Lactoferrin Against SARS-CoV-2: In Vitro and In Silico Evidences.

Lactoferrin (Lf) is a cationic glycoprotein synthetized by exocrine glands and is present in all human secretions. It is also secreted by neutrophils in infection and inflammation sites. This glycoprotein possesses antimicrobial activity due to its capability to chelate two ferric ions per molecule, as well as to interact with bacterial and viral anionic surface components. The cationic features of Lf bind to cells, protecting the host from bacterial and viral injuries. Its anti-inflammatory activity is mediated by the ability to enter inside the nucleus of host cells, thus inhibiting the synthesis of proinflammatory cytokine genes. In particular, Lf down-regulates the synthesis of IL-6, which is involved in iron homeostasis disorders and leads to intracellular iron overload, favoring viral replication and infection. The well-known antiviral activity of Lf has been demonstrated against DNA, RNA, and enveloped and naked viruses and, therefore, Lf could be efficient in counteracting also SARS-CoV-2 infection. For this purpose, we performed in vitro assays, proving that Lf exerts an antiviral activity against SARS-COV-2 through direct attachment to both SARS-CoV-2 and cell surface components. This activity varied according to concentration (100/500 µg/ml), multiplicity of infection (0.1/0.01), and cell type (Vero E6/Caco-2 cells). Interestingly, the in silico results strongly supported the hypothesis of a direct recognition between Lf and the spike S glycoprotein, which can thus hinder viral entry into the cells. These in vitro observations led us to speculate a potential supplementary role of Lf in the management of COVID-19 patients.

Serum iPTH range in a reference population: From an integrated approach to vitamin D prevalence impact evaluation.

BACKGROUND: The iPTH upper reference limit (URL) reported by our laboratory provider (Abbott Laboratories) at Tor Vergata University Hospital was evaluated by internal verification procedures as not representative of our population and resulting as underestimated. In this study, a new reference interval has been investigated and established by comparing a direct and an indirect method based on a statistical reduction from results stored in the laboratory database. METHODS: For reference interval calculation from the healthy population, we analyzed a cohort of 100 blood donors (84% males and 16% females) screened with no bone-related and malabsorption diseases. We analyzed a cohort of 495 patients retrieved from more than 800 iPTH results by excluding subjects with pathological measurement for calcium, phosphorus, and creatinine for the reference interval evaluation. Patients with vitamin D results were included in the analysis. Vitamin D sufficiency status during the period from January to September 2020 was also evaluated by investigating 3,050 patients. RESULTS: The iPTH reference interval of a healthy blood donor population was measured as 25.2-109.1 pg/mL (2.7-11.6 pmol/L) at 2.5 and 97.5 distribution percentile. The iPTH reference interval from data stored in the laboratory database was 19.3-112.5 pg/mL (2.0-11.9 pmol/L). Furthermore, 60% of the whole population had prevalently insufficient vitamin D concentration (<30 ng/dL; <75 nmol/L). The impact of vitamin D concentration on the iPTH reference interval was measured for insufficient vitamin D (<30 ng/dL; <75 nmol/L) as 15.2-127.7 pg/mL (1.6-13.5 pmol/L), desirable vitamin D (30-40 ng/ml; 75-100 nmol/L) as 25.6-105 pg/mL (2.7-10.7 pmol/L) and optimal vitamin D (>40 ng/ml; >100 nmol/L) as 26.2-89.2 pg/mL (2.8-9.4 pmol/L), respectively. CONCLUSIONS: The URL reported in manufacturer datasheets likely refers to a normal population with non-pathological vitamin D levels. On the contrary, the considered population was mostly vitamin D insufficient, resulting in a URL shift. On this basis, we suggest describing in medical reports the iPTH range for vitamin D deficiency for diagnosis of primary hyperparathyroidism even when a specific vitamin D request is lacking. On the other hand, reporting optimal vitamin D-based iPTH reference interval could be clinically relevant in supplemented patients as a marker of treatment efficacy.

Dietary Flaxseed Mitigates Impaired Skeletal Muscle Regeneration: in Vivo, in Vitro and in Silico Studies.

BACKGROUND: Diets enriched with n-3 polyunsaturated fatty acids (n-3 PUFAs) have been shown to exert a positive impact on muscle diseases. Flaxseed is one of the richest sources of n-3 PUFA acid α-linolenic acid (ALA). The aim of this study was to assess the effects of flaxseed and ALA in models of skeletal muscle degeneration characterized by high levels of Tumor Necrosis Factor-α (TNF). METHODS: The in vivo studies were carried out on dystrophic hamsters affected by muscle damage associated with high TNF plasma levels and fed with a long-term 30% flaxseed-supplemented diet. Differentiating C2C12 myoblasts treated with TNF and challenged with ALA represented the in vitro model. Skeletal muscle morphology was scrutinized by applying the Principal Component Analysis statistical method. Apoptosis, inflammation and myogenesis were analyzed by immunofluorescence. Finally, an in silico analysis was carried out to predict the possible pathways underlying the effects of n-3 PUFAs. RESULTS: The flaxseed-enriched diet protected the dystrophic muscle from apoptosis and preserved muscle myogenesis by increasing the myogenin and alpha myosin heavy chain. Moreover, it restored the normal expression pattern of caveolin-3 thereby allowing protein retention at the sarcolemma. ALA reduced TNF-induced apoptosis in differentiating myoblasts and prevented the TNF-induced inhibition of myogenesis, as demonstrated by the increased expression of myogenin, myosin heavy chain and caveolin-3, while promoting myotube fusion. The in silico investigation revealed that FAK pathways may play a central role in the protective effects of ALA on myogenesis. CONCLUSIONS: These findings indicate that flaxseed may exert potent beneficial effects by preserving skeletal muscle regeneration and homeostasis partly through an ALA-mediated action. Thus, dietary flaxseed and ALA may serve as a useful strategy for treating patients with muscle dystrophies.

