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- Ronquist, Göran K., et al.
(författare)
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Prostasomal DNA Characterization and Transfer Into Human Sperm
- 2011
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Ingår i: Molecular Reproduction and Development. - : Wiley. - 1040-452X .- 1098-2795. ; 78:7, s. 467-476
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Tidskriftsartikel (refereegranskat)abstract
- Human prostasomes, exosome-like microvesicles secreted by acinar cells of the prostate gland, contain chromosomal DNA. Agarose gel electrophoresis of DNA from seminal prostasomes displayed fragments of over 12 kb and smaller, with a distinct band around 1 kb that was excised, cloned, and sequenced. The sequences showed 8 out of 25 clones (32%) originating from genes. We elaborated the concept further by carrying out a genome-wide DNA copy number analysis of prostasomal DNA, hypothesizing that human prostasomes contain fragments of DNA randomly selected from the entire genome. Acridine orange-stained prostasomes were incubated with freshly prepared sperm for different times, and a transfer of acridine orange-stained prostasomal DNA to sperm (preferentially the head region) was observed. Fluorescence microscopy of slices in the center of 14 optical slides of the sperm head displayed an even fluorescence rather than a halo-like one, indicating DNA-uptake rather than just binding along the sperm head membrane.
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- Ronquist, Göran K, et al.
(författare)
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Prostasomes are heterogeneous regarding size and appearance but affiliated to one DNA-containing exosome family
- 2012
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Ingår i: The Prostate. - : Wiley. - 0270-4137 .- 1097-0045. ; 72:16, s. 1736-1745
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND:Prostate acinar epithelial cells release microvesicles (prostasomes) that possess pleiotropic biological effects relevant for successful fertilization. Prostasomes are formed in a similar way as exosomes but are heterogeneous as regards size and appearance. Like exosomes they are thought to be mediators of intercellular communication.METHODS:We prepared seminal prostasomes in accordance with the prevailing protocol for exosome preparation including passage through a 0.2 µm filter and centrifugation in a sucrose gradient.RESULTS:We compared the "filterable prostasomes" with those trapped on the filter ("nonfilterable prostasomes") and, qualitatively, no conspicuous differences were apparent regarding ultrastructure and SDS-PAGE banding pattern. Moreover, both types of prostasomes contained DNA fragments and Western blot revealed presence of prostate specific membrane antigen (PSMA), CD38, and annexin A1.CONCLUSIONS: Reasonably, prostasomes could be included in the exosome family and be regarded as one entity containing chromosomal DNA.
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- Ronquist, K Göran, et al.
(författare)
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Human prostasomes express glycolytic enzymes with capacity for ATP production
- 2013
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Ingår i: American Journal of Physiology. Endocrinology and Metabolism. - : American Physiological Society. - 0193-1849 .- 1522-1555. ; 304:6, s. E576-E582
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Tidskriftsartikel (refereegranskat)abstract
- Prostasomes are prostate-derived, exosome-like microvesicles that transmit signaling complexes between the acinar epithelial cells of the prostate and sperm cells. A vast majority of prostasomes has a diameter of 30 - 200 nm and they are generally surrounded by a classical membrane bilayer. Using a selected proteomic approach, it became increasingly clear that prostasomes harbor distinct subsets of proteins that may be linked to adenosine triphosphate (ATP) metabolic turnover that in turn might be of importance in the role of prostasomes as auxiliary instruments in the fertilization process. Among the 21 proteins identified most of the enzymes of anaerobic glycolysis were represented and three of the glycolytic enzymes present are among the ten top proteins found in most exosomes, once again linking prostasomes to the exosome family. Other prostasomal enzymes involved in ATP turnover were adenylate kinase, ATPase, 5'-nucleotidase and hexose transporters. The identified enzymes in their prostasomal context were operational for ATP formation when supplied with substrates. The net ATP production was low due to a high prostasomal ATPase activity that could be partially inhibited by vanadate that was utilized in order to profile the ATP forming ability of prostasomes. Glucose and fructose were equivalent as glycolytic substrates for prostasomal ATP formation and the enzymes involved were apparently surface-located on prostasomes, since an alternative substrate not being membrane-permeable (glyceraldehyde 3-phosphate) was operative, too. There is no clear cut function linked to this subset of prostasomal proteins but some possible roles are discussed.
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| 5. |
- Ronquist, K Göran, et al.
(författare)
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Prostasomes from four different species are able to produce extracellular adenosine triphosphate (ATP)
- 2013
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Ingår i: Biochimica et Biophysica Acta - General Subjects. - : Elsevier BV. - 0304-4165 .- 1872-8006. ; 1830:10, s. 4604-4610
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND:Prostasomes are extracellular vesicles. Intracellularly they are enclosed by another larger vesicle, a so called "storage vesicle" equivalent to a multivesicular body of late endosomal origin. Prostasomes in their extracellular context are thought to play a crucial role in fertilization.METHODS:Prostasomes were purified according to a well worked-out schedule from seminal plasmas obtained from human, canine, equine and bovine species. The various prostasomes were subjected to SDS-PAGE separation and protein banding patterns were compared. To gain knowledge of the prostasomal protein systems pertaining to prostasomes of four different species proteins were analyzed using a proteomic approach. An in vitro assay was employed to demonstrate ATP formation by prostasomes of different species.RESULTS:The SDS-PAGE banding pattern of prostasomes from the four species revealed a richly faceted picture with most protein bands within the molecular weight range of 10-150kDa. Some protein bands seemed to be concordant among species although differently expressed and the number of protein bands of dog prostasomes seemed to be distinctly fewer. Special emphasis was put on proteins involved in energy metabolic turnover. Prostasomes from all four species were able to form extracellular adenosine triphosphate (ATP). ATP formation was balanced by ATPase activity linked to the four types of prostasomes.CONCLUSION:These potencies of a possession of functional ATP-forming enzymes by different prostasome types should be regarded against the knowledge of ATP having a profound effect on cell responses and now explicitly on the success of the sperm cell to fertilize the ovum.GENERAL SIGNIFICANCE:This study unravels energy metabolic relationships of prostasomes from four different species.
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