Opportunities and challenges of Nephrin research (Part 1)

Unexplained pFSGS

Recent studies have shown that anti-Nephrin autoantibodies (Anti-nephrin Ab) play an important role in the occurrence of minimal change disease (MCD) and focal segmental glomerulosclerosis (FSGS). This article outlines the research progress of anti-Nephrin antibodies as blood circulation penetration factors in MCD and primary FSGS (pFSGS), and provides prospects for future research directions.

The causes of MCD and FSGS are mainly divided into three categories: caused by genetic mutations in podocyte-related genes, secondary factors such as infection or drug use, and unexplained primary factors. Genetic mutations only account for about 8-14% [1-3], secondary factors account for 20-30% [4-5], and the rest are basically caused by unknown primary factors.

Circulating Factor Theory and FSGS

When Hoyer et al. [6] first reported patients with recurrent FSGS after kidney transplantation in 1972, researchers speculated that certain circulating osmotic factors in the blood might cause the production of FSGS. Subsequent epidemiological studies have shown that FSGS after transplantation has a high recurrence rate, sometimes occurring within hours after transplantation. This result further prompted researchers to believe that the causative factor may not be a "local phenomenon within the kidney", but a blood circulation permeability factor that damages podocytes by changing the glomerular barrier [7-8]. Until 2012 Gallon et al[9]In a case study of a patient with recurrence of FSGS, it was found that when the transplanted kidney was removed and retransplanted to another recipient without a history of FSGS, the function of the kidney was restored, and the histopathological damage unique to FSGS was also reversed, which basically determined that certain circulating osmotic factors present in the blood caused the occurrence of FSGS.

In the following years, a large amount of research resources were devoted to finding circulating osmotic factors that play a key role in the formation of proteinuria in MCD and pFSGS [10-11]. Despite the investment of very large resources and research efforts, the identification of these unknown circulating permeability factors has been arduous and lengthy, and researchers have not found reliable and consistent biomarkers.

Some candidate biomarker molecules thought to increase serum protein vascular permeability (Table 1) include soluble urokinase-type plasminogen activator receptor (suPAR) [12], cardiotrophin-like cytokine-1 (CLCF-1) [13], soluble CD40 ligand [14], etc., have been found to be associated with the progression and recurrence of MCD and pFSGS, but studies to verify their clinical utility have shown that these markers do not play a decisive role in the development of MCD and FSGS, and they can also be detected in the serum of healthy and non-nephrotic syndrome patients [15-17].

table1. FSGS and MCD related circulating permeability factors and their limitations

Nephrin protein introduction

In 1966, in a group of Finnish families with congenital nephrotic syndrome (NPHS1) [39], it was discovered that this is a kidney disease caused by a rare genetic mutation, but the specific gene causing this disease is unknown. It was not until 1998 that the key chromosomal region where the NPHS1 gene was located was identified and sequenced. The expression product of the NPHS1 gene was named Nephrin, and its locus was located on chromosome 19q12-q13.1 [40-41]. Although the precise structure and function of the Nephrin protein remain unknown at this time, it is speculated that its domain structure is similar to a large class of cell adhesion receptors belonging to the immunoglobulin family, and that it can function as an adhesion receptor and signaling protein [40]. In subsequent studies [42-43], confirmed that Nephrin is a component of the podocyte slit diaphragm (SD)-related complex, which together with glomerular endothelial cells and basement membrane (GBM) forms the glomerular filtration barrier.(Figure 1)

Figure 1. Nephrin antibodies induce mediastinal changes, podocyte damage, and protein leakage into urine.

Nephrin is a transmembrane protein with an extracellular fragment at its N-terminus and an intracellular domain at its C-terminus. The detection of an antibody specific for the N-terminus of human Nephrin confirmed that it is located in the mediastinum of the kidney [44]. The entire protein consists of eight immunoglobulin-like extracellular domains, a fibronectin type III-like domain, a transmembrane domain and a short intracellular domain. It maintains the tissue morphology, structure and function of podocytes through interaction with the actin cytoskeleton [40,45]. As a cell surface receptor protein of the immunoglobulin superfamily, Nephrin is also involved in intercellular adhesion and signaling functions. When Fyn of the Src kinase family phosphorylates one or more of the six tyrosine residues (Tyr1114, Tyr1136, Tyr1176, Tyr1183, Tyr1193, Tyr1217) of the intracellular domain of Nephrin, downstream signaling mediated by the intracellular tail of Nephrin is activated [46]. The specific performance is as followsThe phosphorylated intracellular domain interacts with several podocyte cytosolic proteins, including podocin, CD2AP, NEPH1, and phosphatidylinositol 3-kinase (PI3K), to transmit downstream signals to the actin cytoskeleton to regulate podocyte structural integrity and glomerular filtration.narrowThe function of sewing[47-48]. Phosphorylated Nephrin promotes the recruitment and interaction of Nck adapter proteins, ZO-1, catenin, barrel proteins, podocytes, CD2-AP, and PI3K. Nck adapter proteins send signals for actin remodeling and participate in the regulation of multiple intracellular signaling pathways, thereby affecting the polymerization dynamics of actin that is critical for podocyte stability [48-49] (Figure 2).

Figure 2. Schematic diagram of the gap layer between adjacent foot processes. The intracellular domain interacts with cytosolic proteins through cascade reactions when phosphorylated, inducing signaling, modulating actin polymerization dynamics and adjusting podocyte structure. The interaction between anti-Nephrin antibodies and Nephrin results in the loss of slit barrier function by causing Nephrin to relocate away from the slit layer.

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