Reactions to Participation in Research on Violence Experiences and Minority Stress Among Sexual and Gender Minority Young Adults.

Given the growing body of research seeking to examine adverse childhood experiences (ACEs) and intimate partner violence (IPV) among sexual and gender minority (SGM) individuals, Institutional Review Boards must consider whether participating in violence research is emotionally distressing for SGM people. Yet, little research has studied SGM participants' reactions to participating in research on ACEs, IPV, and minority stress. Thus, the current study examined reactions, including negative emotional reactions, to participating in violence research among SGM young adults. In total, 230 participants who self-identified as a sexual minority (30.1% also identified as a gender minority) in a dating relationship completed a cross-sectional assessment on ACEs, IPV (including identity abuse victimization and perpetration), minority stress (i.e., internalized homo/bi/transphobia), and reactions to research participation. Results indicated that participants identifying as a gender minority had significantly higher negative emotional reactions to study participation compared to cisgender participants, but this increase among gender minority individuals was small. In addition, gender minority participants and those with higher minority stress (i.e., internalized trans/bi/homo-negativity) and ACEs reported significantly higher negative emotional reactions to participation. Furthermore, gender minority participants scored worse on a scale indicating appreciation for contributing to research. Finally, reporting IPV victimization and perpetration was not associated with negative emotional reactions. Findings suggest that questions assessing minority stress and negative childhood experiences may be more emotionally salient or stressful for gender minority participants compared to questions measuring IPV.

Specific knockdown of OCT4 in human embryonic stem cells by inducible short hairpin RNA interference.

Manipulation of gene function in embryonic stem cells by either over expression or downregulation is critical for understanding their subsequent cell fate. We have developed a tetracycline-inducible short hairpin RNA interference (shRNAi) for human embryonic stem cells (hESCs) and demonstrated doxycycline dose-dependent knockdown of the transcription factor OCT4 and the cell surface antigen beta2-microglobulin. The induced knockdown of OCT4 resulted in rapid differentiation of hESCs with a significant increase in transcription of genes associated with trophoblast and endoderm lineages, the extent of which was controlled by the degree of induction. Transgene toxicity, which may occur in conditional over-expression strategies with hESCs, was not observed with wild-type Tet repressor protein. The system allows efficient, reversible, and long-term downregulation of target genes in hESCs and enables the generation of stable transfectants for the knockdown of genes essential for cell survival and self-renewal, not necessarily possible by nonconditional shRNAi methods. STEM CELLS 2009;27:776-782.

Sphingosine-1-phosphate mediates transcriptional regulation of key targets associated with survival, proliferation, and pluripotency in human embryonic stem cells.

Human embryonic stem cells (hESCs) replicate in vitro by the process of self-renewal, whilst maintaining their pluripotency. Understanding the pathways involved in the regulation of this process will assist in developing fully-defined conditions for the robust proliferation of hESCs necessary for therapeutic applications. We previously demonstrated that sphingosine-1-phosphate (S1P) plays an important role in survival and proliferation of hESCs. and here the key signaling pathways and downstream targets of S1P were investigated in a representative cell line (Shef 4). A significant rise in ERK1/2 activation with S1P treatment was witnessed in hESCs maintained on murine embryonic fibroblasts (MEFs) exhibiting significantly higher levels of active ERK1/2 than those grown on Matrigel. RT-PCR and microarray analysis of micro-dissected, non-differentiated hESC revealed 1049 differentially expressed genes in S1P treated preparations compared with controls (n = 3). S1P regulated apoptosis through several BCL-2 family members, including BAX and BID, with increased expression of cell cycle progression genes associated with proliferation of hESC cultures. S1P treatment also increased expression of cell adhesion genes specifically cadherins and integrins. However, gene expression associated with pluripotency was decreased with S1P treatment indicating that an increased rate of hESC turnover (higher proliferation and lower apoptosis) may be balanced by an increased susceptibility to differentiate.