Lactoferrin Induces Erythropoietin Synthesis and Rescues Cognitive Functions in the Offspring of Rats Subjected to Prenatal Hypoxia.

The protective effects of recombinant human lactoferrin rhLF (branded "CAPRABEL™") on the cognitive functions of rat offspring subjected to prenatal hypoxia (7% O, 3 h, 14th day of gestation) have been analyzed. About 90% of rhLF in CAPRABEL was iron-free (apo-LF). Rat dams received several injections of 10 mg of CAPRABEL during either gestation (before and after the hypoxic attack) or lactation.

Lactoferrin modified by hypohalous acids: Partial loss in activation of human neutrophils.

Previously we have shown that lactoferrin (LTF), a protein of secondary neutrophilic granules, can be efficiently modified by hypohalous acids (HOCl and HOBr), which are produced at high concentrations during inflammation and oxidative/halogenative stress by myeloperoxidase, an enzyme of azurophilic neutrophilic granules. Here we compared the effects of recombinant human lactoferrin (rhLTF) and its halogenated derivatives (rhLTF-Cl and rhLTF-Br) on functional responses of neutrophils. Our results demonstrated that after halogenative modification, rhLTF lost its ability to induce mobilization of intracellular calcium, actin cytoskeleton reorganization, and morphological changes in human neutrophils.

Bovine lactoferrin and Lentinus edodes mycelia polysaccharide complex: The formation and the activity to protect islet β cells.

The formation of complexes may be used for the development of delivery systems in foods field. The aim of this study was to explore the interaction mechanism between Lentinus edodes mycelia polysaccharide (LMP) and bovine lactoferrin (BLF), and the activity of LMP-BLF complex to inhibit oxidative stress in islet β cells. The interaction mechanisms of LMP with BLF were investigated with multi-spectroscopic techniques.

Interaction of Lactoferrin with Unsaturated Fatty Acids: In Vitro and In Vivo Study of Human Lactoferrin/Oleic Acid Complex Cytotoxicity.

As shown recently, oleic acid (OA) in complex with lactoferrin (LF) causes the death of cancer cells, but no mechanism(s) of that toxicity have been disclosed. In this study, constitutive parameters of the antitumor effect of LF/OA complex were explored. Complex LF/OA was prepared by titrating recombinant human LF with OA.

Characterization of serotonin and N-acetylserotonin systems in the human epidermis and skin cells.

Tryptophan hydroxylase (TPH) activity was detected in cultured epidermal melanocytes and dermal fibroblasts with respective Km of 5.08 and 2.83 mM and Vmax of 80.

Effects of recombinant human lactoferrin on calcium signaling and functional responses of human neutrophils.

Lactoferrin is a non-heme iron-binding glycoprotein with multiple health-beneficial functions including antimicrobial, antioxidant, anticarcinogenic, and immunomodulatory effects. There is emerging evidence that neutrophils may serve as targets of lactoferrin in vivo, and here we show how recombinant human lactoferrin (rhLf) can contribute to this regulation. Indeed, our results demonstrate that rhLf binds efficiently to human neutrophils and induces a variety of early cellular responses such as mobilization of intracellular Ca, remodeling of actin cytoskeleton, and degranulation (release of lysozyme and myeloperoxidase).

Surface Enhanced Raman Spectroscopy of Lactoferrin Adsorbed on Silvered Porous Silicon Covered with Graphene.

We registered surface enhanced Raman scattering (SERS) spectra of the human lactoferrin molecules adsorbed on a silvered porous silicon (-Si) from 10⁻10 M solutions. It was found that the -Si template causes a negative surface potential of silver particles and their chemical resistivity to oxidation. These properties provided to attract positively charged lactoferrin molecules and prevent their interaction with metallic particles upon 473 nm laser excitation.

Melatonin, mitochondria, and the skin.

The skin being a protective barrier between external and internal (body) environments has the sensory and adaptive capacity to maintain local and global body homeostasis in response to noxious factors. An important part of the skin response to stress is its ability for melatonin synthesis and subsequent metabolism through the indolic and kynuric pathways. Indeed, melatonin and its metabolites have emerged as indispensable for physiological skin functions and for effective protection of a cutaneous homeostasis from hostile environmental factors.

Metabolism of melatonin in the skin: Why is it important?

Melatonin is produced in almost all living taxa and is probably 2-3 billion years old. Its pleiotropic activities are related to its local concentration that is secondary to its local synthesis, delivery from distant sites and metabolic or non-enzymatic consumption. This consumption generates metabolites through indolic, kynuric and cytochrome P450 (CYP) mediated hydroxylations and O-demethylation or non-enzymatic processes, with potentially diverse phenotypic effects.

Novel activities of CYP11A1 and their potential physiological significance.

