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torsdag 7 december 2023

ONCOLOGIA: Sfingosiini-1 fosfaattilyaasin säätelyn merkityksestä myeliinin aineenvaihdunnassa

 

. 2011 Mar;1811(3):119-28.
doi: 10.1016/j.bbalip.2010.12.005. Epub 2010 Dec 22.

Heterogeneous sphingosine-1-phosphate lyase gene expression and its regulatory mechanism in human lung cancer cell lines

Hiromi Ito  1 , Kayo Yoshida, Masashi Murakami, Kazumi Hagiwara, Noriko Sasaki, Misa Kobayashi, Akira Takagi, Tetsuhito Kojima, Sayaka Sobue, Motoshi Suzuki, Keiko Tamiya-Koizumi, Mitsuhiro Nakamura, Yoshiko Banno, Yoshinori Nozawa, Takashi Murate
Affiliations

Abstract

The role of sphingolipid metabolic pathway has been recognized in determining cellular fate. Although sphingolipid degradation has been extensively studied, gene expression of human sphingosine 1-phosphate lyase (SPL) catalyzing sphingosine 1-phosphate (S1P) remains to be determined. Among 5 human lung cancer cell lines examined, SPL protein levels paralleled the respective mRNA and enzyme activities. Between H1155 and H1299 cells used for further experiments, higher cellular S1P was observed in H1155 with higher SPL activity compared with H1299 with low SPL activity. GATA-4 has been reported to affect SPL transcription in Dictyostelium discoideum. GATA-4 was observed in H1155 but not in other cell lines. Overexpression of GATA-4 in H1299 increased SPL expression. However, promoter analysis of human SPL revealed that the most important region was located between -136bp and -88bp from the first exon, where 2 Sp1 sites exist but no GATA site. DNA pull-down assay of H1155 showed increased DNA binding of Sp1 and GATA-4 within this promoter region compared with H1299. Electrophoresis mobility shift assay (EMSA), chromatin immunoprecipitation (ChIP) assay, reporter assay using mutated binding motif, and mithramycin A, a specific Sp1 inhibitor, suggest the major role of Sp1 in SPL transcription and no direct binding of GATA-4 with this 5' promoter region. The collaborative role of GATA-4 was proved by showing coimmunoprecipitation of Sp1 and GATA-4 using GST-Sp1 and overexpressed GATA-4. Thus, high SPL transcription of H1155 cells was regulated by Sp1 and GATA-4/Sp1 complex formation, both of which bind to Sp1 sites of the 5'-SPL promoter.

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torsdag 11 maj 2023

Kolesterolin (Chol) transgalaktosyloituminen (GalChol) ja transglykosyloituminen (GlcChol)

 

Glucocerebrosidases (GBA)  catalyze a transgalactosylation reaction that yields a newly-identified brain sterol metabolite, galactosylated cholesterol.
Akiyama H, Ide M, Nagatsuka Y, Sayano T, Nakanishi E, Uemura N, Yuyama K, Yamaguchi Y, Kamiguchi H, Takahashi R, Aerts JMFG, Greimel P, Hirabayashi Y. J Biol Chem. 2020 Apr 17;295(16):5257-5277. doi: 10.1074/jbc.RA119.012502. Epub 2020 Mar 6. PMID: 32144204 Free PMC article.

β-Glucocerebrosidase (GBA) hydrolyzes glucosylceramide (GlcCer) to generate ceramide(Cer). Previously, we demonstrated that lysosomal GBA1 and nonlysosomal GBA2 possess not only GlcCer hydrolase activity, but also transglucosylation activity to transfer the glucose residue from GlcCer to cholesterol to form β-cholesterylglucoside (β-GlcChol) in vitro β-GlcChol is a member of sterylglycosides present in diverse species. 

How GBA1 and GBA2 mediate β-GlcChol metabolism in the brain is unknown. 

