Publications

Rescue of platinum-damaged oocytes from programmed cell death through inactivation of the p53 family signaling network.

Cell Death Differ. 2013 Apr 19;

Authors: Kim SY, Cordeiro MH, Serna VA, Ebbert K, Butler LM, Sinha S, Mills AA, Woodruff TK, Kurita T

Abstract

Non-proliferating oocytes within avascular regions of the ovary are exquisitely susceptible to chemotherapy. Early menopause and sterility are unintended consequences of chemotherapy, and efforts to understand the oocyte apoptotic pathway may provide new targets for mitigating this outcome. Recently, the c-Abl kinase inhibitor imatinib mesylate (imatinib) has become the focus of research as a fertoprotective drug against cisplatin. However, the mechanism by which imatinib protects oocytes is not fully understood, and reports of the drug's efficacy have been contradictory. Using in vitro culture and subrenal grafting of mouse ovaries, we demonstrated that imatinib inhibits the cisplatin-induced apoptosis of oocytes within primordial follicles. We found that, before apoptosis, cisplatin induces c-Abl and TAp73 expression in the oocyte. Oocytes undergoing apoptosis showed downregulation of TAp63 and upregulation of Bax. While imatinib was unable to block cisplatin-induced DNA damage and damage response, such as the upregulation of p53, imatinib inhibited the cisplatin-induced nuclear accumulation of c-Abl/TAp73 and the subsequent downregulation of TAp63 and upregulation of Bax, thereby abrogating oocyte cell death. Surprisingly, the conditional deletion of Trp63, but not ΔNp63, in oocytes inhibited apoptosis, as well as the accumulation of c-Abl and TAp73 caused by cisplatin. These data suggest that TAp63 is the master regulator of cisplatin-induced oocyte death. The expression kinetics of TAp63, c-Abl and TAp73 suggest that cisplatin activates TAp63-dependent expression of c-Abl and TAp73 and, in turn, the activation of TAp73 by c-Abl-induced BAX expression. Our findings indicate that imatinib protects oocytes from cisplatin-induced cell death by inhibiting c-Abl kinase, which would otherwise activate TAp73-BAX-mediated apoptosis. Thus, imatinib and other c-Abl kinase inhibitors provide an intriguing new way to halt cisplatin-induced oocyte death in early follicles and perhaps conserve the endocrine function of the ovary against chemotherapy.Cell Death and Differentiation advance online publication, 19 April 2013; doi:10.1038/cdd.2013.31.

PMID: 23598363 [PubMed - as supplied by publisher]

Imaging and elemental mapping of biological specimens with a dual-EDS dedicated scanning transmission electron microscope.

Ultramicroscopy. 2013 Feb 4;128C:24-31

Authors: Wu JS, Kim AM, Bleher R, Myers BD, Marvin RG, Inada H, Nakamura K, Zhang XF, Roth E, Li SY, Woodruff TK, O'Halloran TV, Dravid VP

Abstract

A dedicated analytical scanning transmission electron microscope (STEM) with dual energy dispersive spectroscopy (EDS) detectors has been designed for complementary high performance imaging as well as high sensitivity elemental analysis and mapping of biological structures. The performance of this new design, based on a Hitachi HD-2300A model, was evaluated using a variety of biological specimens. With three imaging detectors, both the surface and internal structure of cells can be examined simultaneously. The whole-cell elemental mapping, especially of heavier metal species that have low cross-section for electron energy loss spectroscopy (EELS), can be faithfully obtained. Optimization of STEM imaging conditions is applied to thick sections as well as thin sections of biological cells under low-dose conditions at room and cryogenic temperatures. Such multimodal capabilities applied to soft/biological structures usher a new era for analytical studies in biological systems.

PMID: 23500508 [PubMed - as supplied by publisher]

Follicle microenvironment-associated alterations in gene expression in the mouse oocyte and its polar body.

Fertil Steril. 2013 Jan 8;

Authors: Jiao ZX, Woodruff TK

Abstract

OBJECTIVE: To determine whether the follicle environment modulates oocyte-specific gene transcript levels in cultured oocytes and polar bodies (PBs). DESIGN: Animal study. SETTING: Large academic research center. ANIMAL(S): CD1 mice. INTERVENTION(S): In vitro growth of secondary mouse follicles in 0.25% or 1.5% alginate (ALG) in a three-dimensional culture system. MAIN OUTCOME MEASURE(S): Relative transcript levels of Gdf9, Bmp15, Nlrp5, Tcl1, and Zp3 were measured by real-time quantitative reverse transcriptase-polymerase chain reaction in oocytes during in vitro follicle development and oocyte maturation and in their first PBs after removal from metaphase II (MII) eggs. RESULT(S): All transcripts decreased earlier in oocytes cultured in 1.5% ALG compared with 0.25% ALG. Transcript levels were lower in MII eggs cultured in 1.5% ALG compared with in 0.25% ALG. All genes were expressed in PBs, and transcript levels were lower in PBs cultured in 1.5% ALG compared with in 0.25% ALG. Abundance of all transcripts was lower in PBs than in their sibling oocytes. CONCLUSION(S): Local follicle environment modulates oocyte-specific gene expression in the oocyte and first PB. There is a significant difference in the transcript levels of oocyte-specific genes in PBs of 1.5% versus 0.25% ALG that correlates with ovarian environment-related decreases in oocyte competence.

PMID: 23312223 [PubMed - as supplied by publisher]

Microarray analysis identifies COMP as the most differentially regulated transcript throughout in vitro follicle growth.

Mol Reprod Dev. 2012 Dec 14;

Authors: Skory RM, Bernabé BP, Galdones E, Broadbelt LJ, Shea LD, Woodruff TK

Abstract

In vitro follicle growth has emerged as a technology that can provide new information about folliculogenesis and serve as part of a suite of methods currently under development to assist women whose fertility is threatened by cancer treatments. Though it has been shown that in vitro-grown follicles secrete peptide and steroid hormones, much of the follicular transcriptome remains unknown. Thus, microarray analysis was performed to characterize the transcriptome and secretome of in vitro-grown follicles. One prominently regulated gene product was cartilage oligomeric matrix protein (Comp): its mRNA was upregulated during the final four days of culture (P < 0.05), and COMP protein could be detected in medium from individual follicles. COMP expression localized to mural granulosa cells of large antral follicles both in vitro and in vivo, with maximal expression immediately preceding ovulation in cycling and chorionic gonadotropin-primed female mice. COMP was co-expressed with two known markers of follicle maturation, inhibin β(A) and gremlin, and was expressed only in TUNEL-negative follicles. In addition to other gene products identified in the microarray, COMP has potential utility as a marker of follicle maturation. Mol. Reprod. Dev. © 2012 Wiley Periodicals, Inc.

PMID: 23242557 [PubMed - as supplied by publisher]