CYP11A1, found only in vertebrates, catalyzes the first step of steroidogenesis where cholesterol is converted to pregnenolone. The purified enzyme, also converts desmosterol and plant sterols including campesterol and β-sitosterol, to pregnenolone. Studies, initially with purified enzyme, reveal that 7-dehydrocholesterol (7DHC), ergosterol, lumisterol 3, and vitamins D3 and D2 also serve as substrates for CYP11A1, with 7DHC being better and vitamins D3 and D2 being poorer substrates than cholesterol.

Local melatoninergic system as the protector of skin integrity.

The human skin is not only a target for the protective actions of melatonin, but also a site of melatonin synthesis and metabolism, suggesting an important role for a local melatoninergic system in protection against ultraviolet radiation (UVR) induced damages. While melatonin exerts many effects on cell physiology and tissue homeostasis via membrane bound melatonin receptors, the strong protective effects of melatonin against the UVR-induced skin damage including DNA repair/protection seen at its high (pharmocological) concentrations indicate that these are mainly mediated through receptor-independent mechanisms or perhaps through activation of putative melatonin nuclear receptors. The destructive effects of the UVR are significantly counteracted or modulated by melatonin in the context of a complex intracutaneous melatoninergic anti-oxidative system with UVR-enhanced or UVR-independent melatonin metabolites.

Novel vitamin D photoproducts and their precursors in the skin.

Novel metabolic pathways initiated by the enzymatic action of CYP11A1 on 7DHC (7-dehydrocholesterol), ergosterol, vitamins D3 and D2 were characterized with help of chemical synthesis, UV and mass spectrometry and NMR analyses. The first pathway follows the sequence 7DHC→22(OH)7DHC → 20,22(OH)27DHC → 7DHP (7-dehydropregnenolone), which can further be metabolized by steroidogenic enzymes. The resulting 5,7-dienes can be transformed by UVB to corresponding, biologically active, secosteroids.

In vivo production of novel vitamin D2 hydroxy-derivatives by human placentas, epidermal keratinocytes, Caco-2 colon cells and the adrenal gland.

We investigated the metabolism of vitamin D2 to hydroxyvitamin D2 metabolites ((OH)D2) by human placentas ex-utero, adrenal glands ex-vivo and cultured human epidermal keratinocytes and colonic Caco-2 cells, and identified 20(OH)D2, 17,20(OH)₂D2, 1,20(OH)₂D2, 25(OH)D2 and 1,25(OH)₂D2 as products. Inhibition of product formation by 22R-hydroxycholesterol indicated involvement of CYP11A1 in 20- and 17-hydroxylation of vitamin D2, while use of ketoconazole indicated involvement of CYP27B1 in 1α-hydroxylation of products. Studies with purified human CYP11A1 confirmed the ability of this enzyme to convert vitamin D2 to 20(OH)D2 and 17,20(OH)₂D2.

Cytochromes p450 and skin cancer: role of local endocrine pathways.

Skin is the largest body organ forming a metabolically active barrier between external and internal environments. The metabolic barrier is composed of cytochromes P450 (CYPs) that regulate its homeostasis through activation or inactivation of biologically relevant molecules. In this review we focus our attention on local steroidogenic and secosteroidogenic systems in relation to skin cancer, e.

In vivo evidence for a novel pathway of vitamin D₃ metabolism initiated by P450scc and modified by CYP27B1.

We define previously unrecognized in vivo pathways of vitamin D(3) (D3) metabolism generating novel D3-hydroxyderivatives different from 25-hydroxyvitamin D(3) [25(OH)D3] and 1,25(OH)(2)D3. Their novel products include 20-hydroxyvitamin D(3) [20(OH)D3], 22(OH)D3, 20,23(OH)(2)D3, 20,22(OH)(2)D3, 1,20(OH)(2)D3, 1,20,23(OH)(3)D3, and 17,20,23(OH)(3)D3 and were produced by placenta, adrenal glands, and epidermal keratinocytes. We detected the predominant metabolite [20(OH)D3] in human serum with a relative concentration ∼20 times lower than 25(OH)D3.

Sequential metabolism of 7-dehydrocholesterol to steroidal 5,7-dienes in adrenal glands and its biological implication in the skin.

Since P450scc transforms 7-dehydrocholesterol (7DHC) to 7-dehydropregnenolone (7DHP) in vitro, we investigated sequential 7DHC metabolism by adrenal glands ex vivo. There was a rapid, time- and dose-dependent metabolism of 7DHC by adrenals from rats, pigs, rabbits and dogs with production of more polar 5,7-dienes as detected by RP-HPLC. Based on retention time (RT), UV spectra and mass spectrometry, we identified the major products common to all tested species as 7DHP, 22-hydroxy-7DHC and 20,22-dihydroxy-7DHC.

Metabolism of melatonin by cytochrome P450s in rat liver mitochondria and microsomes.