 Here, we purified and characterized sterylglycosides from rodent and fish brains. Although glucose is thought to be the sole carbohydrate component of sterylglycosides in vertebrates, structural analysis of rat brain sterylglycosides revealed the presence of galactosylated cholesterol (β-GalChol), in addition to β-GlcChol. 

Analyses of brain tissues from GBA2-deficient mice and GBA1- and/or GBA2-deficient Japanese rice fish (Oryzias latipes) revealed that GBA1 and GBA2 are responsible for β-GlcChol degradation and formation, respectively, and that both GBA1 and GBA2 are responsible for β-GalChol formation. 

 Liquid chromatography-tandem MS revealed that β-GlcChol and β-GalChol are present throughout development from embryo to adult in the mouse brain. We found that β-GalChol expression depends on galactosylceramide (GalCer), and developmental onset of β-GalChol biosynthesis appeared to be during myelination.

 We also found that β-GlcChol and β-GalChol are secreted from neurons and glial cells in association with exosomes. 

In vitro enzyme assays confirmed that GBA1 and GBA2 have transgalactosylation activity to transfer the galactose residue from GalCer to cholesterol to form β-GalChol. This is the first report of the existence of β-GalChol in vertebrates and how β-GlcChol and β-GalChol are formed in the brain.

Keywords: brain; cholesterol; galactosylated cholesterol; glucocerebrosidase; glycolipid; mass spectrometry (MS); sterol; sterylglycoside; transglycosylation; β-cholesterylgalactoside
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GBA1, GBA2 ja GBA3, glukosyylikeramidaasien perhe

GBA1  (1q22)  Lysosomaalinen glykosyylikeramidaasi, Glukokerebrosidaasi

 https://www.genecards.org/cgi-bin/carddisp.pl?gene=GBA1&keywords=GBA1

GBA2 (9p13.3) Non-lysosomaalinen glykosyylikeramidaasi

 https://www.genecards.org/cgi-bin/carddisp.pl?gene=GBA2&keywords=GBA2

GBA3  (4p15.2), Sytosolinen glukosyylikeramidaasi beeta 3  pystyy käsittelemään dietäärisiä glykosyyliyhdisteitäkin.

https://www.genecards.org/cgi-bin/carddisp.pl?gene=GBA3&keywords=GBA3

Entrez Gene Summary for GBA3 Gene

  • The protein encoded by this gene is a cytosolic enzyme that can hydrolyze several types of glycosides. The enzyme has its highest activity at neutral pH and is predominantly expressed in human liver, kidney, intestine, and spleen. This gene is a polymorphic pseudogene, with the most common allele being the functional allele that encodes the full-length protein. Some individuals contain a single nucleotide polymorphism that results in a premature stop codon in the coding region, and therefore this allele is pseudogenic due to the failure to produce a functional full-length protein. Alternative splicing of this gene results in multiple transcript variants. [provided by RefSeq, Apr 2022]

GeneCards Summary for GBA3 Gene

GBA3 (Glucosylceramidase Beta 3 (Gene/Pseudogene)) is a Protein Coding gene. Diseases associated with GBA3 include Gaucher's Disease. Among its related pathways are Sphingolipid metabolism and Metabolism. Gene Ontology (GO) annotations related to this gene include hydrolase activity, hydrolyzing O-glycosyl compounds and beta-glucosidase activity. An important paralog of this gene is LCT.

