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A tissue retrieval and postharvest processing regimen for rodent reproductive tissues compatible with long-term storage on the international space station and postflight biospecimen sharing program.

Gupta V, Holets-Bondar L, Roby KF, Enders G, Tash JS - Biomed Res Int (2015)

Bottom Line: Collection and processing of tissues to preserve space flight effects from animals after return to Earth is challenging.Postfixation processing was also standardized for safe shipment back to our laboratory.Our strategy can be adapted for other tissues under NASA's Biospecimen Sharing Program or similar multi-investigator tissue sharing opportunities.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular & Integrative Physiology, University of Kansas Medical Center, Mail Stop 3050, 3901 Rainbow Boulevard, HLSIC 3098, Kansas City, KS 66160, USA.

ABSTRACT
Collection and processing of tissues to preserve space flight effects from animals after return to Earth is challenging. Specimens must be harvested with minimal time after landing to minimize postflight readaptation alterations in protein expression/translation, posttranslational modifications, and expression, as well as changes in gene expression and tissue histological degradation after euthanasia. We report the development of a widely applicable strategy for determining the window of optimal species-specific and tissue-specific posteuthanasia harvest that can be utilized to integrate into multi-investigator Biospecimen Sharing Programs. We also determined methods for ISS-compatible long-term tissue storage (10 months at -80°C) that yield recovery of high quality mRNA and protein for western analysis after sample return. Our focus was reproductive tissues. The time following euthanasia where tissues could be collected and histological integrity was maintained varied with tissue and species ranging between 1 and 3 hours. RNA quality was preserved in key reproductive tissues fixed in RNAlater up to 40 min after euthanasia. Postfixation processing was also standardized for safe shipment back to our laboratory. Our strategy can be adapted for other tissues under NASA's Biospecimen Sharing Program or similar multi-investigator tissue sharing opportunities.

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Mouse and gerbil ovarian and uterine horn histology. HE staining was used to evaluate quality of oocyte and follicles. Rows represent HE staining of: (a) mouse ovary up to 3 hr after euthanasia (4x objective, magnification bar is 500 μm); (b) mouse uteri up to 3 hr after euthanasia (10x objective, magnification bar is 500 μm); (c) gerbil ovary up to 1.5 hr after euthanasia (4x objective; magnification bar is 500 μm); (d) significantly high number of vacuoles are indicated in the yellow circles in gerbil ovaries from 1.5 h after euthanasia (40x objective, magnification bar is 100 μm); (e) gerbil uteri up to 2.5 hr after euthanasia (10x objective, magnification bar is 500 μm).
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fig3: Mouse and gerbil ovarian and uterine horn histology. HE staining was used to evaluate quality of oocyte and follicles. Rows represent HE staining of: (a) mouse ovary up to 3 hr after euthanasia (4x objective, magnification bar is 500 μm); (b) mouse uteri up to 3 hr after euthanasia (10x objective, magnification bar is 500 μm); (c) gerbil ovary up to 1.5 hr after euthanasia (4x objective; magnification bar is 500 μm); (d) significantly high number of vacuoles are indicated in the yellow circles in gerbil ovaries from 1.5 h after euthanasia (40x objective, magnification bar is 100 μm); (e) gerbil uteri up to 2.5 hr after euthanasia (10x objective, magnification bar is 500 μm).

Mentions: The ovaries (Figure 3(a)) and uteri (Figure 3(b)) harvested from female mice up to 3 hr after euthanasia showed excellent histological properties devoid of apparent tissue degradation.


A tissue retrieval and postharvest processing regimen for rodent reproductive tissues compatible with long-term storage on the international space station and postflight biospecimen sharing program.

Gupta V, Holets-Bondar L, Roby KF, Enders G, Tash JS - Biomed Res Int (2015)

Mouse and gerbil ovarian and uterine horn histology. HE staining was used to evaluate quality of oocyte and follicles. Rows represent HE staining of: (a) mouse ovary up to 3 hr after euthanasia (4x objective, magnification bar is 500 μm); (b) mouse uteri up to 3 hr after euthanasia (10x objective, magnification bar is 500 μm); (c) gerbil ovary up to 1.5 hr after euthanasia (4x objective; magnification bar is 500 μm); (d) significantly high number of vacuoles are indicated in the yellow circles in gerbil ovaries from 1.5 h after euthanasia (40x objective, magnification bar is 100 μm); (e) gerbil uteri up to 2.5 hr after euthanasia (10x objective, magnification bar is 500 μm).
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4309301&req=5

fig3: Mouse and gerbil ovarian and uterine horn histology. HE staining was used to evaluate quality of oocyte and follicles. Rows represent HE staining of: (a) mouse ovary up to 3 hr after euthanasia (4x objective, magnification bar is 500 μm); (b) mouse uteri up to 3 hr after euthanasia (10x objective, magnification bar is 500 μm); (c) gerbil ovary up to 1.5 hr after euthanasia (4x objective; magnification bar is 500 μm); (d) significantly high number of vacuoles are indicated in the yellow circles in gerbil ovaries from 1.5 h after euthanasia (40x objective, magnification bar is 100 μm); (e) gerbil uteri up to 2.5 hr after euthanasia (10x objective, magnification bar is 500 μm).
Mentions: The ovaries (Figure 3(a)) and uteri (Figure 3(b)) harvested from female mice up to 3 hr after euthanasia showed excellent histological properties devoid of apparent tissue degradation.

Bottom Line: Collection and processing of tissues to preserve space flight effects from animals after return to Earth is challenging.Postfixation processing was also standardized for safe shipment back to our laboratory.Our strategy can be adapted for other tissues under NASA's Biospecimen Sharing Program or similar multi-investigator tissue sharing opportunities.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular & Integrative Physiology, University of Kansas Medical Center, Mail Stop 3050, 3901 Rainbow Boulevard, HLSIC 3098, Kansas City, KS 66160, USA.

ABSTRACT
Collection and processing of tissues to preserve space flight effects from animals after return to Earth is challenging. Specimens must be harvested with minimal time after landing to minimize postflight readaptation alterations in protein expression/translation, posttranslational modifications, and expression, as well as changes in gene expression and tissue histological degradation after euthanasia. We report the development of a widely applicable strategy for determining the window of optimal species-specific and tissue-specific posteuthanasia harvest that can be utilized to integrate into multi-investigator Biospecimen Sharing Programs. We also determined methods for ISS-compatible long-term tissue storage (10 months at -80°C) that yield recovery of high quality mRNA and protein for western analysis after sample return. Our focus was reproductive tissues. The time following euthanasia where tissues could be collected and histological integrity was maintained varied with tissue and species ranging between 1 and 3 hours. RNA quality was preserved in key reproductive tissues fixed in RNAlater up to 40 min after euthanasia. Postfixation processing was also standardized for safe shipment back to our laboratory. Our strategy can be adapted for other tissues under NASA's Biospecimen Sharing Program or similar multi-investigator tissue sharing opportunities.

Show MeSH