Circulating sRANKL, Periostin, and Osteopontin as Biomarkers for the Assessment of Activated Osteoclastogenesis in Myeloma Related Bone Disease.

The hallmark of multiple myeloma is myeloma related bone disease. Interactions between myeloma plasma cells (MPCs), stromal cells, and the bone marrow (BM) microenvironment play a critical role in the pathogenesis of MBD. Bone remodeling is severely dysregulated with the prevalence of osteoclast activity. We aimed to assess circulating levels of sRANKL, periostin, and osteopontin as osteoclast activators in NDMM patients at diagnosis and in the course of treatment, correlations with clinical and laboratory data, and to evaluate their potential as additional biomarkers for the assessment of MBD. The current study involved 74 subjects (41 NDMM patients, 33 controls). MBD was assessed by whole-body low-dose computed tomography. sRANKL, periostin, and osteopontin were assayed by commercial ELISA kits. At diagnosis, all tested parameters were significantly higher in NDMM patients compared to the controls (p < 0.0001), correlating with disease stage, MBD grade, and BM infiltration by MPCs. During therapy, the serum levels of all tested proteins decrease, most prominently after autologous stem cell transplantation (p < 0.0001). A significant reduction was established in patients achieving complete and very-good partial response compared to all others (p < 0.05). In conclusion, sRANKL, periostin, and osteopontin reflect MBD severity and could be promising markers for MBD monitoring and the effect of myeloma treatment.

Dynamics of Bone Disease Biomarkers Dickkopf-1 and Sclerostin in Patients with Multiple Myeloma.

Dickkopf-1 (DKK-1) and sclerostin are essential Wnt/ß-catenin pathway inhibitors, playing an important role in multiple myeloma bone disease (MBD). We aimed to examine the serum DKK-1 and sclerostin variations in newly diagnosed multiple myeloma (NDMM) patients at diagnosis and in the course of therapy, including autologous stem cell transplantation (ASCT). This study included 41 NDMM-patients and 33 controls. MBD was assessed by whole-body low-dose computed tomography. DKK-1 and sclerostin were assayed by commercial ELISA kits. At diagnosis, NDMM-patients revealed significantly higher DKK-1 and sclerostin values (p < 0.0001), showing dependence on disease stage (lowest in ISS-I and highest in ISS-III: p < 0.0012 and p < 0.025, respectively, for both proteins). Bone lesions revealed significant positive correlation with both DKK-1 (p < 0.05) and sclerostin (p < 0.0001). In the course of therapy, significant reduction, more prominent after ASCT, was observed for both parameters in each treatment point compared to the baseline (p < 0.0001). Markedly lower sclerostin (p < 0.01) and DKK-1 (p < 0.05) values were observed in patients with complete and very good partial response compared to those with partial response, stable, or progressive disease. Sclerostin and DKK-1 in NDMM patients reflect the MBD severity and the effect of therapy. Both proteins could represent a novel tool for better disease monitoring and effectiveness of therapy.

Sambucus ebulus (Elderberry) Fruits Modulate Inflammation and Complement System Activity in Humans.

Sambucus ebulus (SE) fruits are used for immune stimulation and amelioration of gastrointestinal inflammatory conditions. Currently, there is no scientific evidence of their effects on various aspects of the immune response mechanisms in humans. The purpose of this study was to evaluate the immunomodulatory potential of SE fruit infusion intake in healthy humans. Anthocyanin content was determined with UPLC-ESI-MS/MS. Fifty-three volunteers enrolled in a 4-week SE infusion intake intervention. Blood count, serum total protein, Interleukin 1 beta (IL-1ß), Interleukin 6 (IL-6), Tumor Necrosis Factor Alpha (TNFα), High-sensitivity C-reactive protein (hs-CRP), C3, and C4 levels were measured on automatic analyzers, and Interleukin 8 (IL-8) was measured manually with an ELISA kit. Cyanidin-3-O-galactoside (48.15 mg/g DW), followed by cyaniding-3-sambubioside (43.41 ± 1.07 mg/g DW), were the most abundant anthocyanins in SE samples. A significant decrease in total protein (2.82%), IL-6 (20.15%), TNFα (5.38%), IL-8 (5.50%), C3 (4.16%), and C4 (14.29%) was established in the whole group. Total protein, IL-8, TNFα, and C4 decreased in women (3.11%, 4.76%, 5.09%, and 11.11%), and IL-6 decreased (40.61%) in men. Hb (1.20%) and hematocrit (1.55%) levels decreased in the whole group and in the women group (1.61% and 2.20%). SE fruits exert immune-modulatory activity as revealed by decreased pro-inflammatory status and complement activity markers in healthy volunteers after a 4-week intervention.

