DESIGN AND SYNTHESIS OF NOVEL SYNTHETIC ANTIOXIDANTS FOR THE TREATMENT OF OXIDATIVE STRESS RELATED DISEASES

Free radicals play an important role in many physiological processes that occur in the human body such as cellular defense responses to infectious agents and a variety of cellular signaling pathways. While at low concentrations free radicals are involved in many significant metabolic reactions, high levels of free radicals can have deleterious effects on biomolecules like proteins, lipids, and DNA. Many physiological disorders such as diabetes, ageing, neurodegenerative diseases, and ischemia-reperfusion (I/R) injury are associated with oxidative stress.1 In particular, the deleterious effects caused by I/R injury developed during organ transplantation, cardiac infarct, and stroke have become the main cause of death in the United States and Europe.1,2 In this context, we synthesized and characterized a series of novel indole-amino acid conjugates as potential antioxidants for I/R injury. The synthesis of indole-phenol conjugate compounds is also discussed. Phenolic derivatives such as caffeic acid, butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA), resveratrol, and its analogues are known for their significant antioxidative properties. A series of resveratrol analogues have been designed and synthesized as potential antioxidants. The radical scavenging mechanisms for potential antioxidants and assays for the in vitro evaluation of antioxidant activities are also discussed.

Metabolic predictors of inflammation, adhesion, and coagulability in healthy younger-aged adults

Elevated levels of inflammatory biomarkers are associated with the pathophysiology of cardiovascular diseases and are predictors of cardiovascular events. The objective of this study was to determine the unique contributions of metabolic factors as predictors of inflammation (C-reactive protein (CRP) and interleukin-6 (IL-6)), adhesion (soluble intercellular adhesion molecule-1 (sICAM-1)), and coagulation (D-dimer) in healthy younger-aged adults. Participants were 83 women and 92 men (mean age 30.04 years, s.d. +/- 4.8, range 22-39) of normal weight to moderate obese weight (mean BMI 24.4 kg/m(2), s.d. +/- 3.35, range 17-32). The primary data analytical approaches included Pearson correlation and multiple linear regression. Circulating levels of CRP, IL-6, sICAM-1, and D-dimer were determined in plasma. Higher levels of CRP were independently associated with higher BMI, a greater waist-to-hip ratio, female gender, and higher triglycerides (P < 0.001). Higher IL-6 levels were independently associated with a greater waist-to-hip ratio (P < 0.01). Higher levels of sICAM-1 were independently associated with higher BMI, higher triglycerides, and lower insulin resistance (P < 0.001). Higher D-dimer levels were independently associated with higher BMI and being female (P < 0.001). Having a higher BMI was most consistently associated with elevated biomarkers of inflammation, adhesion, and coagulation in this sample of healthy younger-aged adults, although female gender, insulin resistance, and lipid levels were also related to the biomarkers. The findings provide insight into the adverse cardiovascular risk associated with elevated body weight in younger adults.

Low bone mineral density, renal dysfunction, and fracture risk in HIV infection: a cross-sectional study

BACKGROUND: Reduced bone mineral density (BMD) is common in adults infected with human immunodeficiency virus (HIV). The role of proximal renal tubular dysfunction (PRTD) and alterations in bone metabolism in HIV-related low BMD are incompletely understood. METHODS: We quantified BMD (dual-energy x-ray absorptiometry), blood and urinary markers of bone metabolism and renal function, and risk factors for low BMD (hip or spine T score, -1 or less) in an ambulatory care setting. We determined factors associated with low BMD and calculated 10-year fracture risks using the World Health Organization FRAX equation. RESULTS: We studied 153 adults (98% men; median age, 48 years; median body mass index, 24.5; 67 [44%] were receiving tenofovir, 81 [53%] were receiving a boosted protease inhibitor [PI]). Sixty-five participants (42%) had low BMD, and 11 (7%) had PRTD. PI therapy was associated with low BMD in multivariable analysis (odds ratio, 2.69; 95% confidence interval, 1.09-6.63). Tenofovir use was associated with increased osteoblast and osteoclast activity (P< or = .002). The mean estimated 10-year risks were 1.2% for hip fracture and 5.4% for any major osteoporotic fracture. CONCLUSIONS: In this mostly male population, low BMD was significantly associated with PI therapy. Tenofovir recipients showed evidence of increased bone turnover. Measurement of BMD and estimation of fracture risk may be warranted in treated HIV-infected adults.

