Immunomodulatory role of vitamin D and selenium supplementation in newly diagnosed Graves' disease patients during methimazole treatment.

Introduction: Methimazole (MMI) represents the conventional therapeutic agent for Graves' disease (GD) hyperthyroidism, but MMI efficacy is limited since it marginally affects the underlying autoimmune process. In a previous study, we randomly assigned 42 newly diagnosed GD patients with insufficient vitamin D (VitD) and selenium (Se) levels to treatment with MMI alone (standard) or combined with selenomethionine and cholecalciferol (intervention) and observed a prompter resolution of hyperthyroidism in the intervention group. Methods: In the present study, we aimed to explore changes in peripheral T regulatory (Treg) and circulating natural killer (NK) cell frequency, circulating NK cell subset distribution and function, during treatment. Results: At baseline, circulating total CD3-CD56+NK cells and CD56bright NK cells were significantly higher in GD patients than in healthy controls (HC) (15.7 ± 9.6% vs 9.9 ± 5.6%, p=0.001; 12.2 ± 10.3% vs 7.3 ± 4.1%, p=0.02, respectively); no differences emerged in Treg cell frequency. Frequencies of total NK cells and CD56bright NK cells expressing the activation marker CD69 were significantly higher in GD patients than in HC, while total NK cells and CD56dim NK cells expressing CD161 (inhibitory receptor) were significantly lower. When co-cultured with the K562 target cell, NK cells from GD patients had a significantly lower degranulation ability compared to HC (p<0.001). Following 6 months of treatment, NK cells decreased in both the intervention and MMI-alone groups, but significantly more in the intervention group (total NK: -10.3%, CI 95% -15.8; -4.8% vs -3.6%, CI 95% -9; 1.8%, p=0.09 and CD56bright NK cells: -6.5%, CI 95% -10.1; -3 vs -0.9%, CI 95% -4.4; 2%, p=0.03). Compared to baseline, CD69+ NK cells significantly decreased, while degranulation ability slightly improved, although no differences emerged between the two treatment groups. Compared to baseline, Treg cell frequency increased exclusively in the intervention group (+1.1%, CI 95% 0.4; 1.7%). Discussion: This pilot study suggested that VitD and Se supplementation, in GD patients receiving MMI treatment, modulates Treg and NK cell frequency, favoring a more pronounced reduction of NK cells and the increase of Treg cells, compared to MMI alone. Even if further studies are needed, it is possible to speculate that this immunomodulatory action might have facilitated the prompter and better control of hyperthyroidism in the supplemented group observed in the previous study.

Add-On Effect of Selenium and Vitamin D Combined Supplementation in Early Control of Graves' Disease Hyperthyroidism During Methimazole Treatment.

Prompt and stable control of hyperthyroidism is fundamental to avoid the detrimental effects of thyroid hormone excess, and antithyroid drugs, mainly methimazole (MMI), represent the first-line treatment for Graves' disease (GD) hyperthyroidism. Decreased serum concentrations of selenium (Se) and calcifediol (25(OH)D, VitD) have been reported in newly diagnosed GD patients in observational studies. Low Se levels might exacerbate oxidative stress by compromising the antioxidant machinery's response to reactive oxygen species, and low VitD levels might hamper the anti-inflammatory immune response. We performed a randomized controlled clinical trial (EudraCT 2017-00505011) to investigate whether Se and cholecalciferol (VitD) addition to MMI is associated with a prompter control of hyperthyroidism. Forty-two consecutive patients with newly-onset GD and marginal/insufficient Se and VitD levels were randomly assigned to treatment with either MMI monotherapy or MMI combined with Se and VitD. Se treatment was withdrawn after 180 days, while the other treatments were continued. Combination therapy resulted in a significantly greater reduction in serum FT4 concentration at 45 days (-37.9 pg/ml, CI 95%, -43.7 to -32.2 pg/ml) and 180 days (-36.5 pg/ml, CI 95%, -42 to -30.9 pg/ml) compared to MMI monotherapy (respectively: -25.7 pg/ml, CI 95%, -31.6 to -19.7 pg/ml and -22.9 pg/ml, CI 95%, -28 to -17.3 pg/ml, p 0.002). Data at 270 days confirmed this trend (-37.8 pg/ml, CI 95%, -43.6 to -32.1 pg/ml vs -24.4 pg/ml, CI 95%, -30.3 to -18.4 pg/ml). The quality of life (QoL) score was investigated by the validated "Thyroid-related Patient-Reported Outcome" questionnaire (ThyPRO). ThyPRO composite score showed a greater improvement in the intervention group at 45 days (-14.6, CI 95%, -18.8 to -10.4), 180 (-9, CI 95%, -13.9 to -4.2) and 270 days (-14.3, CI 95%, -19.5 to -9.1) compared to MMI group (respectively, -5.2, CI 95%, -9.5 to -1; -5.4, CI 95%, -10.6 to -0.2 and -3.5, CI 95%, -9 to -2.1, p 0-6 months and 6-9 months <0.05). Our results suggest that reaching optimal Se and VitD levels increases the early efficacy of MMI treatment when Se and VitD levels are suboptimal.

