Noradrenergic signaling controls Alzheimer's disease pathology via activation of microglial ß2 adrenergic receptors.

In Alzheimer's disease (AD) pathophysiology, plaque and tangle accumulation trigger an inflammatory response that mounts positive feed-back loops between inflammation and protein aggregation, aggravating neurite damage and neuronal death. One of the earliest brain regions to undergo neurodegeneration is the locus coeruleus (LC), the predominant site of norepinephrine (NE) production in the central nervous system (CNS). In animal models of AD, dampening the impact of noradrenergic signaling pathways, either through administration of beta blockers or pharmacological ablation of the LC, heightened neuroinflammation through increased levels of pro-inflammatory mediators. Since microglia are the resident immune cells of the CNS, it is reasonable to postulate that they are responsible for translating the loss of NE tone into exacerbated disease pathology. Recent findings from our lab demonstrated that noradrenergic signaling inhibits microglia dynamics via ß2 adrenergic receptors (ß2ARs), suggesting a potential anti-inflammatory role for microglial ß2AR signaling. Thus, we hypothesize that microglial ß2 adrenergic signaling is progressively impaired during AD progression, which leads to the chronic immune vigilant state of microglia that worsens disease pathology. First, we characterized changes in microglial ß2AR signaling as a function of amyloid pathology. We found that LC neurons and their projections degenerate early and progressively in the 5xFAD mouse model of AD; accompanied by mild decrease in the levels of norepinephrine and its metabolite normetanephrine. Interestingly, while 5xFAD microglia, especially plaque-associated microglia, significant downregulated ß2AR gene expression early in amyloid pathology, they did not lose their responsiveness to ß2AR stimulation. Most importantly, we demonstrated that specific microglial ß2AR deletion worsened disease pathology while chronic ß2AR stimulation resulted in attenuation of amyloid pathology and associated neuritic damage, suggesting microglial ß2AR might be used as potential therapeutic target to modify AD pathology.

Degradation kinetics and mechanism of pentoxifylline by ultraviolet activated peroxydisulfate.

Degradation of pentoxifylline (PTX) by sodium peroxydisulfate (SPDS) assisted by UV irradiation has been investigated in deionized water. The treatment was more favorable over direct photolysis or peroxydisulfate oxidation alone. The effects of various parameters, including different dosage of oxidant agent, PTX concentration, initial solution pH levels, and the presence of inorganic ions like chloride, nitrate and carbonate have been evaluated. The rate of PTX decomposition depends on the oxidant agent dose. The highest degradation was determined at pH 10.5, which can be explained by the generation of additional hydroxyl radicals (HO˙) in the reaction between sulfate radicals and hydroxide ions. The presence of inorganic ions, especially the carbonate ions quench valuable sulfate radicals and have successfully retarded the PTX decomposition. Six PTX oxidation products were identified using UPLC-QTOF-MS for trials in a basic environment. The main degradation product (3,7-dimethyl-6-(5-oxohexyloxy)-3,7-dihydro-2H-purin-2-one) was isolated by column chromatography and identified by 1HNMR and LC MS analyzes.

Common polymorphism (81Val>Ile) and rare mutations (257Arg>Ser and 335Ile>Ser) of the MC3R gene in obese Polish children and adolescents.

The predisposing role to human obesity of the MC3R gene polymorphism is controversial. In this report we present the first study focused on the search for the MC3R polymorphism in the Polish population. Altogether 257 obese children and adolescents (RBMI>120) and 94 adults, who were never obese or overweight (BMI<25), were studied. For all subjects the entire coding sequence was analyzed by direct DNA sequencing. One common polymorphism (81Val>Ile) and two rare mutations (257Arg>Ser and 335Ile>Ser) were identified. The common polymorphism was widely distributed in the obese and control cohorts, while the mutations were identified in four obese subjects only. In case of the 335Ile>Ser substitution a three-generation family, consisting of 20 members, was also analyzed. It was found that all carriers of the 335Ser mutation were obese, but among non-carriers obese subjects also were found. Our study suggests that the predisposing effect to obesity of the 81Ile polymorphic variant is rather unlikely. With regard to the studied rare mutations we suggest that the 335Ser allele may have a small predisposing effect.

Epigenetic mechanisms in anti-cancer actions of bioactive food components--the implications in cancer prevention.