New Insights into the Research of Bioactive Compounds from Plant Origins with Nutraceutical and Pharmaceutical Potential.

Plant bioactive compounds are essential for human health due to their multiple biological effects, such as antioxidant, anticarcinogenic, antiallergenic, anti-inflammatory, antimutagenic, and antimicrobial activities, which can have beneficial effects on various noncommunicable diseases, such as autoimmune, inflammatory, cardiovascular, cancer, metabolic, and neurodegenerative diseases [...].

Comparative Phytochemical Analysis of Aronia melanocarpa L. Fruit Juices on Bulgarian Market.

Aronia melanocarpa L. (black chokeberry), belonging to the Rosaceae family, contains high amounts of polyphenolics and therefore exhibits one of the highest antioxidant and anti-inflammatory activities among berry fruits. Chokeberries are used in the food industry for juice, nectar, and wine production and as colorants. We aimed to compare the phytochemical composition of three chokeberry juices commercially available in the local market as sources of beneficial phytochemicals. Using GC-MS and LC-MS/MS, we performed the identification and quantitation of polar compounds and polyphenolics. The concentrations of 13 amino acids, including 6 essential amino acids, 10 organic acids, 20 sugar alcohols and derivatives, 14 saccharides, 12 fatty acids and esters, and 38 polyphenols, were estimated. One of the analyzed juices had the highest polyphenolic content (5273.87 ± 63.16 µg/mL), possibly due to 2.9 times higher anthocyanin concentration compared to anthocyanins in other tested juices. This study provides new data concerning phytochemical composition in terms of amino acids, organic acids, sugar acids, fatty acids and their esters, and polyphenols as phytocomponents of commercially available chokeberry juices. Results show that after all processing techniques and possibly different plant growth conditions, chokeberry juices are a valuable source of health-promoting phytochemicals such as phenolic acids, pro-anthocyanins, and anthocyanins, thus considering them as functional foods. We demonstrated a diversity of the active substances in bioactive foods marketed as "same"; therefore, the standardized therapeutic effect could be expected only by the utilization of food supplements with guaranteed constant content.

Circulating uncarboxylated matrix Gla protein in patients with atrial fibrillation or heart failure with preserved ejection fraction.

CONTEXT: Circulating uncarboxylated matrix Gla protein (ucMGP) is possibly related to coronary arterial calcification (CAC) in cardiovascular disease (CVD) patients. OBJECTIVE: We aimed to evaluate the relationships between circulating ucMGP, CVD pathology and CAC and its interplay with CVD risk factors. MATERIALS AND METHODS: ucMGP was measured in 99 CVD-patients. CAC score was determined by multislice computed tomography. Circulating ucMGP, uncarboxylated (ucOC) and carboxylated osteocalcin (cOC) were assayed by ELISA kits. Vitamin-K status was evaluated by ucOC/cOC ratio. RESULTS: A tendency for decreased ucMGP was observed for CAC ≥ 100 AU vs. CAC = 1-99 AU after exclusion of the patients on vitamin K-antagonist anticoagulants. Significant inverse correlations between ucMGP and vitamin-K status were indicated for the entire cohort and according to CAC score. Significant associations were found between ucMGP and risk factors for CVD. CONCLUSION: Circulating ucMGP may reflect certain stages of CVD and CAC. Future studies are needed to clarify its role as potential biomarker.

High resolution LC-MS/MS characterization of polyphenolic composition and evaluation of antioxidant activity of Sambucus ebulus fruit tea traditionally used in Bulgaria as a functional food.