In Vitro Evaluation of Antioxidant Potential of the Invasive Seagrass Halophila stipulacea.

Marine organisms with fast growth rates and great biological adaptive capacity might have biotechnological interests, since ecological competitiveness might rely on enhanced physiological or biochemical processes' capability promoting protection, defense, or repair intracellular damages. The invasive seagrass Halophila stipulacea, a non-indigenous species widespread in the Mediterranean Sea, belongs to this category. This is the premise to investigate the biotechnological interest of this species. In this study, we investigated the antioxidant activity in vitro, both in scavenging reactive oxygen species and in repairing damages from oxidative stress on the fibroblast human cell line WI-38. Together with the biochemical analysis, the antioxidant activity was characterized by the study of the expression of oxidative stress gene in WI-38 cells in presence or absence of the H. stipulacea extract. Concomitantly, the pigment pool of the extracts, as well as their macromolecular composition was characterized. This study was done separately on mature and young leaves. Results indicated that mature leaves exerted a great activity in scavenging reactive oxygen species and repairing damages from oxidative stress in the WI-38 cell line. This activity was paralleled to an enhanced carotenoids content in the mature leaf extracts and a higher carbohydrate contribution to organic matter. Our results suggest a potential of the old leaves of H. stipulacea as oxidative stress damage protecting or repair agents in fibroblast cell lines. This study paves the way to transmute the invasive H. stipulacea environmental threat in goods for human health.

Acetyl-L-Carnitine downregulates invasion (CXCR4/CXCL12, MMP-9) and angiogenesis (VEGF, CXCL8) pathways in prostate cancer cells: rationale for prevention and interception strategies.