The hallmarks of carcinogenesis are aberrations in gene expression and protein function caused by both genetic and epigenetic modifications. Epigenetics refers to the changes in gene expression programming that alter the phenotype in the absence of a change in DNA sequence. Epigenetic modifications, which include amongst others DNA methylation, covalent modifications of histone tails and regulation by non-coding RNAs, play a significant role in normal development and genome stability. The changes are dynamic and serve as an adaptation mechanism to a wide variety of environmental and social factors including diet. A number of studies have provided evidence that some natural bioactive compounds found in food and herbs can modulate gene expression by targeting different elements of the epigenetic machinery. Nutrients that are components of one-carbon metabolism, such as folate, riboflavin, pyridoxine, cobalamin, choline, betaine and methionine, affect DNA methylation by regulating the levels of S-adenosyl-L-methionine, a methyl group donor, and S-adenosyl-L-homocysteine, which is an inhibitor of enzymes catalyzing the DNA methylation reaction. Other natural compounds target histone modifications and levels of non-coding RNAs such as vitamin D, which recruits histone acetylases, or resveratrol, which activates the deacetylase sirtuin and regulates oncogenic and tumour suppressor micro-RNAs. As epigenetic abnormalities have been shown to be both causative and contributing factors in different health conditions including cancer, natural compounds that are direct or indirect regulators of the epigenome constitute an excellent approach in cancer prevention and potentially in anti-cancer therapy.

Effect of three common SNPs in 5'-flanking region of LEP and ADIPOQ genes on their expression in Polish obese children and adolescents.

Genes encoding adipokines are considered as candidates for human obesity. In this study we analyzed the expression of leptin (LEP) and adiponectin (ADIPOQ) genes in relation to common 5'-flanking or 5'UTR variants: -2548G>A (LEP), 19A>G (LEP) and -11377C>G (ADIPOQ) in Polish obese children and adolescents. Relative transcription levels in the subcutaneous adipose tissue (real time RT-PCR) and serum protein concentrations (RIA) were measured in 48 obese subjects with known genotypes at three polymorphic sites and in five non-obese controls. None of the studied polymorphisms altered significantly the expression. Significantly elevated relative transcription levels of the LEP gene (P < 0.05) and serum leptin concentrations (P < 0.01) were recorded in obese patients, when compared with the non-obese controls, but such differences were not found for the ADIPOQ gene. Interestingly, the leptin to adiponectin protein concentration ratio (L/A) was approximately sevenfold higher in obese children and adolescents when compared with the non-obese controls (P < 0.001). Taking into consideration the observed relationship between the genotypes and the gene expression level we suggest that these SNPs are not conclusive markers for predisposition to obesity in Polish children and adolescents. On the other hand, we confirmed that the leptin to adiponectin gene expression ratio (L/A) is an informative index characterizing obesity.

The effects of nucleoside analogues on promoter methylation of selected tumor suppressor genes in MCF-7 and MDA-MB-231 breast cancer cell lines.

The effects of 2-chloro-2'-deoxyadenosine, 9-beta-D-arabinofuranosyl-2-fluoroadenine, and 5-aza-2'-deoxycytidine on promoter methylation of the selected tumor suppressor genes (i.e., ERalpha, BRCA1, RARbeta2, E-cadherin, PTEN, and APC) were estimated using methylation-sensitive restriction analysis. The studies were carried out in hormone-responsive, low-invasive cell line MCF-7 and hormone-insensitive, highly invasive cell line MDA-MB-231. The results demonstrate an implication of the tested adenosine analogues and 5-aza-dCyd in regulation of DNA methylation process. Moreover, the effects of nucleoside analogues on PTEN promoter methylation suggest distinct mechanism of regulation of the epigenetic DNA modification in low-invasive compared to highly invasive breast cancer cells.

Studies on the methylation status of CpG sequences located in promoters of selected tumour suppressor genes in breast cancer cells.

In the tested samples of sporadic breast cancer (100 cases), hypermethylation of CpG sequences located in ERalpha promoter was observed in 62 cases. It correlated with: (i) deficiency of ERalpha protein in 45%, (ii) hypermethylation of BRCA1 promoter in 95%, and (iii) nonmethylated E-cadherin promoter in 90%. Fifty-eight percent of the patients with nonmethylated E-cadherin promoter (56 cases) did not show metastasis to lymphatic nodes. The analysis of the methylation level of the promoter of ERalpha, BRCA1, and E-cadherin, frequently connected with their activity, shows that it can be an important parameter in the diagnosis and therapeutic strategies in sporadic breast cancer.

