71 research outputs found
Melanogenesis in Visceral Tissues of Salmo salar. A link between Immunity and Pigment production?
No full textThe Atlantic salmon (Salmo salar L.) is an ichthyic breeding species of considerable economic importance. The accumulation of pigments in salmon visceral organs and musculature (1) adversely affects the quality of fish products and is a problem for the aquaculture industry. Moreover, a close relationship between nutrition products and human health has been widely demonstrated.
Liver, spleen, kidney and other organs of lower vertebrates harbour a visceral pigment cell system with an embryonic origin that differs from that of melanocytes (2). In teleosts, melanin-containing cells, named melano-macrophages, occur in the reticulo-endothelial system and are mainly located in kidney and spleen. In these organs, the pigment content may increase in different physiological conditions and as a response to pathological and inflammatory states (2).
With the aim to reveal novel functions and behaviour of the poorly understood teleost extracutaneous pigment system, we undertook the investigation of aspects of the melanogenic systems of Atlantic salmon tissues as well as of leukocyte salmon head-kidney cells (SHK-1 cells). SHK-1 cells is a long-term cell line derived from macrophages of the Atlantic salmon head-kidney (3). Using a radiometric method we demonstrated that a tyrosinase activity is present in SHK-1 cells and salmon tissues. Western blotting showed the presence of a tyrosinase protein. Moreover differing melanin-producing activities in salmon tissues were demonstrated suggesting that pigmented cells in the organs undergo a tissue-specific differentiation. These results were confirmed by a tyrosinase protein expression in lymphoid organs, also highlighted by immunohistochemical analysis.
As teleosts lack lymph nodes and Peyer’s patches, the head-kidney and spleen are regarded as the most important secondary lymphoid organs (4). The detection of tyrosinase activity in lymphoid organs indicates that a link may exist between the extracutaneous pigmentary system and the immune system in salmon.
The extracutaneous melanogenetic system could be used as a biomarker for the health status of farmed fishes.
References:
1. Koppang, E.O., Haugarvoll, E., Hordvik, I., Aune, L., Poppe, T. (2005) Journal of Fish Diseases 28: 13-22.
2. Agius, C., Roberts, R.J. (2003) Journal of Fish Diseases 26: 499-509.
3. Dannevig, B., Falk, K., Narmork, E. (1995) Journal of General Virology 76: 1353-1359.
4. Zapata, A., Amemiya, CT. (2000) Curr Top Microbiol Immunol 248: 67-107
Bioactive paper platform for colorimetric phenols detection.
No full textPolyphenols, as food antioxidants, are of great interest due to their health benefits as they decrease the risks of cancer and coronary cardiopathy (1). Moreover they influence the quality and organoleptic characteristics of foods (2). Lastly, some neurotransmitters are phenolic compounds.
Hence the need to work out a sensitive, portable and inexpensive detection methods to monitor these compounds (3). We developed a disposable paper-based bioassay for the detection of phenolic compounds; the assay was successfully applied for the determination of polyphenols in a real matrix such as wine. The colorimetric quantification of the analyte is based on an enzymatic assay. The tyrosinase enzyme has been immobilized on a filter paper by simple over-spotting with 3-methyl-2-benzothiazolinone hydrazone (MBTH), that allows the detection of phenols by forming stable colored adducts with their enzymatic oxidation products. The color intensity of the adduct (developed after 5 min of reaction) was found to increase proportionally with the increase of the phenolic substrate concentrations. Analyte detection can be achieved by eye and quantification can be simply obtained by using a camera phone and an image analysis software.
The response, characteristics of the sensor were determined using l-3,4-dihydroxyphenyl-alanine (l-DOPA), an archetype substrate of tyrosinase, as the analyte.