BACKGROUND: Prostate cancer (PCa) is a leading cause of cancer-related death in males worldwide. Exacerbated inflammation and angiogenesis have been largely demonstrated to contribute to PCa progression. Diverse naturally occurring compounds and dietary supplements are endowed with anti-oxidant, anti-inflammatory and anti-angiogenic activities, representing valid compounds to target the aberrant cytokine/chemokine production governing PCa progression and angiogenesis, in a chemopreventive setting. Using mass spectrometry analysis on serum samples of prostate cancer patients, we have previously found higher levels of carnitines in non-cancer individuals, suggesting a protective role. Here we investigated the ability of Acetyl-L-carnitine (ALCAR) to interfere with key functional properties of prostate cancer progression and angiogenesis in vitro and in vivo and identified target molecules modulated by ALCAR. METHODS: The chemopreventive/angiopreventive activities ALCAR were investigated in vitro on four different prostate cancer (PCa) cell lines (PC-3, DU-145, LNCaP, 22Rv1) and a benign prostatic hyperplasia (BPH) cell line. The effects of ALCAR on the induction of apoptosis and cell cycle arrest were investigated by flow cytometry (FC). Functional analysis of cell adhesion, migration and invasion (Boyden chambers) were performed. ALCAR modulation of surface antigen receptor (chemokines) and intracellular cytokine production was assessed by FC. The release of pro-angiogenic factors was detected by a multiplex immunoassay. The effects of ALCAR on PCa cell growth in vivo was investigated using tumour xenografts. RESULTS: We found that ALCAR reduces cell proliferation, induces apoptosis, hinders the production of pro inflammatory cytokines (TNF-α and IFN-γ) and of chemokines CCL2, CXCL12 and receptor CXCR4 involved in the chemotactic axis and impairs the adhesion, migration and invasion capabilities of PCa and BPH cells in vitro. ALCAR exerts angiopreventive activities on PCa by reducing production/release of pro angiogenic factors (VEGF, CXCL8, CCL2, angiogenin) and metalloprotease MMP-9. Exposure of endothelial cells to conditioned media from PCa cells, pre-treated with ALCAR, inhibited the expression of CXCR4, CXCR1, CXCR2 and CCR2 compared to those from untreated cells. Oral administration (drinking water) of ALCAR to mice xenografted with two different PCa cell lines, resulted in reduced tumour cell growth in vivo. CONCLUSIONS: Our results highlight the capability of ALCAR to down-modulate growth, adhesion, migration and invasion of prostate cancer cells, by reducing the production of several crucial chemokines, cytokines and MMP9. ALCAR is a widely diffused dietary supplements and our findings provide a rational for studying ALCAR as a possible molecule for chemoprevention approaches in subjects at high risk to develop prostate cancer. We propose ALCAR as a new possible "repurposed agent' for cancer prevention and interception, similar to aspirin, metformin or beta-blockers.

Microalgal Derivatives as Potential Nutraceutical and Food Supplements for Human Health: A Focus on Cancer Prevention and Interception.

Epidemiological studies are providing strong evidence on beneficial health effects from dietary measures, leading scientists to actively investigate which foods and which specific agents in the diet can prevent diseases. Public health officers and medical experts should collaborate toward the design of disease prevention diets for nutritional intervention. Functional foods are emerging as an instrument for dietary intervention in disease prevention. Functional food products are technologically developed ingredients with specific health benefits. Among promising sources of functional foods and chemopreventive diets of interest, microalgae are gaining worldwide attention, based on their richness in high-value products, including carotenoids, proteins, vitamins, essential amino acids, omega-rich oils and, in general, anti-inflammatory and antioxidant compounds. Beneficial effects of microalgae on human health and/or wellness could in the future be useful in preventing or delaying the onset of cancer and cardiovascular diseases. During the past decades, microalgal biomass was predominately used in the health food market, with more than 75% of the annual microalgal biomass production being employed for the manufacture of powders, tablets, capsules or pastilles. In this review, we report and discuss the present and future role of microalgae as marine sources of functional foods/beverages for human wellbeing, focusing on perspectives in chemoprevention. We dissected this topic by analyzing the different classes of microalgal compounds with health outputs (based on their potential chemoprevention activities), the biodiversity of microalgal species and how to improve their cultivation, exploring the perspective of sustainable food from the sea.

Downregulation of Pro-Inflammatory and Pro-Angiogenic Pathways in Prostate Cancer Cells by a Polyphenol-Rich Extract from Olive Mill Wastewater.

Dietary phytochemicals are particularly attractive for chemoprevention and are able to modulate several signal transduction pathways linked with cancer. Olive oil, a major component of the Mediterranean diet, is an abundant source of phenolic compounds. Olive oil production is associated with the generation of a waste material, termed 'olive mill wastewater' (OMWW) that have been reported to contain water-soluble polyphenols. Prostate cancer (PCa) is considered as an ideal cancer type for chemopreventive approaches, due to its wide incidence but relatively long latency period and progression time. Here, we investigated activities associated with potential preventive properties of a polyphenol-rich olive mill wastewater extract, OMWW (A009), on three in vitro models of PCa. A009 was able to inhibit PCa cell proliferation, adhesion, migration, and invasion. Molecularly, we found that A009 targeted NF-κB and reduced pro-angiogenic growth factor, VEGF, CXCL8, and CXCL12 production. IL-6/STAT3 axis was also regulated by the extract. A009 shows promising properties, and purified hydroxytyrosol (HyT), the major polyphenol component of A009, was also active but not always as effective as A009. Finally, our results support the idea of repositioning a food waste-derived material for nutraceutical employment, with environmental and industrial cost management benefits.