Alteration of DNA methylation status in K562 and MCF-7 cancer cell lines by nucleoside analogues.

The effects of 2-chloro-2'-deoxyadenosine, beta-D-arabinofuranosyl-2-fluoroadenine, and 5-aza-2'-deoxycytidine on promoter methylation of the selected tumor suppressor genes (i.e., ERalpha, BRCA1, E-cadherin, PTEN, and APC) were estimated using methylation-sensitive restriction analysis (MSRA) in K562 cells (human erythroleukemic cell line) and MCF-7 cells (human breast cancer cell line). In both cell lines all tested drugs completely reduced methylation of PTEN and APC promoters. The results indicate that the tested nucleoside analogues, which are known inhibitors of DNA synthesis, also are implicated in indirect (or direct in the case of 5-aza-dCyd) regulation of post-replicative DNA modifications (i.e., DNA methylation).

Bone mineral density, chemical composition and biomechanical properties of the tibia of female rats exposed to cadmium since weaning up to skeletal maturity.

The influence of exposure to cadmium (Cd) during skeletal development on the risk of bone fractures at the stage of skeletal maturity was investigated on a female rat model of human exposure. The tibias of rats treated with 1, 5 or 50 mg Cd/l in drinking water for 3, 6, 9 and 12 months (since weaning) were used. The exposure to Cd dose- and time-dependently influenced the tibia bone mineral density (BMD) and chemical composition. In skeletally matured animals, at each level of the exposure to Cd, the BMD at the whole tibia and its diaphysis as well as the percentage of minerals content in the bone, including the content of zinc, copper and iron, were decreased compared to control. Moreover, in the 50 mg Cd/l group, the percentage of organic components content increased. The Cd-induced changes, at all levels of exposure, resulted in weakening in the yield strength and fracture strength of the tibia (a three-point bending test of the diaphysis and compression test with vertical loading) of the skeletally matured females. A very important and clinically useful finding of this study is that a decrease (even by several percent) in the tibia BMD results in weakness in the bone biomechanical properties and that the BMD may predict the risk of its fracture at the exposure to Cd. Moreover, the results together with our previous findings seem to suggest that tibia, due to higher vulnerability of its diaphysis, compared to the femoral diaphysis, to damage by Cd may be more useful than femur to investigate the effect of Cd on the cortical bone. The present study revealed that a low exposure to Cd (1 mg Cd/l), corresponding to low human environmental exposure, during the skeletal development affects the tibia mineral status leading to weakening in its mechanical properties at the skeletal maturity. The findings allow for the conclusion that environmental exposure to Cd during childhood and adolescence may enhance the risk of low BMD and fractures at adulthood.

Mechanical properties of femoral diaphysis and femoral neck of female rats chronically exposed to various levels of cadmium.

The effect of chronic exposure to cadmium (Cd) on the mechanical properties of femoral diaphysis and femoral neck was investigated on a rat model of human exposure. Three-week-old female Wistar rats were exposed to Cd in drinking water at concentrations of 1, 5, 50, or 100 mg/L for 12 months. Biomechanical properties of the femoral diaphysis were evaluated in a three-point bending test and those of the femoral neck in a bending test with vertical loading of the head. Bone mineral content (BMC) and bone mineral density (BMD) at the whole femur, and BMD at the diaphysis and proximal femur (head and neck region) of the Cd-treated rats decreased in a dose-dependent manner, except for the diaphyseal BMD at a Cd concentration of 1 mg/L. Exposure to Cd concentrations of 1 and 5 mg/L had only little effect on the diaphyseal mechanical properties (decreased yield load with unchanged bending strength, stiffness, yield stress, ultimate stress, and Young modulus), whereas the bending strength and stiffness of the neck decreased and the yield load clearly tended to decline or declined. The effect of Cd at the two locations was more marked in the 50 and 100 mg/L groups, and changes in the bone geometry were observed in these animals. The results clearly revealed that chronic, even low-level, exposure to Cd results in demineralization and weakening of the femur. The femoral neck seems to be more vulnerable than the diaphysis to failure from Cd. We conclude that environmental exposure to Cd may be an important risk factor for femoral neck fracture.

Mineral status and mechanical properties of lumbar spine of female rats chronically exposed to various levels of cadmium.