This disposable paper-based biosensor relies on a rapid and simple method, without need of sophisticated instrumentation and trained personnel and could be extremely useful in remote locations or developing countries which does not have ready access to laboratory facilities and where simple, sensitive and low cost bioassays are essentials
Seasonal variations of Rana esculenta L. skin tyrosinase
No full textVarious enzymes are known to be involved in melanin biosynthesis, but the key role appertains to tyrosinase. In amphibians this enzyme displays peculiar characteristics: i) it requires an activation process; ii) its level of enzymatic activity in the animal skin changes depending on the season. In this work, by using chymotrypsin, subtilisin and SDS as putative activators, we studied the activation process of the skin pro-tyrosinase of Rana esculenta L. in different seasons over a period of two years. We found that chymotrypsin and subtilisin were able to yield an active enzyme, but not SDS. The maximum levels of tyrosinase activity were recorded in winter and the minimum in summer. We detected tyrosinase activity in the melanosomal fraction, where the enzyme form was least sensitive to proteolytic activation, probably corresponding to a "mature" tyrosinase. The enzyme forms found in the microsomal and soluble fractions were more sensitive to proteolytic activation, probably corresponding to "immature" tyrosinase. On SDS-PAGE, the tyrosinase activity assays showed a dopa-positive band at 200 kDa and a second aggregated band with a still higher molecular mass. The significance of these results in frog melanogenesis regulation is discussed
Extracutaneous melanogenetic system in Dicentrarchus labrax and Sparus aurata (Linnaeus, 1758): a preliminary study.
No full textMelanogenesis has been mainly studied in mammalian melanocytes and melanoma cells. In heterothermic vertebrates, melanin-containing cells, named melano-macrophages, occur in visceral organs (1). In Teleosts, these cells are copious and dispersed in the stroma of haemopoietic tissues in kidney and spleen. In these organs, pigment content increases in particular states such as pathological and inflammatory conditions (1).
To gain a more extensive knowledge of the extracutaneous melanin-containg cell system, we investigated for the enzymes responsible of melanogenesis in two fish species: sea bass (Dicentrarchus labrax, L.) and gilthead seabream (Sparus aurata, L.). We looked for DOPA oxidase (DO) and peroxidase (POD) activity in kidney and spleen of breeding specimens caught in spring and winter.
Kidney melanosomal protein fractions revealed a very high DO activity in both species. A seasonal variation of the enzymatic activity (higher in winter and lower in spring) was found. Kidney melano-macrophages also showed a POD activity with seasonal variations similar to those observed in DO activity. Assays on spleen tissue extract revealed only a POD activity in spring, whereas in winter both activities were detected.
In conclusion we demonstrate, in these two fish species, that the melano-macrophages from kidney and spleen possess a melanogenetic capability that share common features with the melano-macrophages studied in other low vertebrate species.
The extracutaneous melanogenetic system could be used as a biomarker for the health status of farmed fishes. Great interest is directed to human diet because of the close relationship between nutrition and health.
References
1)Agius C., Roberts R. J., Journal of Fish Diseases, 200326, 499-509
Putting an "end" to the motor cortex representations of action words
No full textLanguage processing is an example of implicit learning of multiple statistical cues that provide probabilistic information regarding word structure and use. Much of the current debate about language embodiment is devoted to how action words are represented in the brain, with motor cortex activity evoked by these words assumed to selectively reflect conceptual content and/or its simulation. We investigated whether motor cortex activity evoked by manual action words (e.g., caress) might reflect sensitivity to probabilistic orthographic-phonological cues to grammatical category embedded within individual words. We first review neuroimaging data demonstrating that nonwords evoke activity much more reliably than action words along the entire motor strip, encompassing regions proposed to be action category specific. Using fMRI, we found that disyllabic words denoting manual actions evoked increased motor cortex activity compared with non-body-part-related words (e.g., canyon), activity which overlaps that evoked by observing and executing hand movements. This result is typically interpreted in support of language embodiment. Crucially, we also found that disyllabic nonwords containing endings with probabilistic cues predictive of verb status (e.g., -eve) evoked increased activity compared with nonwords with endings predictive of noun status (e.g., -age) in the identical motor area. Thus, motor cortex responses to action words cannot be assumed to selectively reflect conceptual content and/or its simulation. Our results clearly demonstrate motor cortex activity reflects implicit processing of ortho-phonological statistical regularities that help to distinguish a word's grammatical class
Analisi del sistema tirosinasico nella linea cellulare SHK-1 di Salmo Salar L. (Teleostei)
No full text
Analisi del sistema tirosinasico nella linea cellulare SHK-1 di Salmo Salar L. (Teleostei)
No full textMELANOGENESIS IN VISCERAL TISSUES OF SALMO SALAR. A LINK BETWEEN IMMUNITY AND PIGMENT PRODUCTION?