UniProtKB/Swiss-Prot Summary for GBA3 Gene

Neutral cytosolic beta-glycosidase with a broad substrate specificity that could play a role in the catabolism of glycosylceramides (PubMed:11389701, 11784319, 20728381, 26724485, 17595169, 33361282). Has a significant glucosylceramidase activity in vitro (PubMed:26724485, 17595169). However, that activity is relatively low and its significance in vivo is not clear (PubMed:26724485, 17595169, 20728381). Hydrolyzes galactosylceramides/GalCers, glucosylsphingosines/GlcSphs and galactosylsphingosines/GalSphs (PubMed:17595169). However, the in vivo relevance of these activities is unclear (PubMed:17595169). It can also hydrolyze a broad variety of dietary glycosides including phytoestrogens, flavonols, flavones, flavanones and cyanogens in vitro and could therefore play a role in the metabolism of xenobiotics (PubMed:11784319). Possesses transxylosylase activity in vitro using xylosylated ceramides/XylCers (such as beta-D-xylosyl-(1<->1')-N-acylsphing-4-enine) as xylosyl donors and cholesterol as acceptor (PubMed:33361282). Could also play a role in the catabolism of cytosolic sialyl free N-glycans (PubMed:26193330). ( GBA3_HUMAN,Q9H227 )

Tocris Summary for GBA3 Gene

  • Glycosylases are a group of enzymes that includes glucosidases, mannosidases and heparanases. There are two glucosidase subtypes, both found in the gut. They hydrolyze terminal (1,4)alpha-glucosidic linkages and (1,6)beta-glucosidic linkages, liberating alpha-glucose and beta-glucose.

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GlcCer, GalCer , geramide, glukosyl, galaktosyl , GBA1 ja GBA2 transglykosyloivat ja myös transgalaktosyloivat , GlcChol, GalChol

 AIEMPI tekstini:

Synteesi KERAMIDISTA kohti glykosfingolipidejä.KERAMIDI (Cer) syntyy endoplasmisessa retikulumissa ja se kulkeutuu Golgin laitteeseen.. Keramidia purkautuu myös sfingolipidien kataboliasta ja käytetään uudestaan.

GlcCer, glucosylceramide, glukosyylikeramidi

GlcCer syntaasientsyymi lisää glukoosin beta-sidoksella keramidin OH- ryhmään 1- asemassa.Glukosyylikeramidiin tapahtuva jatkosyntetisoiminen tapahtuu sen jälkeen kun karakterisoimattoman flippaasi-entsyymin avulla on tapahtunut vaihde Golgin laitteen ontelon puolelle.

http://link.springer.com/chapter/10.1007%2F978-4-431-67877-9_1#page-1

GalCer, galactosylceramide, galaktosyylikeramidi muodostusta voi myös joskus tapahtua keramidista. Tämä tapahtuu endoplasmisen retikulumin ontelon puolella, vaikkakin galaktosyylikeramidien synteesitietä tapahtuu vain hyvin rajoitetusti.

http://link.springer.com/chapter/10.1007%2F978-4-431-67877-9_8#page-1

PÄIVITYSTÄ GlcCer ja GalCer synteesiasiaan  11.5. 2023

On uudempaa tietoa on GlcCer ja GalCer muodostumisesta aineenvaihdunnassa vuodelta 2020

https://www.jbc.org/article/S0021-9258(17)48548-6/fulltext

β-Glucocerebrosidase (GBA) hydrolyzes glucosylceramide (GlcCer) to generate ceramide. Previously, we demonstrated that lysosomal GBA1 and nonlysosomal GBA2 possess not only GlcCer hydrolase activity, but also transglucosylation activity to transfer the glucose residue from GlcCer to cholesterol to form β-cholesterylglucoside (β-GlcChol) in vitro. β-GlcChol is a member of sterylglycosides present in diverse species. How GBA1 and GBA2 mediate β-GlcChol metabolism in the brain is unknown. Here, we purified and characterized sterylglycosides from rodent and fish brains. Although glucose is thought to be the sole carbohydrate component of sterylglycosides in vertebrates, structural analysis of rat brain sterylglycosides revealed the presence of galactosylated cholesterol (β-GalChol), in addition to β-GlcChol. Analyses of brain tissues from GBA2-deficient mice and GBA1- and/or GBA2-deficient Japanese rice fish (Oryzias latipes) revealed that GBA1 and GBA2 are responsible for β-GlcChol degradation and formation, respectively, and that both GBA1 and GBA2 are responsible for β-GalChol formation. Liquid chromatography–tandem MS revealed that β-GlcChol and β-GalChol are present throughout development from embryo to adult in the mouse brain.