The purpose of this study was to perform phytochemical analysis of tea from Sambucus ebulus fruits concerning hydroxycinnamic acids, flavonol glucosides, stilbenes and proanthocyanidin mono-, di- and trimers content. In total, 33 compounds were identified and quantified using UPLC-DAD-ESI/MS/MS system and the results are presented in mg/g dry weight (DW). Among analyzed hydroxycinnamic acids, 5-Caffeoylquinic acid (114.17 mg/g) was most abundant, followed by 3-p-Coumaroylquinic acid (50.33 mg/g) and 3-p-Feruloylquinic acid, p-Coumaric acid glucoside and 4-p-Coumaroylquinic acid (31.36 mg/g, 29.78 mg/g and 27.70 mg/g, respectively). Flavonol glucosides were represented predominantly by Quercetin-3-O-galactoside, Quercetin-3-O-rhamnosyl-galactoside Quercetin-3-O-glucoside and Quercetin-3-O-rhamnosyl-glucoside (3.68 mg/g, 3.22 mg/g, 2.87 mg/g and 2.56 mg/g, respectively). trans-Resveratrol-3-O-glucoside, epicatechin (40.62 mg/g) and proanthocyanidin di- and -trimers (19.90 mg/g - 31.42 mg/g) also were present in the tea. ABTS cation decolorization assay revealed 1.248 mM UAE activity and the percent of DPPH radical scavenging was 14.25%, corresponding to 39.07 µM Trolox equivalents.

Phytochemical Composition, Anti-Inflammatory and ER Stress-Reducing Potential of Sambucus ebulus L. Fruit Extract.

Sambucus ebulus L. (SE) fruits are used for their immunostimulation, hematopoietic and antiviral potential. Recently, we focused on analyzing the mechanism underlying SE fruit aqueous extract's (FAE) immunomodulation and anti-inflammatory activities, with attention to its endoplasmic reticulum (ER) stress-reducing potential. J774A.1 macrophages were treated with SE FAE alone or in conditions of lipopolysaccharides (LPS) stimulation. Using GC-MS and LC-MS/MS, its phytochemical composition was analyzed. To measure transcription and protein levels, we used qPCR and Western blot, respectively. The prevailing phytochemicals in SE FAE were hydroxycinnamic acids, proanthocyanidins and anthocyanins. The content of some amino acids, organic acids, alcohols, fatty acids and esters were newly reported. Extracts exerted an immunostimulation potential by stimulating IL-6, TNFα, Ccl2, COX2 and iNOS transcription, without inducing ER stress. SE FAE suppressed the LPS-induced transcription of inflammation related genes (IL-1ß, IL-6, TNFα, Ccl2, Icam-1, Fabp4, COX2, iNOS, Noxo1, IL-1ra, Sirt-1) and reduced the protein levels of iNOS, peIF2α, ATF6α and CHOP. The effects were comparable to that of salicylic acid. SE suppresses LPS-stimulated inflammatory markers on the transcription and translation levels. Targeting ER stress is possibly another mechanism underlying its anti-inflammatory potential. These findings reveal the potential of SE fruits as a beneficial therapeutic of inflammation and ER stress-related pathological conditions.

Effects of sulphur-containing mineral water intake on oxidative status and markers for inflammation in healthy subjects.

CONTEXT: Sulphurous mineral waters (SMW) have a wide range of applications. Sulphur content of mineral waters is considered as possible determinant for their anti-inflammatory or pro-inflammatory effects. OBJECTIVE: To explore the healing properties of Varna basin mineral water by analysing possible antioxidative and anti-inflammatory effects. MATERIALS AND METHODS: An intervention with Varna SMW intake was performed with healthy volunteers. Total thiols, total glutathione and its fractions, reactive oxygen metabolites, malondialdehyde, intracellular adhesion molecule (ICAM-1) and vascular cell adhesion molecule (VCAM-1) were measured. Expression of γ-gluthamyl-cysteinyl ligase (GCL) and sICAM-1 genes was also analysed. RESULTS: A significantly increased total glutathione and total thiols were observed at the end of the intervention. GCL and sICAM-1 gene expressions were increased after the intervention. CONCLUSION: SMW consumption improved redox status of the body. We suggested that these beneficial effects may be attributed to the established high levels of sulphur-containing compounds in Varna mineral water.

Adenosine Deaminase as a Useful Biomarker for Diagnosis and Monitoring of Inflammatory Bowel Disease.

