Cellular and molecular effects of Baccharis dracunculifolia D.C. and Plectranthus barbatus Andrews medicinal plant extracts on retinoid metabolism in the human hepatic stellate cell LX-2.

BACKGROUND: Chronic hepatic diseases are serious problems worldwide, which may lead to the development of fibrosis and eventually cirrhosis. Despite the significant number of people affected by hepatic fibrosis, no effective treatment is available. In the liver, hepatic stellate cells are the major fibrogenic cell type that play a relevant function in chronic liver diseases. Thus, the characterization of components that control the fibrogenesis in the hepatic stellate cells is relevant in supporting the development of innovative therapies to treat and/or control liver fibrosis. The present study investigated the effects of Baccharis dracunculifolia D.C. and Plectranthus barbatus Andrews medicinal plant extracts in LX-2 transdifferentiation. METHODS: LX-2 is a human immortalized hepatic stellate cell that can transdifferentiate in vitro from a quiescent-like phenotype to a more proliferative and activated behavior, and it provides a useful platform to assess antifibrotic drugs. Then, the antifibrotic effects of hydroalcoholic extracts of Baccharis dracunculifolia and Plectranthus barbatus medicinal plants on LX-2 were evaluated. RESULTS: The results in our cellular analyses, under the investigated concentrations of the plant extracts, indicate no deleterious effects on LX-2 metabolism, such as toxicity, genotoxicity, or apoptosis. Moreover, the extracts induced changes in actin filament distribution of activated LX-2, despite not affecting the cellular markers of transdifferentiation. Consistent effects in cellular retinoid metabolism were observed, supporting the presumed activity of the plant extracts in hepatic lipids metabolism, which corroborated the traditional knowledge about their uses for liver dysfunction. CONCLUSION: The combined results suggested a potential hepatoprotective effect of the investigated plant extracts reinforcing their safe use as coadjuvants in treating imbalanced liver lipid metabolism.

Toxicity of blue led light and A2E is associated to mitochondrial dynamics impairment in ARPE-19 cells: implications for age-related macular degeneration.

Age-related macular degeneration (AMD) is a multifactorial retinal disease characterized by a progressive loss of central vision. Retinal pigment epithelium (RPE) degeneration is a critical event in AMD. It has been associated to A2E accumulation, which sensitizes RPE to blue light photodamage. Mitochondrial quality control mechanisms have evolved to ensure mitochondrial integrity and preserve cellular homeostasis. Particularly, mitochondrial dynamics involve the regulation of mitochondrial fission and fusion to preserve a healthy mitochondrial network. The present study aims to clarify the cellular and molecular mechanisms underlying photodamage-induced RPE cell death with particular focus on the involvement of defective mitochondrial dynamics. Light-emitting diodes irradiation (445 ± 18 nm; 4.43 mW/cm2) significantly reduced the viability of both unloaded and A2E-loaded human ARPE-19 cells and increased reactive oxygen species production. A2E along with blue light, triggered apoptosis measured by MC540/PI-flow cytometry and activated caspase-3. Blue light induced mitochondrial fusion/fission imbalance towards mitochondrial fragmentation in both non-loaded and A2E-loaded cells which correlated with the deregulation of mitochondria-shaping proteins level (OPA1, DRP1 and OMA1). To our knowledge, this is the first work reporting that photodamage causes mitochondrial dynamics deregulation in RPE cells. This process could possibly contribute to AMD pathology. Our findings suggest that the regulation of mitochondrial dynamics may be a valuable strategy for treating retinal degeneration diseases, such as AMD.

The Visual Cycle in the Inner Retina of Chicken and the Involvement of Retinal G-Protein-Coupled Receptor (RGR).

The vertebrate retina contains typical photoreceptor (PR) cones and rods responsible for day/night vision, respectively, and intrinsically photosensitive retinal ganglion cells (ipRGCs) involved in the regulation of non-image-forming tasks. Rhodopsin/cone opsin photopigments in visual PRs or melanopsin (Opn4) in ipRGCs utilizes retinaldehyde as a chromophore. The retinoid regeneration process denominated as "visual cycle" involves the retinal pigment epithelium (RPE) or Müller glial cells. Opn4, on the contrary, has been characterized as a bi/tristable photopigment, in which a photon of one wavelength isomerizes 11-cis to all-trans retinal (Ral), with a second photon re-isomerizing it back. However, it is unknown how the chromophore is further metabolized in the inner retina. Nor is it yet clear whether an alternative secondary cycle occurs involving players such as the retinal G-protein-coupled receptor (RGR), a putative photoisomerase of unidentified inner retinal activity. Here, we investigated the role of RGR in retinoid photoisomerization in Opn4x (Xenopus ortholog) (+) RGC primary cultures free of RPE and other cells from chicken embryonic retinas. Opn4x (+) RGCs display significant photic responses by calcium fluorescent imaging and photoisomerize exogenous all-trans to 11-cis Ral and other retinoids. RGR was found to be expressed in developing retina and in primary cultures; when its expression was knocked down, the levels of 11-cis, all-trans Ral, and all-trans retinol in cultures exposed to light were significantly higher and those in all-trans retinyl esters lower than in dark controls. The results support a novel role for RGR in ipRGCs to modulate retinaldehyde levels in light, keeping the balance of inner retinal retinoid pools.