In the present study we provide direct evidence for the involvement of rat microsomal cytochrome P450s in melatonin O-demethylation and hydroxylation at two different positions: 2 and 6, as well as generation of N(1)-acetyl-N(2)-formyl-5-methoxy-kynuramine (AFMK) and two unknown products. Moreover, we found that mitochondrial cytochrome P450s also converts melatonin into AFMK, N-acetylserotonin, 2-hydroxymelatonin, 6-hydroxymelatonin and the same two unknown products. Eadie-Hofstee plots for 6-hydroxylation and O-demethylation reactions were curvilinear for all tested fractions, suggestive of involvement of at least two components, one with a high affinity and low capacity, and another with a low affinity and high capacity.

An alternative pathway of vitamin D metabolism. Cytochrome P450scc (CYP11A1)-mediated conversion to 20-hydroxyvitamin D2 and 17,20-dihydroxyvitamin D2.

We report an alternative, hydroxylating pathway for the metabolism of vitamin D2 in a cytochrome P450 side chain cleavage (P450scc; CYP11A1) reconstituted system. NMR analyses identified solely 20-hydroxyvitamin D2 and 17,20-dihydroxyvitamin D2 derivatives. 20-Hydroxyvitamin D2 was produced at a rate of 0.

Constitutive and UV-induced metabolism of melatonin in keratinocytes and cell-free systems.

Melatonin, which can be produced in the skin, exerts a protective effect against damage induced by UV radiation (UVR). We have investigated the effect of UVB, the most damaging component of UVR, on melatonin metabolism in HaCaT keratinocytes and in a cell-free system. Four metabolites were identified by HPLC and LC-MS: 6-hydroxymelatonin, N1-acetyl-N2-formyl-5-methoxykynuramine (AFMK), 2-hydroxymelatonin (the main intermediate between melatonin and AFMK), and 4-hydroxymelatonin.

On the role of melatonin in skin physiology and pathology.

Melatonin has been experimentally implicated in skin functions such as hair growth cycling, fur pigmentation, and melanoma control, and melatonin receptors are expressed in several skin cells including normal and malignant keratinocytes, melanocytes, and fibroblasts. Melatonin is also able to suppress ultraviolet (UV)-induced damage to skin cells and shows strong antioxidant activity in UV exposed cells. Moreover, we recently uncovered expression in the skin of the biochemical machinery involved in the sequential transformation of l-tryptophan to serotonin and melatonin.

Enzymatic metabolism of ergosterol by cytochrome p450scc to biologically active 17alpha,24-dihydroxyergosterol.

We demonstrate the metabolism of ergosterol by cytochrome P450scc in either a reconstituted system or isolated adrenal mitochondria. The major reaction product was identified as 17alpha,24-dihydroxyergosterol. Purified P450scc also generated hydroxyergosterol as a minor product, which is probably an intermediate in the synthesis of 17alpha,24-dihydroxyergosterol.

The cytochrome P450scc system opens an alternate pathway of vitamin D3 metabolism.

We show that cytochrome P450scc (CYP11A1) in either a reconstituted system or in isolated adrenal mitochondria can metabolize vitamin D3. The major products of the reaction with reconstituted enzyme were 20-hydroxycholecalciferol and 20,22-dihydroxycholecalciferol, with yields of 16 and 4%, respectively, of the original vitamin D3 substrate. Trihydroxycholecalciferol was a minor product, likely arising from further metabolism of dihydroxycholecalciferol.

A novel metabolic pathway of melatonin: oxidation by cytochrome C.

The indoleamine melatonin is ubiquitously distributed, and because of its small size and amphiphilic nature, it is able to reach easily all cellular compartments. The highest intracellular melatonin concentrations are found in the mitochondria, suggestive of local metabolism and/or direct participation in organelle function. In mitochondria cytochrome c (cyt c) could represent a melatonin target since it has the capability to oxidize organic molecules in the presence of H2O2, and mitochondria are the main site of H2O2 production in nonphagocytic cells.

CRH stimulates POMC activity and corticosterone production in dermal fibroblasts.

It has been previously documented that human skin cells including epidermal keratinocytes and dermal fibroblasts produce and process proopiomelanocortin (POMC), corticotropin releasing hormone (CRH), and express functional CRH receptors type-1 (CRH-R1). The skin also has corticosteroidogenic activity, suggesting a functional connection between these elements. In the current study, we found that human dermal fibroblasts (but not normal epidermal keratinocytes) respond to CRH with stimulation of cAMP, with POMC gene and protein expression, and ACTH production and release.

CRH stimulation of corticosteroids production in melanocytes is mediated by ACTH.

The response to systemic stress is organized along the hypothalamic-pituitary-adrenal axis (HPA), whereas the response to a peripheral stress (solar radiation) is mediated by epidermal melanocytes (cells of neural crest origin) responsible for the pigmentary reaction. Melanocytes express proopiomelanocortin (POMC), corticotropin-releasing hormone (CRH), and CRH receptor-1 (CRH-R1) and can produce corticosterone. In the present study, incubation of normal epidermal melanocytes with CRH was found to trigger a functional cascade structured hierarchically and arranged along the same algorithm as in the HPA axis: CRH activation of CRH-R1 stimulated cAMP accumulation and increased POMC gene expression and production of ACTH.