BACKGROUND: Inflammatory bowel disease (IBD) is a multifactorial immunologically mediated disorder characterized by repeated cycles of exacerbations and remissions. Diagnosis and monitoring of ulcerative colitis (UC) and Crohn's disease (CD) as common types of IBD require the usage of unpleasant invasive methods such as colonoscopy and cross-sectional imaging. Development of a non-invasive biomarker panel including different inflammatory parameters for evaluation of various aspects of gut inflammation and disease course is a priority task. In addition to the well-known inflammatory markers serum C-reactive protein (CRP) and fecal calprotectin (FC), adenosine deaminase (ADA) could be a promising candidate-biomarker. ADA has been shown to increase in several inflammatory conditions, but little is known about its significance in IBD. This preliminary research aims to study the serum levels of ADA in IBD patients and to evaluate its ability to adequately reflect the gut inflammatory process. METHODS: Fifty-four IBD patients (40 with UC and 14 with CD) and 50 controls were enrolled in the study. Routine laboratory parameters such as white blood cells (WBC) count, erythrocyte sedimentation rate (ESR), and CRP were used. The specific biomarker for intestinal inflammation FC was measured by sandwich immunoassay (BÜHLMANN) and ADA activity - by two-step enzyme method (BioSystems). RESULTS: The median [IQR: 25th - 75th percentile] ADA values in IBD patients were significantly higher than those in the controls (18.7 U/L [15.4 - 22.5] vs. 9.10 U/L [7.1 - 11.2] respectively; p < 0.0001). A significant difference was obtained when comparing median ADA values in patients with active disease (20.4 [17.8 - 25.3] U/L) with those in patients with mild form of the disease or in remission (15.3 [13.0 - 16.0]; p < 0.0001). A strong positive correlation between ADA and FC (r = 0.63; p < 0.0001) and moderate positive correlation between ADA and CRP (r = 0.46; p < 0.001) were observed. ROC-curve analysis revealed good ability of ADA to discriminate not only IBD patients from healthy individuals, but also patients with active disease and those in remission/mild form. CONCLUSIONS: The present study revealed that ADA levels were significantly increased in IBD patients. Together with FC and CRP, ADA could be used as an effective biomarker for assessment of intestinal inflammation and as a potential indicator for disease activity.

Metabolomic analysis in ophthalmology.

Modern science takes into account phenotype complexity and establishes approaches to track changes on every possible level. Many "omics" studies have been developed over the last decade. Metabolomic analysis enables dynamic measurement of the metabolic response of a living system to a variety of stimuli or genetic modifications. Important targets of metabolomics is biomarker development and translation to the clinic for personalized diagnosis and a greater understanding of disease pathogenesis. The current review highlights the major aspects of metabolomic analysis and its applications for the identification of relevant predictive, diagnostic and prognostic biomarkers for some ocular diseases including dry eye, keratoconus, retinal diseases, macular degeneration, and glaucoma. To date, possible biomarker candidates for dry eye disease are lipid metabolites and androgens, for keratoconus cytokeratins, urea, citrate cycle, and oxidative stress metabolites. Palmitoylcarnitine, sphingolipids, vitamin D related metabolites, and steroid precursors may be used for distinguishing glaucoma patients from healthy controls. Dysregulation of amino acid and carnitine metabolism is critical in the development and progression of diabetic retinopathy. Further work is needed to discover and validate metabolic biomarkers as a powerful tool for understanding the molecular mechanisms of ocular diseases, to provide knowledge on their etiology and pathophysiology and opportunities for personalized clinical intervention at an early stage.

Serum 25-hydroxy vitamin D levels in Bulgarian patients with prostate cancer: a pilot study.