We found that β-GalChol expression depends on galactosylceramide (GalCer), and developmental onset of β-GalChol biosynthesis appeared to be during myelination. We also found that β-GlcChol and β-GalChol are secreted from neurons and glial cells in association with exosomes.

In vitro enzyme assays confirmed that GBA1 and GBA2 have transgalactosylation activity to transfer the galactose residue from GalCer to cholesterol to form β-GalChol. This is the first report of the existence of β-GalChol in vertebrates and how β-GlcChol and β-GalChol are formed in the brain.


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tisdag 7 mars 2023

Pitkäketjuisten essentiellien rasvahappojen osuudet nykyajan dieeteissä

Pohdittavaa! Onko nykyajan suositukset  evoluutionäkökohdasta käsin aivan parhainta mahdollista tieteellistä tietoa ihmisen perustavista  ravitsemuksellisista tarpeista?  Yleensä perustetaan suositukset  ihmisten käyttämään yleiseen  ravintoon nykyaikana eikä  ihmisen evolutionaaliseen  ravitsemukseen esim  satoja vuosia sitten. tosin  tietämys on vain possulkevaa tietoa. Silloin EI OLLUT nykyaan prosessoituja ja modifioituja ja jalostettuja ym tuotteita  ainakaan, vaikka sodan, kadon, ruton ja genosidisten piirteiden  ravinnonsaantiin vaikuttamat asiat lienevät  kaikkina aikoina samanlaisen alkeellisia. 
 
doi: 10.1016/j.plefa.2006.05.010. Epub 2006 Jul 28.

Long-chain polyunsaturated fatty acids in maternal and infant nutrition

Frits A J Muskiet  1 , Saskia A van Goor, Remko S Kuipers, Francien V Velzing-Aarts, Ella N Smit, Hylco Bouwstra, D A Janneke Dijck-Brouwer, E Rudy Boersma, Mijna Hadders-Algra
Affiliations
Abstract

Homo sapiens has evolved on a diet rich in alpha-linolenic acid  C18:3 n3 (short  omega3)  and long chain polyunsaturated fatty acids (LCP). We have, however, gradually changed our diet from about 10,000 years ago and accelerated this change from about 100 to 200 years ago. The many dietary changes, including lower intake of omega3-fatty acids, are related to 'typically Western' diseases. After a brief introduction in essential fatty acids (EFA), LCP and their functions, this contribution discusses our present low status of notably LCP omega3 in the context of our rapidly changing diet within an evolutionary short time frame. It then focuses on the consequences in pregnancy, lactation and neonatal nutrition, as illustrated by some recent data from our group. We discuss the concept of a 'relative' EFA/LCP deficiency in the fetus as the outcome of high transplacental glucose flux. This flux may in the fetus augment de novo synthesis of fatty acids, which not only dilutes transplacentally transported EFA/LCP, but also causes competition of de novo synthesized oleic acid ( C18:1n9) with linoleic acid for delta-6 desaturation. Such conditions were encountered by us in mothers with high body mass indices, diabetes mellitus and preeclampsia. The unifying factor might be compromised glucose homeostasis. In search of the milk arachidonic acid (AA, C20:4n6) and docosahexaenoic acid (DHA, C22:6n3) contents of our African ancestors, we investigated women in Tanzania with high intakes of freshwater fish as only animal lipid source. These women had milk AA and DHA contents that were well above present recommendations for infant formulae. Both studies stimulate rethinking of 'optimal homeostasis'. Subtle signs of dysbalanced maternal glucose homeostasis may be important and observations from current Western societies may not provide us with an adequate basis for dietary recommendations.  

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Essentiellit rasvahapot AA, EPA ja DHA solukalvoissa.

