FIBRONECTIN glomerulopathy is a rare, autosomal dominant, non-amyloid glomerular disease. It was firstly reported by Burgin and his colleagues in 1980. [1] Thereafter, a few case reports were published describing familial and sporadic patients with fibronectin glomerulopathy. [2,3,4,5] It usually onsets in adolescence with proteinuria, microhematuria, hypertension, and slowly progressive renal failure. In pathology, it is characterized by nonimmune lobular glomerulopathy with massive mesangial and subendothelial deposits composed of fibronectin. In 2008, Castelletti et al[6] identified three mutations in the gene encoding fibronectin 1 (FN1) in the different pedigrees of fibronectin glomerulopathy.
In this article, we reported a Chinese patient of fibronectin glomerulopathy, which was confirmed by kidney biopsy. Genetic sequencing of FN1 detected a previously described heterozygous mutation causing a substitution of the tyrosine at amino acid 973 by cysteine (Y973C) in the affected patient.
CASE DESCRIPTION
A 29-year-old woman was hospitalized due to systemic edema and hypertension. She had cerebral infarction three years ago in past medical history. In family history, her father had diabetes and hypertension. Findings of physical examination included obesity, elevated blood pressure of 150/90 mm Hg and peripheral edema. Urinalysis revealed massive proteinuria and microscopic hematuria with bland sediment, and 24-hour urine protein was 15.43 g. Serum albumin and creatinine levels were 28 g/L and 79.6 μmol/L [corresponding to estimated glomerular filtration rate (eGFR) of 86 ml/min·1.73 m2 calculated by Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) equation], with hypercholesterolemia. Antinuclear antibody and anti-double stranded DNA were negative, as well as hepatitis serologic test. No hypocomplementemia or cryoglobulinemia was detected. Serum protein electrophoresis did not show monoclonal protein. Enlarged kidneys (left was 13.4 cm, and right was 13.2 cm in long diameter) were found in ultrasonography.
Light microscopy revealed enlarged and hyperlobular glomeruli with a homogeneous material in the mesangium and subendothelium (Fig. 1A). There was mild mesangial hyperplasia, without endocapillary or extracapillary proliferation, necrosis, or inflammatory infiltration. The deposited material was period acid-Schiff positive (Fig. 1B) and methenamine silver negative (Fig. 1C). Negative result of Congo red staining excluded amyloidosis.
Immunofluorescence microscopy was negative for immunoglobulin, light chain, or complement components. Electron microscopy showed massive electron-dense deposits in the mesangial and subendothelial space, with some admixture of irregularly arranged fibrils (15 nm in width) (Fig. 1D and 1E). Indirect immunofluorescence staining for fibronectin was performed, and glomerular deposits stained brightly with monoclonal anti-fibronectin antibody (Fig. 1F).
The diagnosis of fibronectin glomerulopathy was established on the basis of renal pathology. As an additional investigation, we performed genetic sequencing of FN1 in the patient and her parents. Genomic DNA was extracted from peripheral blood. The whole exons of FN1 were screened by Sanger sequencing (Sangon Biotech Co., Ltd, Shanghai, China). DNA sequencing showed a previously described heterozygous mutation in exon 19 in which an adenine is substituted by a guanine at nucleotide 2918 of the complementary DNA (c. 2918 A>G) (Fig. 2), leading to replacement of a tyrosine at amino acid 973 by a cysteine (Y973C). No FN1 mutation was detected in her parents.
Anti-hypertensive therapy including angiotensin II receptor antagonist, calcium channel blocker, α and β receptor blocker was given as well as statin for hyperlipidemia. At her last evaluation 12 month after diagnosis, serum creatinine level was 86.6 μmol/L (eGFR 81 ml/min·1.73 m2) while serum albumin level was 30 g/L. 24-hour urine protein was unchanged.
DISCUSSION
Fibronectin is a multifunctional extracellular matrix glycoprotein involved in cellular adhesion, migration, and cytoskeleton maintenance. It normally exists in a cellular form and a plasma soluble form. The deposit in kidneys of fibronectin glomerulopathy is mainly derived from circulating fibronectin, [2] although plasma fibronectin levels are not elevated in these patients.