Nutraceuticals and "Repurposed" Drugs of Phytochemical Origin in Prevention and Interception of Chronic Degenerative Diseases and Cancer.

BACKGROUND: Chronic, degenerative diseases are often characterized by inflammation and aberrant angiogenesis. For these pathologies, including rheumatoid arthritis, cardiovascular and autoimmune diseases, cancer, diabetes, and obesity, current therapies have limited efficacy. OBJECTIVES: The validation of novel (chemo)preventive and interceptive approaches, and the use of new or repurposed agents, alone or in combination with registered drugs, are urgently required. RESULTS: Phytochemicals (triterpenoids, flavonoids, retinoids) and their derivatives, nonsteroidal anti-inflammatory drugs (aspirin) as well as biguanides (metformin and phenformin) originally developed from phytochemical backbones, are multi-target agents showing antiangiogenic and anti-anti-inflammatory proprieties. Many of them target AMPK and metabolic pathways such as the mTOR axis. We summarize the beneficial effects of several compounds in conferring protection and supporting therapy, and as a paradigm, we present data on terpenoids & biquanides on beer hop xanthohumol and hydroxytryrosol from olive mill waste waters. CONCLUSIONS: These molecules could be employed for combinatorial chemoprevention and interception approaches or chemoprevention/therapy regimens for cancer and other chronic complex diseases.

Acetyl-l-carnitine is an anti-angiogenic agent targeting the VEGFR2 and CXCR4 pathways.

Carnitines play an important role in the energy exchange in cells, and are involved in the transport of fatty acids across the inner mitochondrial membrane. l-Acetylcarnitine (ALCAR) is an acetic acid ester of carnitine that has higher bioavailability and is considered a fat-burning energizer supplement. We previously found that in serum samples from prostate cancer (PCa) patients, 3 carnitine family members were significantly decreased, suggesting a potential protective role of carnitine against PCa. Several studies support beneficial effects of carnitines on cancer, no study has investigated the activities of carnitine on tumor angiogenesis. We examined whether ALCAR acts as an "angiopreventive" compound and studied the molecular mechanisms involved. We found that ALCAR was able to limit inflammatory angiogenesis by reducing stimulated endothelial cell and macrophage infiltration in vitro and in vivo. Molecularly, we show that ALCAR downregulates VEGF, VEGFR2, CXCL12, CXCR4 and FAK pathways. ALCAR blocked the activation of NF-κB and ICAM-1 and reduced the adhesion of a monocyte cell line to endothelial cells. This is the first study showing that ALCAR has anti-angiogenic and anti-inflammatory properties and might be an attractive candidate for cancer angioprevention.

Drink your prevention: beverages with cancer preventive phytochemicals.

Specific alimentary habits, including oriental and Mediterranean diets characterized by high consumption of vegetables, fruits, cereals and, for the Mediterranean diet, olive oil, are associated with a reduction of risk of cardiovascular diseases, type 2 diabetes, neurodegenerative diseases, and some cancers. Numerous beverages contain diverse natural compounds, termed phytochemicals, that have been reported to exert antitumor, antiangiogenic, and antioxidant properties. Here we review the chemopreventive and angiopreventive properties of selected phytochemicals found in common beverages: epigallocatechin(green tea), triterpenoids (citrus juices), resveratrol (red wine), xanthohumol (beer), procyanidin (chocolate), and caffeine (coffee), focusing on their molecular mechanisms, providing "ready to drink" prevention approaches.