The effects of chronic exposure to cadmium (Cd) on the mineral status, mechanical properties and incidence of deformities and fractures of the lumbar spine (L1-L5) were studied in a rat model of human exposure. Young female Wistar rats were exposed to 1, 5, 50 or 100 mg Cd/l for 12 months. Cd, dose and time dependently, disturbed the mineral status of the lumbar vertebrae as reflected in decreased bone mineral content (BMC) and density (BMD) at the L1-L5 (DEXA technique) and ash weight (AW) of the fourth lumbar vertebral body (L4). However, the changes were too small to be evident radiographically. Cd had no effect on the ratio of nonorganic to organic component content, except for its decrease at the 100 mg Cd/l. Weakness in the mechanical properties (compression test; Instron machine) of the L4 was noted. At the 1 mg Cd/l, a decrease was observed in the deformation at the yield point, with a simultaneous increase in the L4 stiffness, but not in strength (defined by load at yield or ultimate load). In the 5 mg Cd/l group, similar changes took place and a decrease in the ultimate load was evident as well. At the 50 and 100 mg Cd/l, Cd more seriously affected the L4 mechanical properties. At all levels of Cd exposure, the L4 deformities and/or fractures took place. Intact L4 was noted only in the 1 and 5 mg Cd/l groups. The study clearly revealed that chronic exposure to Cd disturbs the L1-L5 mineral status resulting in weakness in its mechanical properties and in turn in vertebral body (cancellous bone) deformities and fractures. The results allow us to conclude that the critical Cd concentration for these effects is very low [about 0.06-0.09 microg/g dry defatted weight (DW)] and seem to indicate an osteoporotic character of changes. A very important finding of the study is that Cd affects cancellous bone even at low-level intoxication corresponding to the general population exposure. Thus, we hypothesize that environmental exposure to Cd may be a risk factor for the lumbar spine demineralization and increased incidence of vertebral deformities and fractures.

Effect of addition of grit made of crude and expanded amaranth seeds on the quality of canned meat.

Grit made of crude and expanded amaranth seeds was added to meat-fat batter in order to determine its effect on its quality. Amaranthus cruentus seeds were expanded by the nonpressure method and graded by density (degree of expansion) into three fractions: 341, 208 and 135 kg/m3; crude seed density was 800 kg/m3. Then the seeds were ground to a homogeneous mass with granulation of 0.8 mm. The water-holding capacity and viscosity of grit made of crude and expanded seeds were determined. Class II pork was used for the production of meat-fat batter. Cooking losses were determined in model meat-fat batter, pasteurized and sterilized cans. The texture and color parameters of canned meat were also determined, and a taste-panel evaluation was made. It was found that expanded seed grit was characterized by worse hydration and gelling properties than crude seed grit. Grit made of crude amaranth seeds had a positive effect on the water-holding capacity of stuffing and allowed to reduce the cooking losses in cans. Better water-holding capacity resulted in an improvement in the tenderness, juiciness and taste of canned meat. Its quality deteriorated when large amounts of highly-expanded seed grit were used.

[Dynamic of IL-6 and IL-8 concentrations in patients after surgery treated with total parenteral nutrition]. / Dynamika zmian stezenia interleukin 6 i 8 u chorych po zabiegach chirurgicznych leczonych calkowitym zywieniem pozajelitowym.

Exaggerated cytokines productions and development of systemic inflammatory response (SIRS) is the most common cause of postoperative complications and death after major abdominal surgery. The present study was conducted to investigate alterations in systemic production of interleukin-6 (IL-6) and interleukin-8 (IL-8) after total parenteral nutrition (TPN) in surgical patients. Plasma concentrations of IL-6 and IL-8 were measured in 22 patients (10 treated with TPN and 12 without TPN) before major surgery and on the days 1, 3, 7, 10 and 14-16 after, by ELISA test (indications for surgery: stomach, pancreatic and colon carcinoma, complications of IBD and acute pancreatitis). There were no differences between preoperative levels of IL-6 and IL-8 in the examined groups of patients. The highest (on the days 1, 3, 7, 10: 268.3 (p = 0.002), 41.9 (p = 0.03), 122.6 (p = 0.009), 29.3 (p = 0.03) pg/ml respectively) and longer lasting significantly elevated level of IL-6 was observed in the group of patients after major surgery without TPN. In the group of patients received TPN (with glutamine) there was a significantly increased but in comparison with group of patients without TPN, significantly lower level of IL-6 on days 1 and 7 (103.4 and 34.7 pg/ml respectively, p = 0.01). There was no significant change in postoperative concentration of IL-8 after major surgery in the group of patients treated with TPN. The level of IL-8 was significantly elevated (p = 0.01) in the group of patients without TPN on day 1 and 3 following surgery. The IL-8 level in the TPN group vs. group of patients without TPN was significantly lower on day 1 after surgery. After TPN concentration of cholesterol was significantly higher and CRP level significantly lower. We conclude that TPN improved immunological response to major surgical trauma by reduction of the inflammatory response.

