GATA4 Regulates Blood-Testis Barrier Function and Lactate Metabolism in Mouse Sertoli Cells

Abstract

Conditional deletion of Gata4 in Sertoli cells (SCs) of adult mice has been shown to increase permeability of the blood-testis barrier (BTB) and disrupt spermatogenesis. To gain insight into the molecular underpinnings of these phenotypic abnormalities, we assessed the impact of Gata4 gene silencing in cell culture models. Microarray hybridization identified genes dysregulated by siRNA-mediated inhibition of Gata4 in TM4 cells, an immortalized mouse SC line. Differentially expressed genes were validated by quantitative RT-PCR analysis of primary cultures of Gata4(flox/flox) mouse SCs that had been subjected to cre-mediated recombination in vitro. Depletion of GATA4 in TM4 cells and primary SCs was associated with altered expression of genes involved in key facets of BTB maintenance, including tight/adherens junction formation (Tjp1, Cldn12, Vcl, Tnc, Csk) and extracellular matrix reorganization (Lamc1, Col4a1, Col4a5, Mmp10, Mmp23, Timp2). Western blotting and immunocytochemistry demonstrated reduced levels of tight junction protein-1, a prototypical tight junction protein, in GATA4-depleted cells. These changes were accompanied by a loss of morphologically recognizable junctional complexes and a decline in epithelial membrane resistance. Furthermore, Gata4 gene silencing was associated with altered expression of Hk1, Gpi1, Pfkp, Pgam1, Gls2, Pdk3, Pkd4, and Ldhb, genes regulating the production of lactate, a key nutrient that SCs provide to developing germ cells. Comprehensive metabolomic profiling demonstrated impaired lactate production in GATA4-deficient SCs. We conclude that GATA4 plays a pivotal role in the regulation of BTB function and lactate metabolism in mouse SCs.

Probing GATA factor function in mouse Leydig cells via testicular injection of adenoviral vectors

“…We found no immunohistological evidence of caspase-3 activation in our model, and TUNEL staining was negative. In cultured gonadal somatic cell lines, silencing of Gata4 impairs glycolysis and causes other metabolic derangements (Schrade et al 2015, Schrade et al 2016). Thus, it is conceivable that altered metabolism contributes to Leydig cell death in our adenoviral injection model.…”

The Cellular Impact of the ZIKA Virus on Male Reproductive Tract Immunology and Physiology

“…The process of male mature gamete production is called spermatogenesis and consists of the intense proliferation and subsequent differentiation of spermatogonial stem cells to spermatozoa [25]. The crosstalk between constituent cells of the testis is essential in this process [36,37], once the energy source of gametes during differentiation depends on the lactate that is provided by SC. On the other hand, glucose capitation depends on androgen hormone signalization provided by LC, as well as pituitary hormones, insulin sensibility, and paracrine communication [36,37].…”

Validation and automation of a high-throughput multi-targeted method for semi-quantification of endogenous metabolites from different biological matrices using tandem mass spectrometry

“…We have applied our fully validated analytical methodology in various international and national biomedical research projects, epidemiological studies, clinical studies including dietary interventions, and clinical trials. We have successfully implemented our technology in the following research fields, including but not limited to: mitochondrial metabolism/disorders [11,17–23], cancer [24,25], bone metabolism [12], endocrinology [26,27], psychiatric disorders [28], inflammatory bowel disease [29], viral infections [30–33], allergies [34,35], circadian rhythms [36], and pain research [37].…”