Comparative Proteomics Analysis of Mouse Habu Nephritis Models with and without Unilateral Nephrectomy

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1 Original Paper 2016 The Author(s) Published The Author(s) by S. Karger AG, Basel Published online: October 17, 17, Published by S. Karger AG, Basel 1761 Accepted: August 02, 2016 This article is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License (CC BY-NC-ND) ( Usage and distribution Comparative Proteomics Analysis of Mouse Habu Nephritis Models with and without Unilateral Nephrectomy Lei Chen a,b Yang Lu b b b b b b Bo Fu b Zhong Yin b a b a b Department of Nephrology, Chinese PLA General Hospital, Chinese PLA Institute of Nephrology, State Key Laboratory of Kidney Diseases, National Clinical Research Center for Key Words Abstract Background/Aims: Individuals possessing a single kidney are at greater risk of renal injury upon exposure to harmful stimuli. This study aimed to explore the pathogenesis of renal injury Methods: Histological models, respectively). The role of villin 1, a differentially expressed protein (DEP) in mouse mesangial cells, was investigated. Results: Persistent mesangiolysis and focal hypercellularity together with reduced activation of cell proliferation in the HabuU model induced more serious renal injury compared with that in the Habu model. The DEPs between the two models enriched in the HabuU model. The cytoskeleton regulation protein villin 1 was downregulated in the HabuU model, but unchanged in the Habu model. Knockdown of villin 1 promoted apoptosis and inhibited the proliferation of mouse mesangial cells, suggesting villin 1 to be Conclusion: By assessing the proteomic 2016 The Author(s) Published by S. Karger AG, Basel Dept Nephrol, Chinese PLA General Hospital, Chinese PLA Institute of Nephrology, State Key Laboratory of Kidney Diseases, National Clinical Research Center for Kidney Diseases, Fuxing Road 28, Beijing , (PR China); Dialysis Dept of Nephrology Hospital, First /

2 1762 Introduction Individuals may possess a single kidney as a result of various physiological and pathological conditions, such as congenital solitary kidney, obstruction of the unilateral for tumors. Structural and functional changes in the remaining kidney are important concerns for nephrologists. The remaining kidney frequently shows compensatory hypertrophy to satisfy the increased workload and maintain normal kidney function by means of remaining kidney is at greater risk of glomerular disease upon exposure to harmful stimuli. kidney, with adriamycin nephrosis worsened interstitial nephrosis and aggravated the effective means of understanding the biology and the progression of renal disease in the glomerulonephritis in China and is characterized by mesangial cell proliferation accompanied underlying mechanism is unclear. and analyzed. Moreover, a proteomics method was used to screen differentially expressed underlying the changes following unilateral nephrectomy. By comparing the proteomic in the regulatory mechanisms underlying progressive renal disease following unilateral nephrectomy, which may facilitate the development of novel therapeutic strategies for patients possessing a single kidney. Methods and Materials Induction of glomerulonephritis

3 1763 blot analyses. Cardiac puncture was used for collection of blood samples. C were evaluated by light microscopy. The glomerular nucleated cell numbers were determined in thirty randomly selected glomeruli from the renal cortex of each mouse. To quantify glomerular mesangiolysis, from the renal cortex of each mouse were scored. min at room temperature to block endogenous peroxidase activity. The microwave antigen repair technique C. Then, the sections were reacted with a biotinylated secondary microscopy. The percentage of cells with condensed nuclei in each glomerulus was calculated to determine

4 1764 C cells terminated and prepared for subsequent assays.

5 1765 performed in at least triplicate. The percentage of cells with condensed nuclei among the total number of cells was calculated to determine the extent of apoptosis. The assay was performed in at least triplicate. post hoc Results creatinine and blood urea nitrogen levels. The serum creatinine levels were notably higher

6 1766 Fig. 1. Clinical fea tures of the two renal tissues ob tained at the in dicated days after glomerular nuc leated cell counts in the two models per mouse, me mesangiolysis in dex of renal tissu es obtained at the indicated days in the two models. per mouse, mean to glomerular structures, which contributed to serious deterioration of kidney function in Cell proliferation and apoptosis are the primary development and recovery mechanisms may result in deteriorated kidney function. model

7 1767 Fig. 2. Cell p r o l i f e r a tion and a p o p t o s i s in glomeru immunohis tochemistry of glomeruli and the ra p o s i t i v i t y at the indi cated days in the two mo group and staining and the apoptotic p p during the entire process of nephritis, which plays a crucial role in disease progression. Three model. Therefore, these shared pathways exerted differential effects in the two models. acids, butanoate metabolism, lysine degradation and tryptophan metabolism, were enriched progression after unilateral nephrectomy was evaluated.

8 1768 Fig. 3. Clas clusters accor ding to their expression pat downregulated ted only at day upregulated at regulated only gulated at days gulated only at gulated at days regulated only

9 1769 Fig. 4. pathway enrich ment analysis. selected using an The pathways en riched among the enriched among thways enriched The pathways en riched among the son of pathways between the two models.

10 1770 Fig. 5. The expression pat pathways shared by the two models and the two regulated and blue, down inhibited mesangial cell proliferation and promoted their apoptosis, which were associated Discussion model showed more severe renal injury, as indicated by higher levels of serum creatinine The pathological progression of mesangial proliferative nephritis is due to disturbances mesangiolysis. Therefore, the lack of recovery of mesangiolysis aggravated renal function

11 1771 To explore the molecular mechanisms in the two models, a proteomics analysis of glomeruli was performed. Based on in the two models. These pathways regulated energy metabolism during the entire process in both models. The disturbance of glycolysis and valine, leucine and isoleucine energy metabolism can affect cell activities these pathways were different between the two models, which suggests different effects on cell activities. The pathways unique to each model of unsaturated fatty acids and butanoate cytoskeleton regulation may contribute to activation of cell apoptosis and inhibition of cell proliferation during the pathogenesis of proximal tubular cells, whereas few studies have evaluated its expression in mesangial was abundant in glomeruli and mesangial its downregulation may be involved in mesangial proliferation inhibition and Table 1. # # of mesangiolysis via interfering with cell proliferation and promoting apoptosis.

12 1772 Fig. 6. The effect gulation on MMC proliferation and levels in glome expressed as fold changes compa group, which was to detect MMC apoptosis. The percentage of cells with con densed nuclei among the total number of cells represents the degree of apop corporation as say of MMC pro The percentage cells among the total number of cells represents the degree of p p performed in at least triplicate. Conclusion functions including aberrant energy metabolism, regulation of actin cytoskeleton, and

13 1773 Fig. 7. cells. Fig. 8. of kidney disease, particularly for patients with a single kidney. Abbreviations

14 1774 Acknowledgments Disclosure Statement The authors declare that they have no competing interests. References

15 1775 mechanism for resolution of glomerular hypercellularity in experimental mesangial proliferative nephritis.

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