Non-invasive quantitative estimation of bone density in rats throughout the life cycle and in arthritic osteopenia: preliminary results

A longitudinal bone survey was conducted in 86 female Wistar rats in order to assess mineral density kinetics from young age (5 weeks: 115 g) till late adulthood (64 weeks: 586 g). In vivo quantitative radiographic scanning was performed on the caudal vertebrae, taking trabecular mass as the parameter. Measurements were expressed as Relative Optical Density (ROD) units by means of a high resolution densitometric device. Results showed a progressive increase in mineral density throughout the life cycle, with a tendency to level in the higher weight range, indicating that progressive mineral aposition occurs in rats in dependency of age. This phenomenon, however, should be always considered within the context of continuous skeletal growth and related changes typical of this species. Twelve different animals were also examined following induction of articular inflammation with Freund's adjuvant in six of them. Bone survey conducted 12 to 18 days after inoculation revealed a significant (P less than 0.01) reduction in trabecular bone mass of scanned vertebrae in comparison with the weight-matched untreated controls. It is concluded that the in vivo quantitative assessment of bone density illustrated in this report represents a sensitive and useful tool for the long-term survey of naturally occurring or experimentally induced bone changes. Scanning of the same part of the skeleton can be repeated, thereby avoiding sacrifice of the animal and time-consuming preparation of post-mortem material.

Targeted Multistage Delivery of Nanoparticles to the Bone Marrow

Bone marrow is a target organ site involved in multiple diseases including myeloproliferative disorders and hematologic malignancies and metastases from breast and prostate. Most of these diseases are characterized with poor quality of life, and the treatment options are only palliative due to lack of delivery mechanisms for systemically injected drugs which results in dose limitation to protect the healthy hematopoietic cells. Therefore, there is a critical need to develop effective therapeutic strategies that allow for selective delivery of therapeutic payload to the bone marrow. Nanotechnology-based drug delivery systems provide the opportunity to deliver drugs to the target tissue while decreasing exposure to normal tissues. E-selectin is constitutively expressed on the bone marrow vasculature, but almost absent in normal vessels, and therefore, E-selectin targeted drug delivery presents an ideal strategy for the delivery of therapeutic nanoparticles to the bone marrow. The objective of this study was to develop a novel bone marrow targeted multistage vector (MSV) via E-selectin for delivery of therapeutics and imaging agents. To achieve this goal, Firstly, an E-selectin thioaptamer (ESTA) ligand was identified through a two-step screening from a combinatorial thioaptamer library. Next, ESTA-conjugated MSV (ESTA-MSV) were developed and evaluated for their stability and binding to E-selectin expressing endothelial cells. Different types of nanoparticles including liposomes, quantum dots, and iron oxide nanoparticles were loaded into the porous structure of ESTA-MSV. In vivo targeting experiments demonstrated 8-fold higher accumulation of ESTA-MSV in the mouse bone marrow as compared to non-targeted MSV Furthermore, intravenous injection of liposomes loaded ESTA-MSV resulted in a significantly higher accumulation of liposome in the bone marrow space as compared to injection of non-targeted MSV or liposomes alone. Overall this study provides first evidence that E-selectin targeted multistage vector preferentially targets to bone marrow vasculature and delivers larger amounts of nanoparticles. This delivery strategy holds potential for the selective delivery of large amounts of therapeutic payload to the vascular niches in the bone marrow for the treatment of bone marrow associated diseases.

The Interactions of Nonalcoholic Fatty Liver Disease and Cardiovascular Diseases

A complex interaction among metabolic factors, adipose tissue lipolysis, oxidative stress, and insulin resistance results in a deleterious process that may link nonalcoholic fatty liver disease (NAFLD) with severe cardiovascular (CV) outcomes. Patients with NAFLD are at higher risk of atherosclerosis, new onset of CV events, and overall mortality. The strong association between NAFLD and CV disease should affect clinical practice, with screening and surveillance of patients with NAFLD. This review discusses the data linking these major diseases.

In vitro release of dimethyloxaloylglycine and l-mimosine from bovine bone mineral.