BACKGROUND: The antiproliferative effect of the active form of vitamin D on cancer cells and its ability to induce cell differentiation and suppression of tumor-induced angiogenesis in the last decade has provoked enormous research for the elucidation of its role in the prevention of different types of cancer and in slowing down the malignancy progression. The aim of the present pilot study was to determine the circulating 25-hydroxy vitamin D (25OHD) levels in Bulgarian prostate cancer (PCa) patients and to investigate their relationship with various determinants associated with the severity and progression of the disease. METHODS: A total of 53 male patients (mean age 67.0 ± 7.1 years) with clinical suspicion for PCa were enrolled in the study. All patients were subjected to systemic transrectal ultrasound-guided tru-cut prostate biopsies (10 cores at least). Detected tumors were graded using the Gleason grading system. Prostate specific antigen (PSA) serum levels were measured immunochemically. The 25OHD assay was performed by a validated HPLC-UV method. Other covariates (BMI, age, family history of PCa) were collected by interview at the time of hospitalization. One-way ANOVA with Kruskal Wallis statistics was used for comparison of medians of different parameters. The level of significance was set at p < 0.05. RESULTS: Significantly lower 25OHD levels were detected in PCa patients compared to those with benign prostate hyperplasia (BPH) (p < 0.05). Patients with high grade tumors (Gleason score ≥ 7) showed significantly lower 25OHD levels, while those with low grade tumors (Gleason score < 7) revealed better 25OHD status (50.49 vs. 63.17 nmol/L, p < 0.05). A moderate negative correlation between 25OHD levels and the Gleason score was established (Spearman r = -0.46, p < 0.05). Significant seasonal variations in 25OHD levels, both for PCa and BPH patients, were detected (p < 0.01). CONCLUSIONS: This preliminary study shows an association between 25OHD status and classical markers characterizing the severity of PCa. The results might suggest a potential beneficial role of vitamin D for PCa patients. Further prospective studies are needed to strengthen the interrelationships between 25OHD levels and variables related with PCa and to test them for causality.

Impaired mitochondrial Ca2+ homeostasis in respiratory chain-deficient cells but efficient compensation of energetic disadvantage by enhanced anaerobic glycolysis due to low ATP steady state levels.

Energy-producing pathways, adenine nucleotide levels, oxidative stress response and Ca(2+) homeostasis were investigated in cybrid cells incorporating two pathogenic mitochondrial DNA point mutations, 3243A>G and 3302A>G in tRNA(Leu(UUR)), as well as Rho(0) cells and compared to their parental 143B osteosarcoma cell line. All cells suffering from a severe respiratory chain deficiency were able to proliferate as fast as controls. The major defect in oxidative phosphorylation was efficiently compensated by a rise in anaerobic glycolysis, so that the total ATP production rate was preserved. This enhancement of glycolysis was enabled by a considerable decrease of cellular total adenine nucleotide pools and a concomitant shift in the AMP+ADP/ATP ratios, while the energy charge potential was still in the normal range. Further important consequences were an increased production of superoxide which, however, was neither escorted by major changes in the antioxidative defence systems nor was it leading to substantial oxidative damage. Most interestingly, the lowered mitochondrial membrane potential led to a disturbed intramitochondrial calcium homeostasis, which most likely is a major pathomechanism in mitochondrial diseases.

Correlation between the in vitro antioxidant activity and polyphenol content of aqueous extracts from Bulgarian herbs.

The water phase antioxidant activity of extracts from 23 Bulgarian medicinal plants was studied in relation to their polyphenol content in comparison with mate, black tea, honeybush and rooibos foreign species. Antioxidant activity was measured by the ABTS (2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid)) cation radical decolorization assay, and the total polyphenol content was assayed according to the Folin-Ciocalteu method. Five Bulgarian plant extracts exhibited higher antioxidant activity than that of mate, which is 21.7% of all Bulgarian herbs included in this study. These were Alchemilla vulgaris L. (4.79 +/- 0.14 mm), Sambucus ebulus L. (4.03 +/- 0.07 mm), Mentha spicata L. (3.90 +/- 0.03 mm), Fragaria vesca L. (3.74 +/- 0.06 mm), Crataegus monogyna Jacq. (3.63 +/- 0.05 mm). Another eight Bulgarian medicinal plant extracts exhibited an intermediate antioxidant activity - lower than that of mate and higher than that of honeybush, which makes 34.8% of all Bulgarian herbs included in the study. More than half of the herbal extracts included in the present study exhibited antioxidant activity higher than or comparable to the reference foreign plants. A positive correlation (r = 0.92) between antioxidant activity and polyphenol content was found, suggesting that the antioxidant capacity of the aqueous plant extracts is due to a great extent to their polyphenols.