Putrescine treatment reverses α-tocopherol-induced desynchronization of polyamine and retinoid metabolism during rat liver regeneration.

BACKGROUND: The pre-treatment with α-tocopherol inhibits progression of rat liver proliferation induced by partial hepatectomy (PH), by decreasing and/or desynchronizing cyclin D1 expression and activation into the nucleus, activation and nuclear translocation of STAT-1 and -3 proteins and altering retinoid metabolism. Interactions between retinoic acid and polyamines have been reported in the PH-induced rat liver regeneration. Therefore, we evaluated the effect of low dosage of α-tocopherol on PH-induced changes in polyamine metabolism. METHODS: This study evaluated the participation of polyamine synthesis and metabolism during α-tocopherol-induced inhibition of rat liver regeneration. In PH-rats (Wistar) treated with α-tocopherol and putrescine, parameters indicative of cell proliferation, lipid peroxidation, ornithine decarboxylase expression (ODC), and polyamine levels, were determined. RESULTS: Pre-treatment with α-tocopherol to PH-animals exerted an antioxidant effect, shifting earlier the increased ODC activity and expression, temporally affecting polyamine synthesis and ornithine metabolism. Whereas administration of putrescine induced minor changes in PH-rats, the concomitant treatment actually counteracted most of adverse actions exerted by α-tocopherol on the remnant liver, restituting its proliferative potential, without changing its antioxidant effect. Putrescine administration to these rats was also associated with lower ODC expression and activity in the proliferating liver, but the temporally shifting in the amount of liver polyamines induced by α-tocopherol, was also "synchronized" by the putrescine administration. The latter is supported by the fact that a close relationship was observed between fluctuations of polyamines and retinoids. CONCLUSIONS: Putrescine counteracted most adverse actions exerted by α-tocopherol on rat liver regeneration, restoring liver proliferative potential and restituting the decreased retinoid levels induced by α-tocopherol. Therefore interactions between polyamines and retinol, mediated by the oxidant status, should be taken into consideration in the development of new therapeutic strategies for pathologies occurring with liver cell proliferation.

The neurotoxic effects of vitamin A and retinoids.

Vitamin A (retinol) and its congeners - the retinoids - participate in a panoply of biological events, as for instance cell differentiation, proliferation, survival, and death, necessary to maintain tissue homeostasis. Furthermore, such molecules may be applied as therapeutic agents in the case of some diseases, including dermatological disturbances, immunodeficiency, and cancer (mainly leukemia). In spite of this, there is a growing body of evidences showing that vitamin A doses exceeding the nutritional requirements may lead to negative consequences, including bioenergetics state dysfunction, redox impairment, altered cellular signaling, and cell death or proliferation, depending on the cell type. Neurotoxicity has long been demonstrated as a possible side effect of inadvertent consumption, or even under medical recommendation of vitamin A and retinoids at moderate to high doses. However, the exact mechanism by which such molecules exert a neurotoxic role is not clear yet. In this review, recent data are discussed regarding the molecular findings associated with the vitamin A-related neurotoxicity.

Retinoid levels influence enterohemorrhagic Escherichia coli infection and Shiga toxin 2 susceptibility in mice.

Enterohemorrhagic Escherichia coli (EHEC) is a food-borne pathogen that produces Shiga toxin (Stx) and causes hemorrhagic colitis. Under some circumstances, Stx produced within the intestinal tract enters the bloodstream, leading to systemic complications that may cause the potentially fatal hemolytic-uremic syndrome. Although retinoids like vitamin A (VA) and retinoic acid (RA) are beneficial to gut integrity and the immune system, the effect of VA supplementation on gastrointestinal infections of different etiologies has been controversial. Thus, the aim of this work was to study the influence of different VA status on the outcome of an EHEC intestinal infection in mice. We report that VA deficiency worsened the intestinal damage during EHEC infection but simultaneously improved survival. Since death is associated mainly with Stx toxicity, Stx was intravenously inoculated to analyze whether retinoid levels affect Stx susceptibility. Interestingly, while VA-deficient (VA-D) mice were resistant to a lethal dose of Stx2, RA-supplemented mice were more susceptible to it. Given that peripheral blood polymorphonuclear cells (PMNs) are known to potentiate Stx2 toxicity, we studied the influence of retinoid levels on the absolute number and function of PMNs. We found that VA-D mice had decreased PMN numbers and a diminished capacity to produce reactive oxygen species, while RA supplementation had the opposite effect. These results are in line with the well-known function of retinoids in maintaining the homeostasis of the gut but support the idea that they have a proinflammatory effect by acting, in part, on the PMN population.

