title: Does adipose tissue-derived stem cell therapy improve graft quality in freshly grafted ovaries?
abstract: A major concern in ovarian transplants is substantial follicle loss during the initial period of hypoxia. Adipose tissue-derived stem cells (ASCs) have been employed to improve angiogenesis when injected into ischemic tissue. This study evaluated the safety and efficacy of adipose tissue-derived stem cells (ASCs) therapy in the freshly grafted ovaries 30 days after injection.
  
  
  
  Rat ASCs (rASCs) obtained from transgenic rats expressing green fluorescent protein (GFP)-(5 × 10(4) cells/ovary) were injected in topic (intact) or freshly grafted ovaries of 30 twelve-week-old adult female Wistar rats. The whole ovary was grafted in the retroperitoneum without vascular anastomosis, immediately after oophorectomy. Vaginal smears were performed daily to assess the resumption of the estrous cycle. Estradiol levels, grafts morphology and follicular viability and density were analyzed. Immunohistochemistry assays were conducted to identify and quantify rASC-GFP(+), VEGF tissue expression, apoptosis (cleaved caspase-3 and TUNEL), and cell proliferation (Ki-67). Quantitative gene expression (qPCR) for VEGF-A, Bcl2, EGF and TGF-β1 was evaluated using RT-PCR and a double labeling immunofluorescence assay for GFP and Von Willebrand Factor (VWF) was performed.
  
  
  
  Grafted ovaries treated with rASC-GFP(+) exhibited earlier resumption of the estrous phase (p < 0.05), increased VEGF-A expression (11-fold in grafted ovaries and 5-fold in topic ovaries vs. control) and an increased number of blood vessels (p < 0.05) in ovarian tissue without leading to apoptosis or cellular proliferation (p > 0.05). Estradiol levels were similar among groups (p > 0.05). rASC-GFP(+) were observed in similar quantities in the topic and grafted ovaries (p > 0.05), and double-labeling for GFP and vWF was observed in both injected groups.
  
  
  
  rASC therapy in autologous freshly ovarian grafts could be feasible and safe, induces earlier resumption of the estrous phase and enhances blood vessels in rats. This pilot study may be useful in the future for new researches on frozen-thawed ovarian tissue.
authors.affiliation: Laboratório de Ginecologia Estrutural e Molecular (LIM-58), Disciplina de Ginecologia, Departamento de Obstetrícia e Ginecologia, Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo, Dr Arnaldo av 455, 2nd floor, room 2113, Pacaembu, 01246-903, São Paulo, Brazil. lucianadamous@gmail.com.
authors.forename: Luciana L
authors.initials: LL
authors.lastname: Damous
authors.affiliation: Laboratory of Genetics and Molecular Cardiology, Heart Institute (Incor), Faculdade de Medicina da Universidade de São Paulo, Dr Enéas de Carvalho Aguiar Av 44, 10th floor, Cerqueira Cesar, 05403-000, São Paulo, Brazil. juliana.nakamuta@gmail.com.
authors.forename: Juliana S
authors.initials: JS
authors.lastname: Nakamuta
authors.affiliation: Laboratory of Genetics and Molecular Cardiology, Heart Institute (Incor), Faculdade de Medicina da Universidade de São Paulo, Dr Enéas de Carvalho Aguiar Av 44, 10th floor, Cerqueira Cesar, 05403-000, São Paulo, Brazil. anaelisafb@yahoo.com.br.
authors.forename: Ana E T Saturi de
authors.initials: AE
authors.lastname: Carvalho
authors.affiliation: Laboratório de Ginecologia Estrutural e Molecular (LIM-58), Disciplina de Ginecologia, Departamento de Obstetrícia e Ginecologia, Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo, Dr Arnaldo av 455, 2nd floor, room 2113, Pacaembu, 01246-903, São Paulo, Brazil. carvalhokc@gmail.com.
authors.forename: Katia Candido
authors.initials: KC
authors.lastname: Carvalho
authors.affiliation: Laboratório de Ginecologia Estrutural e Molecular (LIM-58), Disciplina de Ginecologia, Departamento de Obstetrícia e Ginecologia, Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo, Dr Arnaldo av 455, 2nd floor, room 2113, Pacaembu, 01246-903, São Paulo, Brazil. jsoares415@hotmail.com.
authors.forename: José Maria
authors.initials: JM
authors.lastname: Soares
authors.affiliation: Department of Morphology and Genetics, Universidade Federal de São Paulo (UNIFESP), Botucatu St 740. Ed. Lemos Torres, 2nd floor, Vila Clementino, 04023-009, São Paulo, Brazil. manuel.morf@epm.br.
authors.forename: Manuel de Jesus
authors.initials: Mde J
authors.lastname: Simões
authors.affiliation: Laboratory of Genetics and Molecular Cardiology, Heart Institute (Incor), Faculdade de Medicina da Universidade de São Paulo, Dr Enéas de Carvalho Aguiar Av 44, 10th floor, Cerqueira Cesar, 05403-000, São Paulo, Brazil. krieger@incor.usp.br.
authors.forename: José Eduardo
authors.initials: JE
authors.lastname: Krieger
authors.affiliation: Laboratório de Ginecologia Estrutural e Molecular (LIM-58), Disciplina de Ginecologia, Departamento de Obstetrícia e Ginecologia, Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo, Dr Arnaldo av 455, 2nd floor, room 2113, Pacaembu, 01246-903, São Paulo, Brazil. ecbaracat@usp.br.
authors.forename: Edmund Chada
authors.initials: EC
authors.lastname: Baracat
mesh_terms: D017667:Adipocytes / Q000637:transplantation*; D000818:Animals; D017209:Apoptosis / Q000502:physiology; D053148:Caspase 3 / Q000378:metabolism; D049109:Cell Proliferation; D030762:Estrous Cycle; D005260:Female; D006080:Ovarian Follicle / Q000378:metabolism / Q000637:transplantation*; D010053:Ovary / Q000378:metabolism / Q000637:transplantation*; D010865:Pilot Projects; D051381:Rats; D055647:Rats, Transgenic; D017208:Rats, Wistar; D033581:Stem Cell Transplantation; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism
chemical_list: D042461:Vascular Endothelial Growth Factor A; D053148:Caspase 3