More than just hormones: H295R cells as predictors of reproductive toxicity

Abstract

Many of the commonly observed reproductive toxicities associated with therapeutic compounds can be traced to a disruption of the steroidogenic pathway. We sought to develop an in vitro assay that would predict reproductive toxicity and be high throughput in nature. H295R cells, previously validated as having an intact and functional steroidogenic pathway, were treated with 83 known-positive and 79 known-negative proprietary and public-domain compounds. The assay measured the expression of the key enzymes STAR, 3βHSD2, CYP17A1, CYP11B2, CYP19A1, CYP21A2, and CYP11A1 and the hormones DHEA, progesterone, testosterone, and cortisol. We found that a Random Forest model yielded a receiver operating characteristic area under the curve (ROC AUC) of 0.845, with sensitivity of 0.724 and specificity of 0.758 for predicting in vivo reproductive toxicity with this in vitro assay system.

Stimulus-selective induction of the orphan nuclear receptor NGFIB underlies different influences of angiotensin II and potassium on the human adrenal gland zona glomerulosa-specific 3β-HSD isoform gene expression in adrenocortical H295R cells

“…Forskolin treatment, on the other hand, could increase expression of HSD3B2 (Fig. 2B) as previously suggested [9][10][11][12][13]. Thus, the responsiveness of this gene relies on the type of agonists administered to the cells.…”

“…Where specified, cycloheximide (CHX; final concentration, 10 µg/ mL) was added to the medium 15 min before AngII or K + treatment. For the analysis of serum-starved cells, cells were cultured as described above except that the medium was replaced with DMEM/F-12 serum-free medium for the last 24 h. Forskolin (cAMP activator) treatment, which has been considered to mimic ACTH stimulation, was also performed to confirm a previously reported response of HSD3B2 to the ACTH signal in H295R cells [9][10][11][12][13].…”

WITHDRAWN: Exposure to Difenoconazole Induces Reproductive Toxicity of Zebrafish by Interfering Reproductive Behavior

“…In the gonads (Fig. 6), E2 and T are produced through a series of enzymatic reactions with cholesterol as the precursor material (Maglich et al, 2014;Yang et al, 2018). In our study, DCZ exposure signi cantly altered the expression of genes (cyp11a, cyp19a, hsd17b, and hsd3b) related to key steroidogenic enzymes.…”

Disruption of steroidogenesis: Cell models for mechanistic investigations and as screening tools

“…In contrast to the testicular and ovarian cell models, there is a human adrenal cell model (H295R) that has been recognized by the regulators for toxicity screening and resulted in an OECD test guideline for the evaluation of EDCs [169]. Therefore, a large number of studies applied the H295R cell model for the assessment of chemicals that cause disturbances of steroidogenesis, including pharmaceuticals, consumer products, food constituents and environmental pollutants [191][192][193][194][195][196][197][198], and it is out of the scope of this review to cover the findings of these studies. Currently, the OECD guideline only focuses on the use of H295R cells in their basal state and on the production of estradiol and testosterone as endpoints [199], two hormones not typically produced by the adrenals.…”