OBJECTIVE Prolyl hydroxylases (PHD) are oxygen sensors and therefore pharmacological targets to stimulate periodontal regeneration. Here we evaluate the release profile of the PHD inhibitors dimethyloxaloylglycine and l-mimosine from bone substitutes. MATERIALS Dimethyloxaloylglycine and l-mimosine were lyophilised onto bone substitutes including bovine bone mineral, beta-tricalcium phosphate, and hydroxyapatite. Release kinetic was evaluated by bioassays with gingival and periodontal ligament fibroblasts. We determined the capacity of PHD inhibitors to provoke VEGF expression and to repress metabolic activity and proliferation as assessed by immunoassay, MTT conversion and (3)[H]thymidine incorporation, respectively. RESULTS We found that the PHD inhibitors are released from bovine bone mineral as indicated by the increase of VEGF production in gingival and periodontal ligament fibroblasts. Supernatants obtained after 1h also decreased metabolic activity and proliferation of the fibroblasts. A fibrin matrix prolonged the release of PHD inhibitors up to 192h. A similar cellular response was found when supernatants from PHD inhibitors loaded beta-tricalcium phosphate and hydroxyapatite embedded in fibrin were assessed. CONCLUSIONS In conclusion bone substitutes can serve as carriers for PHD inhibitors that maintain their capacity to provoke a pro-angiogenic response in vitro. These findings provide the basis for preclinical studies to evaluate if this release kinetic can stimulate periodontal regeneration.

Möller-Barlow disease in archaeology: Preliminary study of biochemical detection

Human bone is the most direct source for reconstructing health and living conditions of ancient populations. However, many diseases remain undetected in palaeopathology. Möller-Barlow disease (scurvy) is a historically well-documented metabolic disease and must have been common in clinical and sub-clinical severity. Due to long incubation periods and the subtle nature of bone changes osteological evidence is relatively rare (Brickley & Ives 2008). Möller-Barlow disease is caused by deficiency of dietary vitamin C (ascorbic acid) and evokes symptoms like fatigue, haemorrhage, inflammations, delayed wound healing and pain. Vitamin C is a cofactor for the hydroxylation of the amino acids proline and lysine which are essential for the production of intact connective tissue by cross-linking the propeptides in collagen. In a preliminary study we tested the detectability of Möller-Barlow disease by analysis of relative quantitative variability of hydroxylated amino acids in collagen (Pendery & Koon 2013). Samples (N=9) were taken from children with (n=3, cranium, femur, tibia) and without (n=4, cranium, femur, tibia) apparent bone reactions indicative of Möller-Barlow disease, as well as from adults with lethal traumata (n=2; negative controls). The skeletal remains originated from two early medieval cemeteries from Switzerland. Gas chromatographic (GC) analysis revealed minor differences between the samples. So far children with no pathologic alterations had fairly same values as negative controls while children with bone reactions paradoxically exhibited even slightly higher values of hydroxyproline and hydroxylysine. Future research demands for larger sample size and has to discuss sampling strategies. Beside possible misdiagnosis of Möller-Barlow disease it is arguable if only the newly built bone should be analysed even though this could lead to problems related to small sample quantity. It also remains to be seen to which extent varying turnover rates of different skeletal elements, especially in children, must be taken into account.

Neutrophils mediate blood-spinal cord barrier disruption in demyelinating neuroinflammatory diseases

Disruption of the blood-brain and blood-spinal cord barriers (BBB and BSCB, respectively) and immune cell infiltration are early pathophysiological hallmarks of multiple sclerosis (MS), its animal model experimental autoimmune encephalomyelitis (EAE), and neuromyelitis optica (NMO). However, their contribution to disease initiation and development remains unclear. In this study, we induced EAE in lys-eGFP-ki mice and performed single, nonterminal intravital imaging to investigate BSCB permeability simultaneously with the kinetics of GFP(+) myeloid cell infiltration. We observed a loss in BSCB integrity within a day of disease onset, which paralleled the infiltration of GFP(+) cells into the CNS and lasted for ∼4 d. Neutrophils accounted for a significant proportion of the circulating and CNS-infiltrating myeloid cells during the preclinical phase of EAE, and their depletion delayed the onset and reduced the severity of EAE while maintaining BSCB integrity. We also show that neutrophils collected from the blood or bone marrow of EAE mice transmigrate more efficiently than do neutrophils of naive animals in a BBB cell culture model. Moreover, using intravital videomicroscopy, we demonstrate that the IL-1R type 1 governs the firm adhesion of neutrophils to the inflamed spinal cord vasculature. Finally, immunostaining of postmortem CNS material obtained from an acutely ill multiple sclerosis patient and two neuromyelitis optica patients revealed instances of infiltrated neutrophils associated with regions of BBB or BSCB leakage. Taken together, our data provide evidence that neutrophils are involved in the initial events that take place during EAE and that they are intimately linked with the status of the BBB/BSCB.