Castelletti et al[6] sequenced FN1 and found that heterozygous FN1 mutations constituted the underlying genetic abnormality in 6 of 15 unrelated pedigrees (40%). Three heterozygous missense mutations were detected, Y973C, W1925R and L1974R that co-segregated with fibronectin glomerulopathy. The Y973 mutation was found in four pedigrees. The four families were of different ethnic origin and did not share a disease haplotype, thus excluding a founder effect. A heterozygous Y973C mutation was found in our patient. However, the mutation of FN1 was not detected in her parents. Thus, she could be a sporadic case of fibronectin glomerulopathy. Ishimoto et al[7] reported a Japanese case of sporadic, elderly-onset fibronectin glomerulopathy, but did not find any missense mutation of FN1 previously reported. In 2015, a retrospective study of ten Chinese patients of fibronectin glomerulopathy showed that only one case had a family history of renal disease. [8] However, the mentioned study in China did not sequence FN1 to investigate the potential mutation. Our patient was the first case reported with Y973C mutation in China.
Functional studies using recombinant proteins suggest that mutations in FN1 impair the process of the assembly of fibronectin into fibrils and the balance between soluble and insoluble form of fibronectin. [6] Y973C mutation affects the Hep-Ⅲ heparin-binding domain of fibronectin, located in the fourth and fifth of fibronectin’s type Ⅲ (Ⅲ4-5) repeats. Hep-Ⅲ has the ability to modulate the cytoskeletal response and induce intracellular signaling. [9] Y973C mutation introduces an additional cysteine in the fourth type Ⅲ repeat, which might affect protein folding and function through the formation of abnormal disulfide bonds. Although mutations in FN1 may be necessary for fibronectin glomerulopathy to develop, they are not sufficient since not all family members with the mutation develop clinical kidney disease.
In conclusion, we presented a rare case of fibronectin glomerulopathy in China. This case attaches the importance to electron microscopy and anti-fibronectin antibody staining when nonimmune, lobular glomerulopathy with a massive homogeneous material deposit in the mesangium and subendothelium is observed. The identification of mutations in FN1 is an advance in understanding of the molecular genetic mechanisms of fibronectin glomerulopathy.
Figure 1.
Figure 1.
Pathological changes of kidney with fibronectin glomerulopathy.
A. Glomeruli were diffusely enlarged with lobular accentuation and minimal hypercellularity (arrows). (HE staining) B. Period acid-Schiff-positive material expanding the mesangium and subendothelium of an enlarged glomerulus (arrows). C. The deposits in the subendothelium and mesangium (arrows) failed to stain with silver. (Jones methenamine silver stain) D. Electron microscopy showed mesangial and subendothelial deposits (arrows). E. The electron dense deposits as well as 15 nm fibrils (arrows) arranged randomly were observed. F. Intense immunofluorescence staining showed glomerular deposits with antisera against fibronectin [primary antibody: mouse monoclonal anti-human fibronectin antibody, clone IST-4 (Sigma-Aldrich); secondary antibody: fluorescin isothiocynanate-conjugated goat anti-mouse]. (×200)
Figure 2.
Figure 2.
Mutation 2918A→G in the fibronectin gene.
The sequencing electropherogram that shows the heterozygous mutation c.2918A→G (reference single-nucleotide polymorphism identifier rs137854488) causing a substitution of tyrosine at amino acid 973 by cysteine in the patient.
参考文献
Label-free quantitative proteomic analysis reveals potential biomarkers and pathways in renal cell carcinoma
Renal cell carcinoma (RCC) is one of the most common malignancies in adults, and there is still no acknowledged biomarker for its diagnosis, prognosis, recurrence monitoring, and treatment stratification. Besides, little is known about the post-translational modification (PTM) of proteins in RCC. Here, we performed quantitative proteomic analysis on 12 matched pairs of clear cell RCC (ccRCC) and adjacent kidney tissues using liquid chromatography-tandem mass spectrometry (nanoLCMS/MS) and Progenesis LC-MS software (label-free) to identify and quantify the dysregulated proteins. A total of 1872 and 1927 proteins were identified in ccRCC and adjacent kidney tissues, respectively. Among these proteins, 1037 proteins were quantified by Progenesis LC-MS , and 213 proteins were identified as dysregulated proteins between ccRCC and adjacent tissues. Pathway analysis using IPA, STRING, and David tools was performed, which demonstrated the enrichment of cancer-related signaling pathways and biological processes such as mitochondrial dysfunction, metabolic pathway, cell death, and acetylation. Dysregulation of two mitochondrial proteins, acetyl- CoA acetyltransferase 1 (ACAT1) and manganese superoxide dismutase (MnSOD) were selected and confirmed by Western blotting and immunohistochemistry assays using another 6 pairs of ccRCC and adjacent tissues. Further mass spectrometry analysis indicated that both ACAT1 and MnSOD had characterized acetylation at lysine residues, which is the first time to identify acetylation of ACAT1 and MnSOD in ccRCC. Collectively, these data revealed a number of dysregulated proteins and signaling pathways by label-free quantitative proteomic approach in RCC, which shed light on potential diagnostic or prognostic biomarkers and therapeutic molecular targets for clinical intervention of RCC.