A diet supplemented with ALA-rich flaxseed prevents cardiomyocyte apoptosis by regulating caveolin-3 expression.

AIMS: n-3 polyunsaturated fatty acids (PUFAs) induce beneficial effects on the heart, but the mechanisms through which these effects are operated are not completely clarified yet. Among others, cardiac diseases are often associated with increased levels of cytokines, such as tumour necrosis factor-α (TNF), that cause degeneration and death of cardiomyocytes. The present study has been carried out to investigate (i) the potential anti-apoptotic effects induced by the n-3 polyunsaturated α-linolenic acid (ALA) in experimental models of cardiac diseases characterized by high levels of TNF, and (ii) the potential role of caveolin-3 (Cav-3) in the mechanisms involved in this process. METHODS AND RESULTS: An ALA-rich flaxseed diet, administered from weaning to hereditary cardiomyopathic hamsters, prevented the onset of myocardial apoptosis associated with high plasma and tissue levels of TNF preserving caveolin-3 expression. To confirm these findings, isolated neonatal mouse cardiomyocytes were exposed to TNF to induce apoptosis. ALA pre-treatment greatly enhanced Cav-3 expression hampering the internalization of the caveolar TNF receptor and, thus, determining the abortion of the apoptotic vs. survival cascade. CONCLUSION: This study unveiled the Cav-3 pivotal role in defending cardiomyocytes against the TNF pro-apoptotic action and the ALA capacity to regulate this mechanism preventing cardiac degenerative diseases.

An omega-3 fatty acid-enriched diet prevents skeletal muscle lesions in a hamster model of dystrophy.

Currently, despite well-known mutational causes, a universal treatment for neuromuscular disorders is still lacking, and current therapeutic efforts are mainly restricted to symptomatic treatments. In the present study, δ-sarcoglycan-null dystrophic hamsters were fed a diet enriched in flaxseed-derived ω3 α-linolenic fatty acid from weaning until death. α-linolenic fatty acid precluded the dystrophic degeneration of muscle morphology and function. In fact, in dystrophic animals fed flaxseed-derived α-linolenic fatty acid, the histological appearance of the muscular tissue was improved, the proliferation of interstitial cells was decreased, and the myogenic differentiation originated new myocytes to repair the injured muscle. In addition, muscle myofibers were larger and cell membrane integrity was preserved, as witnessed by the correct localization of α-, ß-, and γ-sarcoglycans and α-dystroglycan. Furthermore, the cytoplasmic accumulation of both ß-catenin and caveolin-3 was abolished in dystrophic hamster muscle fed α-linolenic fatty acid versus control animals fed standard diet, while α-myosin heavy chain was expressed at nearly physiological levels. These findings, obtained by dietary intervention only, introduce a novel concept that provides evidence that the modulation of the plasmalemma lipid profile could represent an efficacious strategy to ameliorate human muscular dystrophy.

Alpha-linolenic acid-enriched diet prevents myocardial damage and expands longevity in cardiomyopathic hamsters.

Randomized clinical trials have demonstrated that the increased intake of omega-3 polyunsaturated fatty acids significantly reduces the risk of ischemic cardiovascular disease, but no investigations have been performed in hereditary cardiomyopathies with diffusely damaged myocardium. In the present study, delta-sarcoglycan-null cardiomyopathic hamsters were fed from weaning to death with an alpha-linolenic acid (ALA)-enriched versus standard diet. Results demonstrated a great accumulation of ALA and eicosapentaenoic acid and an increased eicosapentaenoic/arachidonic acid ratio in cardiomyopathic hamster hearts, correlating with the preservation of myocardial structure and function. In fact, ALA administration preserved plasmalemma and mitochondrial membrane integrity, thus maintaining proper cell/extracellular matrix contacts and signaling, as well as a normal gene expression profile (myosin heavy chain isoforms, atrial natriuretic peptide, transforming growth factor-beta1) and a limited extension of fibrotic areas within ALA-fed cardiomyopathic hearts. Consequently, hemodynamic indexes were safeguarded, and more than 60% of ALA-fed animals were still alive (mean survival time, 293+/-141.8 days) when all those fed with standard diet were deceased (mean survival time, 175.9+/-56 days). Therefore, the clinically evident beneficial effects of omega-3 polyunsaturated fatty acids are mainly related to preservation of myocardium structure and function and the attenuation of myocardial fibrosis.