Retinal differentiation of human bone marrow-derived stem cells by co-culture with retinal pigment epithelium in vitro.

The goal of this study was to assess the in vitro differentiation capacity of human bone marrow-derived stem cells (hBMSCs) along retinal lineages. Mononuclear cells (MNC) were isolated from bone marrow (BM) and mobilized peripheral blood (mPB) using Ficoll-Paque density gradient centrifugation, and were sorted by magnetic-activated cell sorting (MACS) for specific stem cell subsets (CD34(+)CD38(+)/CD34(+)CD38(-)). These cells were then co-cultured on human retinal pigment epithelial cells (hRPE) for 7 days. The expression of stem cell, neural and retina-specific markers was examined by immunostaining, and the gene expression profiles were assessed after FACS separation of the co-cultured hBMSCs by quantitative reverse transcription polymerase chain reaction (qRT-PCR). Furthermore, in vitro functionality of the differentiated cells was analyzed by quantifying phagocytosis of CY5-labeled photoreceptor outer segments (POS). After 7 days of co-culture, hBMSCs adopted an elongated epithelial-like morphology and expressed RPE-specific markers, such as RPE65 and bestrophin. In addition, these differentiated cells were able to phagocytose OS, one of the main characteristics of native RPE cells. Our data demonstrated that human CD34(+)CD38(-) hBMSC may differentiate towards an RPE-like cell type in vitro and could become a new type of autologous donor cell for regenerative therapy in retinal degenerative diseases.

Continuum damage interactions between tension and compression in osteonal bone

Skeletal diseases such as osteoporosis impose a severe socio-economic burden to ageing societies. Decreasing mechanical competence causes a rise in bone fracture incidence and mortality especially after the age of 65 y. The mechanisms of how bone damage is accumulated under different loading modes and its impact on bone strength are unclear. We hypothesise that damage accumulated in one loading mode increases the fracture risk in another. This study aimed at identifying continuum damage interactions between tensile and compressive loading modes. We propose and identify the material constants of a novel piecewise 1D constitutive model capable of describing the mechanical response of bone in combined tensile and compressive loading histories. We performed several sets of loading–reloading experiments to compute stiffness, plastic strains, and stress-strain curves. For tensile overloading, a stiffness reduction (damage) of 60% at 0.65% accumulated plastic strain was detectable as stiffness reduction of 20% under compression. For compressive overloading, 60% damage at 0.75% plastic strain was detectable as a stiffness reduction of 50% in tension. Plastic strain at ultimate stress was the same in tension and compression. Compression showed softening and tension exponential hardening in the post-yield regime. The hardening behaviour in compression is unaffected by a previous overload in tension but the hardening behaviour in tension is affected by a previous overload in compression as tensile reloading strength is significantly reduced. This paper demonstrates how damage accumulated under one loading mode affects the mechanical behaviour in another loading mode. To explain this and to illustrate a possible implementation we proposed a theoretical model. Including such loading mode dependent damage and plasticity behaviour in finite element models will help to improve fracture risk analysis of whole bones and bone implant structures.

Certolizumab treatment during late pregnancy in patients with rheumatic diseases: Low drug levels in cord blood but possible risk for maternal infections. A case series of 13 patients

OBJECTIVE Due to reduction of immune-suppressive drugs, patients with rheumatic diseases can experience an increase in disease activity during pregnancy. In such cases, TNF-inhibitors may be prescribed. However, monoclonal antibodies with the Fc moiety are actively transported across the placenta, resulting in therapeutic drug levels in the newborn. As certolizumab (CZP) lacks the Fc moiety, it may bear a lower risk for the child. METHOD We report a case series of thirteen patients (5 with rheumatoid arthritis and 8 with spondyloarthritis) treated with CZP during late pregnancy to control disease activity. RESULT CZP measured in cord blood of eleven infants ranged between undetectable levels and 1μg/mL whereas the median CZP level of maternal plasma was 32.97μg/mL. Three women developed an infection during the third trimester, of whom one had a severe infection and one had an infection that resulted in a pre-term delivery. During the postpartum period, 6 patients remained on CZP while breastfeeding. CZP levels in the breast milk of two breastfeeding patients were undetectable. CONCLUSION The lack of the active transplacental transfer of CZP gives the possibility to treat inflammatory arthritis during late gestation without potential harm to the newborn. However, in pregnant women treated with TNF-inhibitors and prednisone, attention should be given to the increased susceptibility to infections, which might cause prematurity. CZP treatment can be continued while breastfeeding.

