Obstet Gynecol Sci.  2015 Sep;58(5):391-396. 10.5468/ogs.2015.58.5.391.

Expression of angiogenic factors in cryopreserved mouse ovaries after heterotopic autotransplantation

• Choi WJ
• Seok JS
• Choi IY
• Park JK
• Shin JK
• Lee SA
• Paik WY
• Lee JH

• KMID: 2058463
• DOI: http://doi.org/10.5468/ogs.2015.58.5.391

Abstract

OBJECTIVE
Revascularization is critical for successful ovarian tissue transplantation. Vascular endothelial growth factor (VEGF) and angiopoietin-2 (angpt-2) are the principal mediators of neovascularization. This study was designed to assess VEGF and angpt-2 levels in cryopreserved ovarian tissue after heterotopic autotransplantation.
METHODS
Ovarian tissues harvested from ICR mice at 5 to 6 weeks of age were stratified as follows: no cryopreservation (controls, group I); vitrification in VFS-40 (vitrification, group II); and gradual freezing in dimethyl sulfoxide (slow-freezing, group III). Frozen specimens were thawed at room temperature, assaying VEGF and angpt-2 levels 1 week after cryopreservation and 2 weeks after autotransplantation.
RESULTS
VEGF and angpt-2 protein levels were significantly lower in cryopreserved ovaries of groups II and III than in controls (group I, P<0.05), whereas groups II and III did not differ significantly in this regard. After autotransplantation of cryopreserved ovarian tissue, VEGF and angpt-2 protein levels did not differ significantly by technique but tended to be lower than corresponding levels in controls.
CONCLUSION
Expression of angiogenic factors in ovarian tissue is thought to vary by method of cryopreservation. Our findings indicate that levels of angiogenic factors expressed in cryopreserved ovarian tissue after autotransplantation do not differ appreciably from control levels, regardless of cryopreservation technique.

Keyword

Angiopoietin-2; Cryopreservation; Ovary; Transplantation; Vascular endothelial growth factor

MeSH Terms

Angiogenesis Inducing Agents*
Angiopoietin-2
Animals
Autografts*
Cryopreservation
Dimethyl Sulfoxide
Female
Freezing
Mice*
Mice, Inbred ICR
Ovary*
Tissue Transplantation
Transplantation
Transplants
Vascular Endothelial Growth Factor A
Vitrification
Angiogenesis Inducing Agents
Angiopoietin-2
Dimethyl Sulfoxide
Vascular Endothelial Growth Factor A

Figure

• 

  Fig. 1 Flow diagram of experimental design: After 1 week of cryopreservation (vitrification or slow freezing), one of the paired mouse ovaries was analyzed for angiogenic factor expression; the other ovary was autotransplanted beneath the abdominal wall. After a 2-week autotransplantation period, angiogenic factors were assayed.

• 

  Fig. 2 Autotransplanted mouse ovary: (A) beneath the abdominal wall (arrow), to be assayed after 2 weeks, and (B) on dish with media.

• 

  Fig. 3 Western blot analysis: quantitative analysis by group of (A) vascular endothelial growth factor (VEGF) and (B) angiopoietin-2 (angpt-2) after 1 week of vitrification or slow freezing (no cryopreservation of controls). β-actin was used as internal standard. Data were expressed as mean±standard error of three independent experiments. *P<0.05 vs. cryopreservation groups (vitrification and slow freezing).

• 

  Fig. 4 Western blot analysis: quantitative analysis by group of (A) vascular endothelial growth factor (VEGF) and (B) angiopoietin-2 (angpt-2) after 2 weeks of autotransplantation beneath the abdominal wall (no cryopreservation of controls). β-actin was used as internal standard. Data expressed as mean±standard error of three independent experiments.

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