Unknown,Transcriptomics,Genomics,Proteomics

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Microarray approach to investigate gene expression in human ovarian tissue after xenografting

ABSTRACT: Cryobanking and transplantation of ovarian tissue is a promising approach to restore fertility in cancer patients. However, ischemic stress following avascular ovarian cortex grafting is known to induce stromal tissue fibrosis and alteration in follicular development. The aim of the study is to analyse the impact of freezing-thawing and grafting procedures on gene expression in human ovarian tissue. Frozen-thawed ovarian tissue from 4 patients was xenografted for 7 days in nude mice and one non-grafted fragment was used as control. Immediately after recovery, grafts were processed for RNA extraction and histological analysis. Their expression profile was screened by whole-genome oligonucleotide array (n=4) and validated by reverse-transcriptase polymerase chain analysis (n=10). 84 of the transcripts were significantly altered after 7 days of grafting including matrix metalloproteinase-9 and -14 and angiogenic factors such as Placental growth factor and C-X-C chemokine receptor type 4 (CXCR4). Major biological processes were related to tissue remodelling including secretory processes, cellular adhesion and response to chemical and hormone stimuli. Angiopoietin signaling, interleukin-8 pathway and peroxisome proliferator-activated receptors activation were shown to be differentially regulated. On day 7, overexpression was confirmed by PCR for interleukin-8, transforming growth factor-beta 1, matrix metalloproteinase-14 and CXCR4 compared to the nongrafted control. In conclusion, new as well as known genes involved in tissue restructuration and angiogenesis were identified after human ovarian tissue transplantation. This will facilitate the development of strategies to optimize grafting techniques. In this study, ovarian tissue was obtained from 4 patients. Each biopsy was divided into 2 fragments. One fragment was frozen/thawed and placed in TRIzol reagent (Invitrogen) for RNA extraction. The other fragment was frozen-thawed and grafted in the intraperitoneal cavity of a nude mice. After 7 days graft were recovered and total RNA was extracted.

ORGANISM(S): Homo sapiens

SUBMITTER: Jérôme Ambroise

PROVIDER: E-GEOD-47555 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Phytophthora blight is a highly destructive soil borne disease caused by Phytophthora capsici Leonian, which seriously threatens global pepper production. Grafting is one of the important means to improve plant disease resistance and prevent soil borne diseases in vegetable production. However, the molecular mechanism by which grafting enhances the resistance of pepper to Phytophthora blight is still unclear. This study used Phytophthora capsici resistant strain ‘ZCM334’ and susceptible strain ‘Early Calwonder’ as rootstocks, and ‘Early Calwonder’ as scions for grafting. Phenotypic observation and cytological analysis showed that compared with the ‘Early Calwonder’ self rooted plants, the ‘ZCM334’ grafted plants had a later onset of disease, stronger resistance, and less damage to leaf cells, indicating that grafting can significantly improve the resistance of pepper to Phytophthora capsici. This study identified differentially expressed proteins (DEPs) in the leaves and roots of ‘ZCM334’ grafted plants and ‘Early Calwonder’ self rooted plants through proteomic analysis based on iTRAQ technology, and 478 and 349 DEPs were identified between their leaves and roots, respectively. These DEPs were mainly involved in metabolic process, cellular process, response to stimulus and catalytic activity processes. We identified and screened 12 DEPs with consistent expression trends in the leaves and roots of ‘ZCM334’ grafted plants and ‘Early Calwonder’ self rooted plants, including seven DEPs related to Phytophthora capsici resistance (CA01g31060, CA02g15780, CA02g30850, CA01g11410, CA05g12260, CA03g35150, and CA03g36980) and five proteins with unknown functions (CA01g26190, CA11g10620, CA12g02730, CA01g20890 and CA02g11340). Through qRT-PCR analysis, a significant correlation was observed between the protein and transcript levels of these 12 DEPs, and their expression characteristics were analyzed in the roots and leaves of ‘ZCM334’ grafted plants and ‘Early Calwonder’ self rooted plants at different stages (0h, 12h, 24h, and 36h) after inoculation with Phytophthora capsici. This study provides valuable information for exploring the molecular mechanisms by which grafting enhances the resistance of pepper to Phytophthora blight and the excavation of these key genes provides research ideas for studying the regulatory network of pepper resistance to Phytophthora capsici.

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Microarray approach to investigate gene expression in human ovarian tissue after xenografting

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