Glomerulopathy associated with predominant fibronectin deposits: a newly recognized hereditary disease
Glomerulopathy associated with predominant fibronectin deposits: A newly recognized hereditary disease. A newly recognized type of familial glomerulopathy observed in patients of both sexes in six families is reported. Proteinuria, often within the nephrotic range, microscopic hematuria, hypertension and a slowly decreasing renal function over several years were common. No underlying systemic diseases were identified. Generally, light microscopy showed enlarged glomeruli with minimal hypercellularity and with extensive deposits in the mesangium and subendothelial space. By electron microscopy, granular deposits with some admixture of fibrils were most common. In one family, the deposits were predominantly fibrillary. Immunoglobulins and complement factors were inconstant or lacking. A main finding was a strong immune reactivity to fibronectin, corresponding to the distribution of the deposits. In one patient, the deposits recurred in a renal transplant. There was no indication of systemic deposition. Abnormalities in the metabolism of circulating fibronectin may play a pathogenetic role in this disease of probably autosomal dominant inheritance.
Label-free quantitative proteomic analysis reveals dysfunction of complement pathway in peripheral blood of schizophrenia patients: evidence for the immune hypothesis of schizophrenia
Familial glomerulopathy with giant fibrillar (fibronectin-positive) deposits: 15-year follow-up in a large kindred
Abstract A 15-year clinical follow-up is reported for a familial glomerulopathy characterized on light microscopy by the glomerular deposition of giant fibrillary deposits (Virchows Arch A Pathol Anat Histol 388:313-326, 1980). On electron microscopy, the deposits consist of randomly oriented fibrils (12 to 16 nm in width and 120 to 170 nm in length). These deposits show positive immunoreactivity for fibronectin. One hundred fifty-seven of 197 family members within five generations were investigated. The disease is characterized by the occurrence of albuminuria in the third to fourth decades of life and slow progression to end-stage renal disease over a period of 15 to 20 years with the occurrence of generalized distal tubular acidosis (renal tubular acidosis type IV), hypertension, and the nephrotic syndrome. The frequent occurrence of otherwise unexplained microalbuminuria in young individuals of generations IV and V could be indicative of incipient glomerular disease. In one affected male individual and in his unaffected sister, renal cell carcinoma was diagnosed, raising the possibility that this familial glomerulopathy might be associated with an increased risk to develop renal cell cancer by direct or indirect (associated genetic predisposition) mechanisms. The disease relapsed in one renal transplant, raising the possibility of the presence of a transferable factor that could be part of the deposited fibrillar material or, alternatively, interfere with the glomerular handling of the deposited material.
Glomerulopathy associated with predominant fibronectin deposits: exclusion of the genes for fibronectin, villin and desmin as causative genes
Glomerulopathy with predominant fibronectin deposits (GFD) is a newly recognized autosomal dominant renal disease that leads to albuminuria, microscopic hematuria, hypertension, renal tubular acidosis type IV, and end-stage renal disease in the second to fourth decade of life. Light microscopy documents extensive deposits in the subendothelial space, which on electron microscopy consist of non-oriented 12 x 125 nm fibers. Deposits are strongly immunoreactive for antibodies to fibronectin. We examined the hypothesis that a genetic defect in the gene for fibronectin is responsible for the disease. In a 197 member pedigree, 13 relatives developed end-stage renal failure from the disease. In 99 individuals haplotype analysis was performed using 6 microsatellite markers spanning a > 56 cM interval in chromosome region 2q34, where fibronectin, villin, and desmin map in close proximity. Haplotype analysis resulted in exclusion of the whole range of 78 cM covered by the markers examined. This result excludes fibronectin, villin, and desmin from being the causative genes for GFD in this large kindred.