 AA, C20:4 omega6 (Eicosatetraeenihappo eli  arakidonihappo  C20:4 omega 6 linjasta on lähtöaine eikosanoideille. Linolihappo (LA)   kasvikunnasta johtaa tähän arakidonihappoon kehoentsyymien avulla.
EPA, C20:5 (eicosapentaeenihappo)   omega 3 linjasta.  Alfa-linoleenihappo (ALA) kasvikunnasta johtaa tähän EPA:5  rasvahappoon. 
DHA, C22:6 omega 3 linjasta. Docosahexaeenihappoa muodustuu  jatkossa ihmiskehon entsyymeilä .  
 omega 6 linjasta. Myös  kalarasvoista saa näitä pitkiä rasvahappoja valmiina EPA ja DHA muotoa.

Tiedetään, että ihmisen kaikki  solut, joissa  vain on membraaneja, keräävät membraanin lipidirakenteeseen arakidonihappoa (C20:4), koska  se on lähtöaine  solun monissa funktioissa ja varsinkin kudosten korjaantumisissa ja immuunivasteessa.  Entä sitten  paralleelin  omega3-linjan  rasvahappojen merkitys? Niitäkin solumembraanin  fosfolipidit(PL) keräävät rakenteeseensa ja tämä rasvahappojen valinta  fosfolipidirakenteeseen on  solu- ja kudosspesifistä. Koska nämä kolme pitkää tärkeää essentielliä rakennetta ovat hieman erilaisia ja  koska niiden  kertymiseen vaikuttaa dieetin antamat  essentiellit  linjaa muodostavat  alkumuodot kuten linolihapon (C18:2  omega 6)  ja alfalinoleenihapon (C18:3 omega 3)  saanti, niin mitä tiedeään membraaniaanirakennevaikutuksesta?
 
. 2021;62:100106.
doi: 10.1016/j.jlr.2021.100106. Epub 2021 Aug 13.

EPA and DHA containing phospholipids have contrasting effects on membrane structure

Samuel C R Sherratt  1 , Rebecca A Juliano  2 , Christina Copland  2 , Deepak L Bhatt  3 , Peter Libby  3 , R Preston Mason  4
Affiliations
Free PMC article
Abstract

Omega-3 FAs EPA and DHA influence membrane fluidity, lipid rafts, and signal transduction. A clinical trial, Reduction of Cardiovascular Events with Icosapent Ethyl-Intervention Trial, demonstrated that high-dose EPA (4 g/d icosapent ethyl) reduced composite cardiovascular events in statin-treated high-risk patients. EPA benefits correlated with on-treatment levels, but similar trials using DHA-containing formulations did not show event reduction. We hypothesized that differences in clinical efficacy of various omega-3 FA preparations could result from differential effects on membrane structure. To test this, we used small-angle X-ray diffraction to compare 1-palmitoyl-2-eicosapentaenoyl-sn-glycero-3-phosphocholine (PL-EPA), 1-palmitoyl-2-docosahexaenoyl-sn-glycero-3-phosphocholine (PL-DHA), and 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (PL-AA) in membranes with and without 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and cholesterol. Electron density profiles (electrons/Å3 vs. Å) were used to determine membrane structure, including membrane width (d-space). PL-EPA and PL-DHA had similar membrane structures without POPC and/or cholesterol but had contrasting effects in the presence of POPC and cholesterol. PL-EPA increased membrane hydrocarbon core electron density over an area of ±0-10 Å from the center, indicating an extended orientation. PL-DHA increased electron density in the phospholipid head group region, concomitant with disordering in the hydrocarbon core and a similar d-space (58 Å). Adding equimolar amounts of PL-EPA and PL-DHA produced changes that were attenuated compared with their separate effects. PL-AA increased electron density centered ±12 Å from the membrane center. The contrasting effects of PL-EPA, PL-DHA, and PL-AA on membrane structure may contribute to differences observed in the biological activities and clinical actions of various omega-3 FAs.

Keywords: X-ray diffraction; arachidonic acid; docosahexaenoic acid; eicosapentaenoic acid; membrane structure; omega-3 FAs.


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