No full textMelanogenesis is mostly studied in melanocytes and melanoma cells, but much less is known about other
pigment cell systems. Liver, spleen, kidney, and other organs of lower vertebrates harbour a visceral pigment cell system
with an embryonic origin that differs from that of melanocytes distinct embryonic origin. In teleosts, melanin-containing
cells occur in the reticulo-endothelial system and are mainly in the kidney and spleen. The Atlantic salmon (Salmo salar
L.) is an ichthyic breeding species of considerable economic importance. The accumulation of pigments in salmon visceral
organs and musculature adversely affects the quality of fish products and is a problem for the aquaculture industry. With
the aim to reveal novel functions and behaviour of the salmonid extracutaneous pigment system, we investigated aspects of
the melanogenic systems in the tissues of Atlantic salmon, as well as in SHK-1 cells, which is a long-term cell line derived
from macrophages of the Atlantic salmon head-kidney. We demonstrate that a melanogenic system is present in SHK-1
cells, head-kidney, and spleen tissues. As teleosts lack lymph nodes and Peyer’s patches, the head-kidney and spleen are
regarded as the most important secondary lymphoid organs. The detection of tyrosinase activity in lymphoid organs
indicates that a link exists between the extracutaneous pigmentary system and the immune system in salmon
CHARACTERIZATION OF THE SKIN MELANOGENETIC SYSTEM OF RANA ESCULENTA L
No full textTyrosinase is the key enzyme in melanin biosynthesis.We have characterized the skin tyrosinase of Rana esculenta L. in extracts of dorsal skin and by cloning the gene for its functional expression in a heterologous system. Analysis of R. esculenta tyrosinase ORF showed that it is particularly well conserved as compared with other amphibian tyrosinases, but the levels of homology remain high with respect to mammal tyrosinases. The frog enzyme maintains the typical characteristics of tyrosinases, namely the presence of an EGF-like domain, inserted in a laminin–like EGF domain, 15 conserved cysteine residues grouped in 3 clusters, two copper binding sites (CuA and CuB), and a peptide signal for its entry into the RER. Tyrosinase is expressed in HEK293T cells as an active enzyme and its activity is not increased by the action of proteases, and is only slightly increased by copper. Assays of tyrosinase activity on nondenaturing SDS-PAGE demonstrate the presence of two bands, with a molecular weight of approximately 63 kDa and 67 kDa. Conversely, the skin extracts of R. esculenta are lightly active after extraction and need to be activated by proteases like nagarse to reach maximal activation levels. The degree to which these extracts can be activated shows seasonal fluctuations, reaching maximum levels in winter. Assays of tyrosinase activity on SDS-PAGE demonstrate a dopa positive band, with a molecular weight of approximately 200 kDa and a high aggregate that does not migrate in electrophoresis gel in samples treated with nagarse. The different behaviour of the two forms of tyrosinase suggest the formation in vivo of an aggregate containing tyrosinase as shown by its dopa positivity. The proteolytic activation of the skin tyrosinase could probably depend on the exposure of domains favoring the formation of active multimeric complexes, like those already hypothesized in mammals or/and removal of some inhibitor
- …