Distinct inhibitory effects of 2-chloro-2'-deoxyadenosine and 9-beta-D-arabinosyl-2-fluoroadenine on DNA methyltransferase in human T-lymphocytes.

The effect of 2-chloro-2'-deoxyadenosine and 9-beta-D-arabinosyl-2-fluoroadenine on DNA methyltransferase activity in stimulated human T-lymphocytes was estimated. In comparative studies 5-aza-deoxycytidine and deoxyadenosine plus deoxycoformycin were used. These antileukemic compounds demonstrated different effects; both 2CdA and dAdo plus dCF, like 5-aza-dCyt, inhibited the enzyme activity by 85-90% after 72 hours activation of lymphocytes, while the effect of F-ara-A, under the same conditions, was insignificant.

Common resistance mechanisms to deoxynucleoside analogues in variants of the human erythroleukaemic line K562.

Resistant variants of the human leukaemic line K562 were developed using selection with the deoxynucleoside analogues cytosine arabinoside, 2-chlorodeoxyadenosine, fludarabine and gemcitabine. The resistant lines displayed a high degree of cross resistance to all deoxynucleoside analogues, with little or no cross resistance to other agents. There was a profound accumulation defect of all nucleoside analogues in the resistant variants but no significant defect in nucleoside transport in any of the variants. 5' nucleotidase activity was strongly increased and deoxycytidine kinase activity was moderately reduced in all of the resistant variants, resulting in reduced accumulation of triphosphate analogues. In addition a deletion in one of the alleles of the deoxycytidine kinase was detected in the fludarabine-resistant line. Ribonucleotide reductase activity was found to be strongly increased in the gemcitabine-selected line and purine nucleoside phosphorylase was increased in the 2-chlorodeoxyadenosine-selected line. Free nucleotide pools were increased in the 2-chlorodeoxyadenosine-selected line. There was no expression of the mdr1 gene by the resistant lines. Karyotypic analysis and FISH experiments using a 6q21 specific probe showed alterations in the 6(q16-q22) region which contains the 5'-nucleotidase gene. Early events in the activation and degradation of deoxynucleoside analogues appear to constitute common mechanisms of resistance to these compounds.

[Clinical evaluation of amino acid solution]. / Ocena kliniczna roztworu aminokwasów.

Glutamine, a conditionally essential amino acid, is important for immune function. It is now being formulated for incorporation in to total parenteral nutrition (TPN). The aims of this study were to examine the effect of glutamine administration on clinical status, body composition, protein synthesis and immune function in critically ill patients with sepsis. Eleven patients were included into the study. Seven of them have been on conventional TPN for 8-63 days without significant clinical and nutritional improvement. Glutamine supplemented TPN was implemented for 10 subsequent days. Before, during and after the study venous blood samples were taken for the cellular immunity examination, and for the measurement of plasma albumin, transferrin and triglycerides levels. Nitrogen balance was calculated every day during the study. No side effects were noted. Patients receiving amino acid solution revealed improved nitrogen balance, body composition and body water distribution. Plasma proteins concentrations and immunological indices significantly increased during ten days TPN with Glamin. We confirm the beneficial effects of amino acid solution-supplemented TPN on nitrogen balance, plasma proteins, and immune status of critically ill patients.

[Parenteral nutrition in postoperative period]. / Zywienie dojelitowe w okresie okolooperacyjnym.

Results of surgical procedures in malnutrished patients are worse than in well-nutrished patients. Perioperative nutritional treatment lead to diminished percentage of postoperative complications and mortality. Authors presents the influence of early postoperative enteral nutrition in 35 malnutrished patients on body composition and anthropometrical and biochemical parameters of their nutritional status. Results shows that enteral nutrition in postoperative phase can decrease catabolic effect of surgical treatment.