Reprogramming of Energy Metabolism in Human PKD1 Polycystic Kidney Disease: A Systems Biology Analysis.

Multiple alterations of cellular metabolism have been documented in experimental studies of autosomal dominant polycystic kidney disease (ADPKD) and are thought to contribute to its pathogenesis. To elucidate the molecular pathways and transcriptional regulators associated with the metabolic changes of renal cysts in ADPKD, we compared global gene expression data from human PKD1 renal cysts, minimally cystic tissues (MCT) from the same patients, and healthy human kidney cortical tissue samples. We found gene expression profiles of PKD1 renal cysts were consistent with the Warburg effect with gene pathway changes favoring increased cellular glucose uptake and lactate production, instead of pyruvate oxidation. Additionally, mitochondrial energy metabolism was globally depressed, associated with downregulation of gene pathways related to fatty acid oxidation (FAO), branched-chain amino acid (BCAA) degradation, the Krebs cycle, and oxidative phosphorylation (OXPHOS) in renal cysts. Activation of mTORC1 and its two target proto-oncogenes, HIF-1α and MYC, was predicted to drive the expression of multiple genes involved in the observed metabolic reprogramming (e.g., GLUT3, HK1/HK2, ALDOA, ENO2, PKM, LDHA/LDHB, MCT4, PDHA1, PDK1/3, MPC1/2, CPT2, BCAT1, NAMPT); indeed, their predicted expression patterns were confirmed by our data. Conversely, we found AMPK inhibition was predicted in renal cysts. AMPK inhibition was associated with decreased expression of PGC-1α, a transcriptional coactivator for transcription factors PPARα, ERRα, and ERRγ, all of which play a critical role in regulating oxidative metabolism and mitochondrial biogenesis. These data provide a comprehensive map of metabolic pathway reprogramming in ADPKD and highlight nodes of regulation that may serve as targets for therapeutic intervention.

Transcriptome Profile of the Rat Adrenal Gland: Parenchymal and Interstitial Cells.

The homeostasis of the adrenal gland plays a decisive role in its proper functioning, both in non-stressful conditions and under the influence of various types of stress. This consists of interactions between all types of cells that make up the organ, including parenchymal and interstitial cells. The amount of available information on this subject in the rat adrenal glands under non-stressful conditions is insufficient; the aim of the research was to determine the expression of marker genes for rat adrenal cells depending on their location. The material for the study consisted of adrenal glands taken from intact adult male rats that were separated into appropriate zones. Transcriptome analysis by means of Affymetrix® Rat Gene 2.1 ST Array was used in the study, followed by real-time PCR validation. Expression analysis of interstitial cell marker genes revealed both the amount of expression of these genes and the zone in which they were expressed. The expression of marker genes for fibroblasts was particularly high in the cells of the ZG zone, while the highest expression of specific macrophage genes was observed in the adrenal medulla. The results of this study, especially with regard to interstitial cells, provide a so far undescribed model of marker gene expression of various cells, both in the cortex and medulla of the sexually mature rat adrenal gland. The interdependence between parenchymal and interstitial cells creates a specific microenvironment that is highly heterogeneous within the gland with respect to some of the interstitial cells. This phenomenon most likely depends on the interaction with the differentiated parenchymal cells of the cortex, as well as the medulla of the gland.

Transcriptome Dynamics of Developing Photoreceptors in Three-Dimensional Retina Cultures Recapitulates Temporal Sequence of Human Cone and Rod Differentiation Revealing Cell Surface Markers and Gene Networks.

