this study, the association of plasma total homocysteine (Hcy), plasma folate, and vitamin B12 as well as methylenetetrahydrofolate reductase C667T polymorphism with bone mineral density at neck of femur and lumbar spine in 271 postmenopausal Iranian women was investigated. Bone mineral density was measured by dual-energy X-ray absorptiometry. Restriction fragment length polymorphism was used for genotyping the methylenetetrahydrofolate reductase polymorphism. Plasma total homocysteine, plasma folate, and vitamin B12 were also determined. The bone mineral densities at the neck of femur and lumbar spine together with other clinical characteristics among methylenetetrahydrofolate reductase genotypes (CC, CT, and TT) were examined. Bone mineral densities at both neck of femur (r = -0.18, P = 0.003) and lumbar spine (r = -0.16, P = 0.01) were significantly and negatively correlated with the logarithm of plasma total homocysteine. Bone mineral density at the lumbar spine was also significantly and positively associated with plasma folate (r = 0.14, P = 0.02). However, no correlation between methylenetetrahydrofolate reductase polymorphism with bone mineral density at neck of femur (r = -0.01, P = 0.81) and lumbar spine (r = -0.04, P = 0.51) was observed. The negative association of plasma total homocysteine with bone mineral density was no longer significant when adjusted for folate and vitamin B12. Plasma folate and age were the main predictors of plasma total homocysteine explaining 15.3% and 5.2% of the variance of plasma total homocysteine, respectively. Methylenetetrahydrofolate reductase polymorphism, however, was not associated with plasma folate (r = 0.086, P = 0.17) or vitamin B12 (r = 0.05, P = 0.4). Plasma folate was one of the main predictors explaining 3.0% and 1.7% of variance of the bone mineral density at femoral neck and lumbar spine, respectively. Results from this study suggest hyperhomocysteinemia as a result of folate deficiency, but not methylenetetrahydrofolate reductase polymorphism, is independently associated with low bone mineral density and may contribute to the pathogenicity of osteoporosis in postmenopausal Iranian women.