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Genes (Basel)
2021 Aug 23;128:. doi: 10.3390/genes12081292.
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Expression of Piwi, MMP, TIMP, and Sox during Gut Regeneration in Holothurian Eupentacta fraudatrix (Holothuroidea, Dendrochirotida).
Dolmatov IY
,
Kalacheva NV
,
Tkacheva ES
,
Shulga AP
,
Zavalnaya EG
,
Shamshurina EV
,
Girich AS
,
Boyko AV
,
Eliseikina MG
.
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Mesodermal cells of holothurian Eupentacta fraudatrix can transdifferentiate into enterocytes during the regeneration of the digestive system. In this study, we investigated the expression of several genes involved in gut regeneration in E. fraudatrix. Moreover, the localization of progenitor cells of coelomocytes, juvenile cells, and their participation in the formation of the luminal epithelium of the digestive tube were studied. It was shown that Piwi-positive cells were not involved in the formation of the luminal epithelium of the digestive tube. Ef-72 kDa type IV collagenase and Ef-MMP16 had an individual expression profile and possibly different functions. The Ef-tensilin3 gene exhibited the highest expression and indicates its potential role in regeneration. Ef-Sox9/10 and Ef-Sox17 in E. fraudatrix may participate in the mechanism of transdifferentiation of coelomic epithelial cells. Their transcripts mark the cells that plunge into the connective tissue of the gut anlage and give rise to enterocytes. Ef-Sox9/10 probably controls the switching of mesodermal cells to the enterocyte phenotype, while Ef-Sox17 may be involved in the regulation of the initial stages of transdifferentiation.
Figure 1. Phylogenetic tree showing the relationships of the Piwi sequence of E. fraudatrix (marked with the asterisk) with homolog proteins of other animals.
Figure 2. Phylogenetic tree showing the relationships of the tensilins of E. fraudatrix with homolog proteins of the other holothurians. The tensilin used in the study is marked with red.
Figure 3. Part of the phylogenetic tree showing the relationships of Sox of the E. fraudatrix with homolog proteins of other animals. Ef-Sox9/10 is marked with a blue star, and Ef-Sox17 is marked with a red star.
Figure 4. mRNA expression profile of Piwi during different days post-evisceration and in normal intestinal tissue in E. fraudatrix. Different lowercase letters indicate significant differences (p < 0.05). The data are reported as the means ± S.D. (n = 5).
Figure 5. Localization of Piwi-positive juvenile cells in tissues of E. fraudatrix. (a) Labeled cell in the coelom at 4 hpe. (b) Labeled cells (arrowheads) in the coelomic epithelium at 4 hpe. (c) Labeled cell in the dermis of the body wall at 4 hpe. (d) Numerous labeled cells in the hypodermis at 4 hpe. (e) Rare-labeled cells (arrowheads) in the hypodermis at 24 hpe. (f) General view of the gut anlage at 7 dpe. (g) Labeled cells in the connective tissue of the gut anlage at 7 dpe. g, gut anlage; m, mesentery. Immunocytochemical staining with antibodies for the PIWI protein (red color) and DAPI-stained nuclear DNA (blue color).
Figure 6. Expression of MMPs during gut regeneration. (a) 72 kDa type IV collagenase expression in forming luminal epithelium of the gut on 5–7 dpe (histological section). (b) 72 kDa type IV collagenase expression in the coelomic epithelium of the mesentery and connective tissue thickening of the gut anlage on 5–7 dpe (histological section). (c) 72 kDa type IV collagenase expression in the posterior part of the gut on 10 dpe (whole mount). (d) 72 kDa type IV collagenase expression in the coelomic epithelium of the mesentery and gut anlage on 10 dpe (histological section). (e) Expression of MMP16 in the anterior part of gut anlage on 5–7 dpe (histological section). (f) Expression of MMP16 in the coelomic epithelium of the posterior part of the mesentery and connective tissue thickening on 5–7 dpe (histological section). (g) Expression of MMP16 in the coelomic epithelium of the mesentery and gut on 10 dpe (histological section). ab, aquapharyngeal bulb; ce, coelomic epithelium; ct, connective tissue; ctt, connective tissue thickening; g, gut; le, luminal epithelium; and m, mesentery; the insets in (b,d,f) show higher magnification views of the boxed areas.
Figure 7. Scheme of spatial distribution of the 72 kDa type IV collagenase, MMP16, tensilin3, Sox9/10, and Sox17 transcripts on 5–7 and 10 dpe. (a–c): Dotted lines indicate the planes of the gut anlage cut and a1–c3: sections of the gut anlage on the corresponding planes; ab, aquapharyngeal bulb; ga, gut anlage; le, luminal epithelium of the gut; and m, mesentery; an arrowhead indicates a site of coelomic epithelium embedding.
Figure 8. Expression of tensilin-3 during gut regeneration. (a) Tensilin-3 expression in the ventral part of the mesentery and gut anlage on 5–7 dpe (whole mount). (b) Tensilin-3 expression in the ventral part of the gut anlage on 5–7 dpe (histological section). (c) Tensilin-3 expression in the growing end of the gut on 10 dpe (whole mount). (d) Tensilin-3 expression in the coelomic and luminal epithelia of the gut anlage on 10 dpe (histological section). (e) Tensilin-3 expression in the ventral part of the growing end of the gut on 10 dpe (histological section). ab, aquapharyngeal bulb; ce, coelomic epithelium; ct, connective tissue; g, gut; le, luminal epithelium; and m, mesentery.
Figure 9. Expression of Sox9/10 and Sox17 during gut regeneration. (a) Expression of Sox9/10 in the coelomic and luminal epithelia of the gut anlage on 5–7 dpe; an arrowhead indicates a site of coelomic epithelium embedding, and arrows show the luminal epithelium (whole mount). (b) Expression of Sox9/10 in the coelomic and luminal epithelia of the gut anlage in the site of coelomic epithelium embedding (arrowhead) on 5–7 dpe (histological section). (c) Expression of Sox9/10 in the luminal epithelium of the posterior part of the gut anlage on 5–7 dpe (histological section). (d) Expression of Sox9/10 in the gut on 10 dpe (whole mount). (e) Expression of Sox9/10 in the luminal epithelium in the middle part of the gut anlage on 10 dpe (histological section). (f) Expression of Sox17 in the ventral part (arrowhead) of the gut anlage on 5–7 dpe (whole mount). (g) Expression of Sox17 in the coelomic epithelium in the site of embedding on 5–7 dpe; red spots indicate the site of the epithelium embedding, and arrowheads in the insert show the embedding epithelium (histological section). (h) Expression of Sox17 in the coelomic epithelium of the lateral and dorsal parts of the gut on 10 dpe (histological section). ab, aquapharyngeal bulb; ce, coelomic epithelium; ct, connective tissue; g, gut; le, luminal epithelium; and m, mesentery; the inset in (g) shows a higher magnification view of the boxed area.