Mutations in FN1 cause glomerulopathy with fibronectin deposits
Abstract Glomerulopathy with fibronectin (FN) deposits (GFND) is an autosomal dominant disease with age-related penetrance, characterized by proteinuria, microscopic hematuria, hypertension, and massive glomerular deposits of FN that lead to end-stage renal failure. The genetic abnormality underlying GFND was still unknown. We hypothesized that mutations in FN1, which encodes FN, were the cause of GFND. In a large Italian pedigree with eight affected subjects, we found linkage with GFND at the FN1 locus at 2q32. We sequenced the FN1 in 15 unrelated pedigrees and found three heterozygous missense mutations, the W1925R, L1974R, and Y973C, that cosegregated with the disease in six pedigrees. The mutations affected two domains of FN (Hep-II domain for the W1925R and the L1974R, and Hep-III domain for the Y973C) that play key roles in FN-cell interaction and in FN fibrillogenesis. Mutant recombinant Hep-II fragments were expressed, and functional studies revealed a lower binding to heparin and to endothelial cells and podocytes compared with wild-type Hep-II and an impaired capability to induce endothelial cell spreading and cytoskeletal reorganization. Overall dominant mutations in FN1 accounted for 40% of cases of GFND in our study group. These findings may help understanding the pathogenesis of proteinuria and glomerular FN deposits in GFND and possibly in more common renal diseases such as diabetic nephropathy, IgA nephropathy, and lupus nephritis. To our knowledge no FN1 mutation causing a human disease was previously reported.
Fibronectin glomerulopathy
Clinical and morphological features of fibronectin glomerulopathy: a report of ten patients from a single institution
Fibronectin glomerulopathy is a rare glomerular disease caused by the progressive deposition of fibronectin. We report 10 Chinese patients with fibronectin glomerulopathy.Renal biopsies were performed on all patients, and the clinical and pathological parameters for all patients were analyzed.There were 6 males and 4 females, with an average age of 29 卤8 years. One patient had a family history of renal disease, all patients presented with proteinuria, and 80% of them suffered nephrotic range proteinuria. No patient demonstrated gross hematuria. The levels of serum creatinine were elevated, and the eGFR was decreased in 5 patients. Renal biopsy revealed a lobulated glomerular tuft. Patients showed numerous periodic acid-Schiff-positive and fuchsinophilic deposits in the mesangial area and along the capillary loops. Immense levels of fibronectin were detected in the glomerulus after immunofluorescence analysis. An electron microscopy scan found numerous electron-dense deposits in the mesangial and subendothelial areas. Immune-electron microscopy confirmed that the deposits consisted of fibronectin.Nephrotic proteinuria and massive intraglomerular fibronectin deposits are the most significant features of fibronectin glomerulopathy. *Huiping Chen and Hao Bao have contributed equally to the work and are to be considered first authors.
Label-free quantitative proteomic analysis of puccinia psidii uredospores reveals differences of fungal populations infecting eucalyptus and guava
Puccinia psidii sensu lato (s.l.) is the causal agent of eucalyptus and guava rust, but it also attacks a wide range of plant species from the myrtle family, resulting in a significant genetic and physiological variability among populations accessed from different hosts. The uredospores are crucial to P. psidii dissemination in the field. Although they are important for the fungal pathogenesis, their molecular characterization has been poorly studied. In this work, we report the first in-depth proteomic analysis of P. psidii s.l. uredospores from two contrasting populations: guava fruits (PpGuava) and eucalyptus leaves (PpEucalyptus). NanoUPLC-MSE was used to generate peptide spectra that were matched to the UniProt Puccinia genera sequences (UniProt database) resulting in the first proteomic analysis of the phytopathogenic fungus P. psidii. Three hundred and fourty proteins were detected and quantified using Label free proteomics. A significant number of unique proteins were found for each sample, others were significantly more or less abundant, according to the fungal populations. In PpGuava population, many proteins correlated with fungal virulence, such as malate dehydrogenase, proteossomes subunits, enolases and others were increased. On the other hand, PpEucalyptus proteins involved in biogenesis, protein folding and translocation were increased, supporting the physiological variability of the fungal populations according to their protein reservoirs and specific host interaction strategies.