High dosing of α-tocopherol inhibits rat liver regeneration by modifying signal transducer and activator of transcription protein expression and its correlation with cell redox state and retinoid metabolism.

Lipid peroxidation (LP) promoted by partial hepatectomy (PH) is qualitatively distinct among subcellular fractions and temporally transient, probably being a necessary physiological event for rat liver regeneration. In fact, α-tocopherol (vitamin E [VE]) exerts adverse effects, partially inhibiting PH-induced rat liver regeneration and inducing decreased cyclin D1 expression. The phosphorylation of signal transducer and activator of transcription (STAT) factors 1 and 3 are involved in DNA synthesis and cyclin D1 expression after PH, which is stimulated by production of retinoic acid (RA). Hence, this study was aimed at addressing these events, and its association with cell redox state and oxidative stress, probably underlying VE effects on rat liver regeneration. PH-enhanced activation of STAT proteins, mainly as activated STAT-3, significantly change the cytoplasmic pool for STATs. The latter was associated to a more reduced cytoplasmic redox state and increased alcohol dehydrogenase (ADH)-mediated retinol oxidation to RA. Whereas α-tocopherol promoted minor changes in the parameters tested when administered to sham (control)-animals, pretreatment with VE blocked the PH-induced increase of reactive oxygen species (ROS), altering the pattern of STAT protein activation, blunting RA formation by decreased ADH activity, inducing higher liver caspase-3 activity and increasing tumor necrosis factor-α concentrations, while levels of interleukin-6 were decreased; altogether coinciding with disturbed PH-promoted changes on the liver redox state. In conclusion, altered activation and translocation of STAT-1 and -3 proteins and inhibited retinoid metabolism seem to be involved in the VE-induced inhibition of rat liver regeneration. Data suggest that a PH-induced increase of ROS could participate in the activation of STAT factors, retinoid metabolism and changes in the cell redox state during proliferation of liver cells.

Hepatic retinoid levels in seven fish species (teleosts) from a tropical coastal lagoon receiving effluents from iron-ore mining and processing.

The present study was undertaken to investigate the possible effects of Fe and trace element exposure on hepatic levels of retinoids in seven fish species. Concentrations of retinoids were measured in fish collected from a coastal lagoon in Brazil that receives effluents from an iron-ore mining and processing plant. Fish from nearby coastal lagoons were also included to assess possible differences related to chemical exposure. Results indicated considerable differences in hepatic retinoid composition among the various species investigated. The most striking differences were in retinol and derivative-specific profiles and in didehydro retinol and derivative-specific profiles. The Perciformes species Geophagus brasiliensis, Tilapia rendalli, Mugil liza, and Cichla ocellaris and the Characiforme Hoplias malabaricus were characterized as retinol and derivative-specific, while the Siluriformes species Hoplosternum littorale and Rhamdia quelen were didehydro retinol and derivative-specific fish species. A negative association was observed between Al, Pb, As, and Cd and hepatic didehydro retinoid levels. Fish with higher levels of hepatic Fe, Cu, and Zn showed unexpectedly significant positive correlations with increased hepatic retinol levels. This finding, associated with the positive relationships between retinol and retinyl palmitate with lipid peroxidation, may suggest that vitamin A is mobilized from other tissues to increase hepatic antioxidant levels for protection against oxidative damage. These data show significant but dissimilar associations between trace element exposure and hepatic retinoid levels in fish species exposed to iron-ore mining and processing effluents, without apparent major impacts on fish health and condition.

Involvement of alcohol and aldehyde dehydrogenase activities on hepatic retinoid metabolism and its possible participation in the progression of rat liver regeneration.

