LUCIANA LAMARAO DAMOUS
Projetos de Pesquisa
Unidades Organizacionais
LIM/58 - Laboratório de Ginecologia Estrutural e Molecular, Hospital das Clínicas, Faculdade de Medicina
5 resultados
Resultados de Busca
Agora exibindo 1 - 5 de 5
- Adipose tissue-derived stem cell therapy in rat cryopreserved ovarian grafts(2015) DAMOUS, Luciana Lamarao; NAKAMUTA, Juliana Sanajotti; CARVALHO, Ana Elisa Teofilo Saturi de; SOARES- JR., Jose Maria; SIMOES, Manuel de Jesus; KRIEGER, Jose Eduardo; BARACAT, Edmund C.The preliminary results of ovarian transplantation in clinical practice are encouraging. However, the follicular depletion caused by ischemic injury is a main concern and is directly related to short-term graft survival. Cell therapy with adipose tissue-derived stem cells (ASCs) could be an alternative to induce early angiogenesis in the graft. This study aimed to evaluate ASCs therapy in rat cryopreserved ovarian grafts. A single dose of rat ASC (rASCs) or vehicle was injected into the bilateral cryopreserved ovaries of twelve adult female rats immediately after an autologous transplant. Daily vaginal smears were performed for estrous cycle evaluation until euthanasia on postoperative day 30. Follicle viability, graft morphology and apoptosis were assessed. No differences were found with respect to estrous cycle resumption and follicle viability (P > 0.05). However, compared with the vehicle-treated grafts, the morphology of the ASCs-treated grafts was impaired, with diffuse atrophy and increased apoptosis (P < 0.05). ASCs direct injected in the stroma of rat cryopreserved ovarian grafts impaired its morphology although may not interfere with the functional resumption on short-term. Further investigations are necessary to evaluated whether it could compromise their viability in the long-term.
- Does adipose tissue-derived stem cell therapy improve graft quality in freshly grafted ovaries?(2015) DAMOUS, Luciana L.; NAKAMUTA, Juliana S.; CARVALHO, Ana E. T. Saturi de; CARVALHO, Katia Candido; SOARES- JR., Jose Maria; SIMOES, Manuel de Jesus; KRIEGER, Jose Eduardo; BARACAT, Edmund ChadaBackground: 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. Methods: Rat ASCs (rASCs) obtained from transgenic rats expressing green fluorescent protein (GFP)-(5 x 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-beta 1 was evaluated using RT-PCR and a double labeling immunofluorescence assay for GFP and Von Willebrand Factor (VWF) was performed. Results: 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. Conclusion: 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.
- Cell-free therapy with the secretome of adipose tissue-derived stem cells in rats' frozen-thawed ovarian grafts(2018) DAMOUS, Luciana Lamarao; CARVALHO, Ana Elisa Teofilo Saturi de; NAKAMUTA, Juliana Sanajotti; SHIROMA, Marcos Eiji; LOUZADA, Andressa Cristina Sposato; SOARES- JR., Jose Maria; KRIEGER, Jose Eduardo; BARACAT, Edmund C.The use of secretome may be a new strand of cell therapy, which is equal to or even superior to the injection of live cells, called cell-free therapy. In ovarian transplantation, this approach may be a therapeutic possibility for the ovarian graft in hypoxia. We designed the present study to evaluate whether the cell-free therapy with the secretome of adipose tissue-derived stem cells (ASCs) in rat frozen-thawed ovarian grafts could protect a graft against ischemic injury. A single dose of rat ASCs secretome or vehicle was injected into the bilateral frozen-thawed ovaries of 18 adult female rats immediately after an autologous transplant. Nine animals were used to control the cryopreservation protocol and were evaluated before and after the cryopreservation process. Daily vaginal smears were performed for estrous cycle evaluation until euthanasia on postoperative day 30. Follicle viability by trypan blue, graft morphology by HE, and apoptosis by TUNEL and cleaved-caspase-3 were assessed. No differences were found with respect to estrous cycle resumption and follicle viability (p > 0.05). However, compared with the vehicle-treated grafts, the morphology of the secretome-treated grafts was impaired, showing reduced follicular population and increased apoptosis (p < 0.05). ASC secretome impaired the rat frozen-thawed ovarian graft from ischemic injury. However, more studies are needed to evaluate the factors involved and the possibility of applying the secretome in scaffolds to optimize its use.