Atypical variants of bovine spongiform encephalopathy: rare diseases with consequences for BSE surveillance and control

Occurring for the first time in 1986 in the United Kingdom, bovine spongiform encephalopathy (BSE), the so-called “mad-cow disease”, has had unprecedented consequences in veterinary public health. The implementation of drastic measures, including the ban of meat-and-bone-meal from livestock feed and the removal of specified risk materials from the food chain has eventually resulted in a significant decline of the epidemic. The disease was long thought to be caused by a single agent, but since the introduction of immunochemical diagnostic techniques, evidence of a phenotypic variation of BSE has emerged. Reviewing the literature available on the subject, this paper briefly summarizes the current knowledge about these atypical forms of BSE and discusses the consequences of their occurrence for disease control measures.

Systematic review of worksite-based behavior change programs that target metabolic syndrome risk factors

Approximately one-third of US adults have metabolic syndrome, the clustering of cardiovascular risk factors that include hypertension, abdominal adiposity, elevated fasting glucose, low high-density lipoprotein (HDL)-cholesterol and elevated triglyceride levels. While the definition of metabolic syndrome continues to be much debated among leading health research organizations, the fact is that individuals with metabolic syndrome have an increased risk of developing cardiovascular disease and/or type 2 diabetes. A recent report by the Henry J. Kaiser Family Foundation found that the US spent $2.2 trillion (16.2% of the Gross Domestic Product) on healthcare in 2007 and cited that among other factors, chronic diseases, including type 2 diabetes and cardiovascular disease, are large contributors to this growing national expenditure. Bearing a substantial portion of this cost are employers, the leading providers of health insurance. In lieu of this, many employers have begun implementing health promotion efforts to counteract these rising costs. However, evidence-based practices, uniform guidelines and policy do not exist for this setting in regard to the prevention of metabolic syndrome risk factors as defined by the National Cholesterol Education Program (NCEP) Adult Treatment Panel III (ATP III). Therefore, the aim of this review was to determine the effects of worksite-based behavior change programs on reducing the risk factors for metabolic syndrome in adults. Using relevant search terms, OVID MEDLINE was used to search the peer-reviewed literature published since 1998, resulting in 23 articles meeting the inclusion criteria for the review. The American Dietetic Association's Evidence Analysis Process was used to abstract data from selected articles, assess the quality of each study, compile the evidence, develop a summarized conclusion, and assign a grade based upon the strength of supporting evidence. The results revealed that participating in a worksite-based behavior change program may be associated in one or more improved metabolic syndrome risk factors. Programs that delivered a higher dose (>22 hours), in a shorter duration (<2 years) using two or more behavior-change strategies were associated with more metabolic risk factors being positively impacted. A Conclusion Grade of III was obtained for the evidence, indicating that studies were of weak design or results were inconclusive due to inadequate sample sizes, bias and lack of generalizability. These results provide some support for the continued use of worksite-based health promotion and further research is needed to determine if multi-strategy, intense behavior change programs targeting multiple risk factors are able to sustain health improvements in the long-term.^

Metabolic Regulation of mTOR Activation and Endoplasmic Reticulum Stress in the Heart

When subjected to increased workload, the heart responds metabolically by increasing its reliance on glucose and structurally by increasing the size of myocytes. Whether changes in metabolism regulate the structural remodeling process is unknown. A likely candidate for a link between metabolism and growth in the heart is the mammalian target of rapamycin (mTOR), which couples energy and nutrient metabolism to cell growth. Recently, sustained mTOR activation has also been implicated in the development of endoplasmic reticulum (ER) stress. We explored possible mechanisms by which acute metabolic changes in the hemodynamically stressed heart regulate mTOR activation, ER stress and cardiac function in the ex vivo isolated working rat heart. Doubling the heart’s workload acutely increased rates of glucose uptake beyond rates of glucose oxidation. The concomitant increase in glucose 6-phosphate (G6P) was associated with mTOR activation, endoplasmic reticulum (ER) stress and impaired contractile function. Both rapamycin and metformin restored glycolytic homeostasis, relieved ER stress and rescued contractile function. G6P and ER stress were also downregulated with mechanical unloading of failing human hearts. Taken together, the data support the hypothesis that metabolic remodeling precedes, triggers, and sustains structural remodeling of the heart and implicate a critical role for G6P in load-induced contractile dysfunction, mTOR activation and ER stress. In general terms, the intermediary metabolism of energy providing substrates provides signals for the onset and progression of hypertrophy and heart failure.