Regulation of macromolecular modulators of urinary stone formation by reactive oxygen species: transcriptional study in an animal model of hyperoxaluria

Abstract

We used an unbiased approach of gene expression profiling to determine differential gene expression of all the macromolecular modulators (MMs) considered to be involved in stone formation, in hyperoxaluric rats, with and without treatment with the NADPH oxidase inhibitor apocynin. Male rats were fed rat chow or chow supplemented with 5% wt/wt hydroxy-l-proline (HLP) with or without apocynin-supplemented water. After 28 days, rats were euthanized and their kidneys explanted. Total RNA was isolated and microarray analysis was conducted using the Illumina bead array reader. Gene ontology analysis and the pathway analyses of the genes were done using Database for Annotation, Visualization of Integrated Discovery enrichment analysis tool. Quantitative RT-PCR of selected genes was carried out to verify the microarray results. Expression of selected gene products was confirmed using immunohistochemistry. Administration of HLP led to crystal deposition. Genes encoding for fibronectin, CD 44, fetuin B, osteopontin, and matrix-gla protein were upregulated while those encoding for heavy chains of inter-alpha-inhibitor 1, 3, and 4, calgranulin B, prothrombin, and Tamm-Horsfall protein were downregulated. HLP-fed rats receiving apocynin had a significant reversal in gene expression profiles: those that were upregulated came down while those that were downregulated stepped up. Apocynin treatment resulted in near complete absence of crystals. Clearly, there are two types of MMs; one is downregulated while the other is upregulated during hyperoxaluria and crystal deposition. Apparently gene and protein expressions of known macromolecular modulators of CaOx crystallization are likely regulated by ROS produced in part through the activation of NADPH oxidase.

Osteogenic changes in kidneys of hyperoxaluric rats

“…[60, 71, 87] Genes encoding for fibronectin, CD-44, fetuin B, osteopontin, and matrix-gla protein, are up-regulated while those encoding for heavy chains of inter-alpha-inhibitor 1, 3 and 4, calgranulin B, prothrombin, and Tamm-Horsfall protein were down-regulated. [88] Gene expression of vimentin, a mesenchymal marker, is also increased. [89]…”

“…Interestingly, the inflammatory molecules altered during CaOx nephrolithiasis and epithelial exposure to oxalate and CaOx crystals, [71, 87, 88] are also integral to the calcification cascade during both the physiological and pathological processes. [49] Production of OPN, FN, MGP, and collagens are indicators of osteoblastic character of the cells.…”

Unified theory on the pathogenesis of Randall’s plaques and plugs

“…[118,123] Renal epithelial cells under oxidative stress may become osteogenic [124] as happens to vascular smooth muscle cells during vascular calcification. [78] The production of OPN,[125,126] MGP,[127,128], collagen, fibronectin, osteonectin and fetuin, [129] by renal epithelial cells of rats with experimentally induced CaOx nephrolithiasis are indicative of such a transition. The presence of OPN, osteocalcin, fibronectin, and collagen [130] in stone matrices also suggests their increased production and excretion into the urine.…”

Evaluation of antiurolithiatic and antioxidant potential ofLepidagathis prostrata: APashanbhedplant

“…Reactive oxygen species (ROS) are known to perturb the oxidant-antioxidant balance in kidney cells resulting in cellular damage (Jyothilakshmi et al, 2014). Renal epithelial cells produce ROS when exposed to different crystals like CaOx, calcium phosphate and uric acid (Khan et al, 2014). Several macromolecular modulators of CaOx crystallization are also regulated by ROS.…”