The derivation of three-dimensional (3D) stratified neural retina from pluripotent stem cells has permitted investigations of human photoreceptors. We have generated a H9 human embryonic stem cell subclone that carries a green fluorescent protein (GFP) reporter under the control of the promoter of cone-rod homeobox (CRX), an established marker of postmitotic photoreceptor precursors. The CRXp-GFP reporter replicates endogenous CRX expression in vitro when the H9 subclone is induced to form self-organizing 3D retina-like tissue. At day 37, CRX+ photoreceptors appear in the basal or middle part of neural retina and migrate to apical side by day 67. Temporal and spatial patterns of retinal cell type markers recapitulate the predicted sequence of development. Cone gene expression is concomitant with CRX, whereas rod differentiation factor neural retina leucine zipper protein (NRL) is first observed at day 67. At day 90, robust expression of NRL and its target nuclear receptor NR2E3 is evident in many CRX+ cells, while minimal S-opsin and no rhodopsin or L/M-opsin is present. The transcriptome profile, by RNA-seq, of developing human photoreceptors is remarkably concordant with mRNA and immunohistochemistry data available for human fetal retina although many targets of CRX, including phototransduction genes, exhibit a significant delay in expression. We report on temporal changes in gene signatures, including expression of cell surface markers and transcription factors; these expression changes should assist in isolation of photoreceptors at distinct stages of differentiation and in delineating coexpression networks. Our studies establish the first global expression database of developing human photoreceptors, providing a reference map for functional studies in retinal cultures.

Potential Marker Pathways in the Endometrium That May Cause Recurrent Implantation Failure.

The aim of this prospective cohort study was to identify altered biologic processes in the endometrium that may be potential markers of receptive endometrium in patients with repeated implantation failure (RIF) as compared with fertile controls. The study was conducted in a university-affiliated in vitro fertilization (IVF) gynecology clinic and molecular biology and genetics laboratory. Healthy fertile controls (n = 24) and patients with RIF (n = 24) were recruited. Window of implantation gene profiling associated with RIF was performed. Six hundred forty-one differentially expressed genes were identified, and 44 pathways were found enriched. Upon clustering of the enriched pathways, 9 representative pathways were established. The important pathways that were identified included circadian rhythm, pathways in cancer, proteasome, complement and coagulation cascades, citrate cycle, adherens junction, immune system and inflammation, cell cycle, and renin-angiotensin system. The involvement of the circadian rhythm pathway and other related pathways may alter the endometrium's functioning to ultimately cause RIF. Furthermore, we found that the pathogenesis of RIF was multifaceted and that numerous processes were involved. We believe that a better understanding of the underlying mechanisms of RIF will ultimately give rise to better treatment opportunities and to better outcomes in IVF.

Transcriptome Profile of Rat Adrenal Evoked by Gonadectomy and Testosterone or Estradiol Replacement.

Sex differences in adrenal cortex structure and function are well known in different species. In the rat, they are manifested as larger adrenal cortex and higher corticosterone secretion by females compared with males. These sex differences depend, among others, on functioning of the hypothalamic-pituitary-adrenal axis (HPA). In this aspect, it is widely accepted that testosterone exerts an inhibitory and estradiol stimulatory effect on the said axis. The molecular bases of these sex-related differences are poorly understood. Therefore, we performed studies aimed to demonstrate the effect of testosterone and estradiol on the expression of differentially regulated genes in rat adrenal gland. The classical method applied in the study-gonadectomy and gonadal hormone replacement-allows obtaining results suggesting a physiological role of the tested hormone (testosterone or estradiol) in the regulation of the specific genes. Adult male and female rats were either gonadectomized or sham operated. Half of orchiectomized rats were replaced with testosterone while ovariectomized ones with estradiol. Transcriptome was identified by means of Affymetrix® Rat Gene 2.1 ST Array. Differentially expressed genes were analyzed by means of DAVID web-based bioinformatic tools and confirmed by means of Gene Set Enrichment Analysis. For selected genes, validation of the results was performed using QPCR. Performed experiments have provided unexpected results. Contrary to expectations, in orchiectomized rats, testosterone replacement stimulates expression of numerous genes, mainly those associated with lipids and cholesterol metabolism. However, in ovariectomized animals, estradiol replacement inhibits the expression of genes, mainly those involved in intracellular signaling pathways. The physiological relevance of these findings awaits further research.