Liver alcohol dehydrogenase (ADH) activity is decreased towards exogenous substrates after partial hepatectomy (PH), probably due to putative endogenous substrates acting as ADH inhibitors. Hence, retinoids could be suitable candidates as such endogenous substrates. Therefore, cytosolic ADH kinetic analysis using several substrates, liver cytosolic and mitochondrial aldehyde dehydrogenase (ALDH) activities, retinal and retinol content, as well as expression of proteins for ADH and CRBPI (a retinol carrier protein) were determined in liver samples, at two stages of liver regeneration (one- or two-thirds PH). The effect of inhibiting in vivo liver ADH by 4-methylpyrazole (4-MP) was also evaluated after 70%-PH. With 70%-PH, in vitro ADH activity towards exogenous alcohols and aldehydes was diminished, but retinol oxidation was increased and retinal reduction was decreased. These activities that be due to the participation of an ADH type which did not correlate with the amount of immunoreactive ADH protein. Cytosolic and mitochondrial ALDH activities oxidized actively retinal, whereas retinol and CBRP-I expression were reduced in these animals. With 30%-PH, these changes were less evident and sometimes opposite to those found with 70%-PH. In addition, retinol readily inhibited ADH-mediated ethanol oxidation. Interestingly, in vivo 4-MP administration inhibited ADH activity in a dose-dependent manner correlating with a progressive inhibition of liver regeneration. In conclusion, PH-induced inhibition of ADH (mainly type I) seems to be related to ADH-mediated retinoid metabolism during liver proliferation. Thus, results suggest a role of ADH in retinoid metabolism, which is apparently required during rat liver regeneration.

Retinoides orales en niños y adolescentes / Oral retinoids in children and adolescent

Los retinoides son muy eficaces en el tratamiento de diversas enfermedades cutáneas que se presentan en niños, como psoriasis, acné e ictiosis. Sus efectos adversos potenciales limitan su uso, especialmente en la población pediátrica. Revisamos la eficacia y riesgos de la terapia con retinoides orales en niños y adolescentes. La toxicidad mucocutánea es el efecto adverso más frecuente, siendo generalmente bien tolerada, fácilmente tratable y reversible al discontinuar el tratamiento. Los efectos adversos sistémicos más graves incluyen la teratogenicidad y los efectos musculoesqueléticos, neurológicos y del sistema gastrointestinal. La educación y control de los pacientes pueden minimizar la ocurrencia de complicaciones.

[Advances in vitamin A and retinoid-binding proteins]. / Avances en vitamina A y proteínas que unen retinoides.

The recent interest in the study of vitamin A is due to the discovery of an increasing variety of functions, actions or associations of this vitamin with cellular differentiation of epithelial tissues, growth, reproduction and vision. This review describes the recent advances on the metabolism of vitamin A especially in regard to the retinoid-binding proteins in the transport of hydrophobic retinoids and their participation in the transformation of the different retinoids in the organism as well as in the interior of the cell. Seven binding proteins have been discovered and well characterized: retinol binding protein (RBP); cellular retinol-binding protein (CRBP); cellular retinol-binding protein type two (CRBP-II); cellular retinoic acid binding protein (CRABP); cellular retinoic acid binding protein type two (CRABP-II); cellular retinal-binding protein (CRALBP) and interphotoreceptor retinol-binding protein (IRBP). Their specific roles are presented. Also the function and mechanisms of the regulation of gene expression by retinoic acid is described.

Interaction of fatty acids, acyl-CoA derivatives and retinoids with microsomal membranes: effect of cytosolic proteins.

This paper reviews characteristics of microsomal membrane structure; long chain fatty acids, acyl CoA derivatives, retinoids and the microsomal formation of acyl CoA derivatives and retinyl esters. It is analyzed how the movement of these molecules at the intracellular level is affected by their respective binding proteins (Fatty acid binding protein, acyl CoA binding protein and cellular retinol binding protein). Studies with model systems using these hydrophobic ligands and the lipid-binding or transfer proteins are also described. This topic is of interest especially because in the esterification of retinol the three substrates and the three binding proteins may interact.

Severe hypervitaminosis A in siblings: evidence of variable tolerance to retinol intake.

A 2-year-old boy had signs and symptoms of chronic hypervitaminosis A. A course of increasing severity led to eventual death. A younger brother later had similar clinical features. Chicken liver spread containing up to 420 IU/g vitamin A was the likely source of intoxication. Markedly elevated circulating retinyl ester levels have persisted in the surviving sibling for 3 subsequent years despite severe restriction of vitamin A intake. A therapeutic trial of the carbohydrate-derived complexing agent 2-hydroxypropyl-beta-cyclodextrin was initiated. Circulating retinyl esters transiently increased during the infusion (from 407 to 4791 micrograms/dL), and urinary total vitamin A excretion, undetectable before infusion, increased to 23 micrograms/dL after infusion. The frequency of hypervitaminotic episodes has decreased somewhat in the 2 years since the infusion, probably related to dietary vitamin A restriction. The occurrence of this syndrome in two brothers, while a sister ingesting the same diet remains completely healthy, suggests an inherited variance in tolerance to vitamin A intake.