- Scaffold-based delivery of adipose tissue-derived stem cells in rat frozen-thawed ovarian autografts: preliminary studies in a rat model(2015) DAMOUS, Luciana Lamarao; NAKAMUTA, Juliana Sanajotti; CARVALHO, Ana Elisa Teofilo Saturi de; CARVALHO, Katia Candido; SOARES- JR., Jose Maria; SIMOES, Manuel de Jesus; KRIEGER, Jose Eduardo; BARACAT, Edmund C.This study aimed to evaluate whether a gelatin-based Gelfoam sponge is feasible as a scaffold for adipose tissue-derived stem cell (ASC) therapy in rat frozen-thawed ovarian autografts. Two sets of studies were performed. The in vitro set evaluated ASCs' viability in the Gelfoam scaffold at different times of co-culturing (after 24, 48, 72, 96, and 120 h). The in vivo set used 20 12-week-old adult female Wistar rats. Frozen-thawed ovarian grafts were treated with ASCs delivered in Gelfoam scaffolds immediately after an autologous retroperitoneal transplant (ASCs-GS, n = 10). The controls received Gelfoam with a culture medium (GS, n = 10). Assessment of graft quality was conducted by vaginal smears (until euthanasia on the 30th postoperative day), histological analyses, follicular density, and viability and fibrosis. Immunohistochemical staining for VEGF-A expression, vascular network (vWF), apoptosis (caspase-3 and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL)), cell proliferation (Ki-67), and hormone receptors (estrogen and progesterone) were performed. The cells remained viable in Gelfoam for up to 120 h of co-culturing. The graft morphology was similar among the groups. ASC therapy promoted the earlier resumption of the estrous phase (GS 16.6 +/- 3 vs. ASCs-GS 12.8 +/- 1.3 days) and enhanced estrogen receptors compared with the controls (p < 0.05) without interfering with the quantity and viability of the ovarian follicles, fibrosis, endothelial cells, VEGF immunoexpression, apoptosis, or cell proliferation (p > 0.05). The Gelfoam scaffold could be a feasible and safe non-invasive technique for ASC delivery in the treatment of frozen-thawed ovarian autografts. Future studies should evaluate the real benefit of this treatment on the survival and endocrine activity of the graft.
- Females transplanted with ovaries subjected to hypoxic preconditioning show impair of ovarian function(2014) DAMOUS, Luciana Lamarao; NAKAMUTA, Juliana Sanajotti; SOARES- JR., Jose Maria; MACIEL, Gustavo Arantes Rosa; SIMOES, Ricardo dos Santos; MONTERO, Edna Frasson de Souza; KRIEGER, Jose Eduardo; BARACAT, Edmund ChadaBackground: Cryopreservation of the ovarian tissue has shown promising results. However, there remain controversial issues such as the short half-life of grafts. In this aspect, there are some evidences that preconditioning the ovarian tissue before transplantation is beneficial. Objective: To determine the effect of hypoxic preconditioning in vitro on ovarian tissue prior to transplantation. Methods: Eighteen female adult Wistar rats, were sorted into three experimental groups. Ovaries were maintained in DMEM low glucose serum free at 37 degrees C with 5% CO2, at atmospheric oxigen concentration (normoxia) or 1% O-2 (hypoxia) for 16 hours. Oxigen concentration was determined by injection of nitrogen in the incubator. Animals submitted to ovarian transplantation immediately after oophorectomy were the Control Group (C). After this, the ovaries were implanted in the retroperitoneum with nonabsorbable suture and animals evaluated for thirty days after transplantation. Beginning on postoperative (PO) day 11, a daily collection of vaginal smear was carried out. Analyses comprised morphological, morphometric (counting ovarian follicles and corpora lutea) and immunohistochemistry for cleaved caspase-3 (apoptosis). Results: In normoxia and control groups all animals recovered their estrous cycles, while in the hypoxia group, two animals did not ovulate but, among those which did, resumption took longer than in the other groups (p < 0.05). The number of ovarian follicles and corpora lutea decreased significantly in the hypoxia group when compared to the other two groups (p < 0.001) and apoptosis was increased in the few ovarian follicles which remained viable (p < 0.001). Conclusion: The hypoxic preconditioning in vitro was not beneficial to the graft and worsened their viability, compromising its functionality or delaying the return of this.