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Thesis Defence

PhD degree for Sami Mohammed

PhD Sami 10 Sep 2018

Sami Mohammad successfully defended his thesis entitled "Going with the flow to elucidate renal electrolyte handling".


Outline of this thesis

External cues such as mechanical forces generated by fluid flow play a crucial role in the normal renal architecture and physiology. And since the discovery of primary cilia as a mechanosensor and failure-prone organelle, many researchers are trying to delineate its physiological role in the context of the different functions of the kidney including electrolyte handling. This thesis aims to understand the functional consequences of mechanosensation of urinary flow in the kidney with a special emphasis on electrolyte handling, and how the primary cilia fit in this process. Chapter 2 aims to identify the primary cilia-dependent transcriptome changes of renal epithelial cells in response to fluid shear stress using RNA-seq analysis on inner medullary collecting duct cells-3 (IMCD3) subjected to physiological relevant FSS. This study resulted in identifying pathways regulated by primary cilia bending due to flow. In Chapter 3, the effect of FSS on transepithelial Ca2+ transport in DCT and CNT was investigated. The DCT/CNT is very important in fine-tuning the final concentration of ions, such as Ca2+ and Mg2+, in serum and urine. Due to the absence of a representative DCT/CNT cell model, the study of FSS-mediated regulation of major electrolyte transporters such as TRPV5 was largely impeded. Here, we developed novel sorting technique to isolate DCT/CNT of transgenic mice. Isolated tubules were exposed to physiological shear stress with the help of a 2D rocker system. This chapter extended our knowledge of the flow-activated events that occur in DCT/CNT segments for maintaining Ca2+ balance. While in Chapter 4, the mechanism

underlying the effect of different types of fluid flow on the Ca2+ channel TRPV5 was investigated. Here the CRISPR/Cas9 gene editing tool was utilized to generate an IMCD3 cortical collecting duct cell line stably expressing TRPV5. Later, these cells were exposed to two types of fluid flow, oscillatory and unidirectional fluid flow, followed by functional analysis of TRPV5.

In the previous three chapters, we employed several in-vitro and ex-vivo techniques in our pursuit to study the role of FSS and primary cilia in renal electrolyte handling. Next, we sought to identify the clinical relevance of renal electrolyte reabsorption regulated by primary cilia. A hallmark of many human ciliopathies is the formation of kidney cysts, such as in PKD, which ultimately leads to renal failure. Mutations in PKD1 account for 80 to 85% of cases of ADPKD, while the remaining cases are ascribed to mutations in PKD2 (100). However, little is known about the implications of dysfunctional PKD1 on the renal electrolyte handling prior to cyst formation. The aim of Chapter 5 was, therefore, to assess the electrolyte balance in kidney-specific Pkd1 knockout mice with a pre-cystic phenotype. This study provided early clinical manifestations concerning electrolyte disturbance of PKD in patients. Finally, in Chapter 6 the findings of this thesis are summarized and discussed to augment our understanding of mechanosensation and primary cilia function in the kidney.

Click here to access Sami's thesis digitally.




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