Thin Basement Membrane Nephropathy Research Articles

Aberrant serum-derived FN1 variants bind to integrin β1 on glomerular endothelial cells contributing to thin basement membrane nephropathy

The glomerular basement membrane (GBM) is a critical component of the glomerular filtration barrier (GFB), with its thickness directly influencing renal function. While a uniformly thinned GBM can cause hematuria while preserving normal renal function, this condition is typically diagnosed as thin basement membrane nephropathy (TBMN). However, the pathogenesis and potential progression to renal insufficiency of TBMN are not fully understood. In this study, we analyzed clinical cohorts presenting with microscopic hematuria who underwent genetic testing and identified five novel pathogenic FN1 mutations. Through bioinformatics analysis of these variants, expression localization analysis of GBM-related molecules in renal biopsies, and functional studies of the mutants, we found that these variants exhibited gain-of-function characteristics. This led to the excessive deposition of aberrant serum-derived FN1 variants on glomerular endothelial cells rather than cell-type-specific variants. The deposition competitively binds FN1 variants to Integrin β1, disrupting the interaction with Laminin α5β2γ1 and subsequently reducing the expression of key GBM components, resulting in TBMN. This study elucidated, for the first time, the genetic pathogenesis of TBMN caused by FN1 variants. It provides a crucial foundation for understanding the progression of renal dysfunction associated with simple hematuria, highlights the potential for targeted therapeutic strategies, and differentiates TBMN from early-stage Alport syndrome.

Genotype-first analysis in an unselected health system-based population reveals variable phenotypic severity of COL4A5 variants.

Our knowledge of X-linked Alport Syndrome [AS] comes mostly from selected cohorts with more severe disease. We examined the phenotypic spectrum of X-linked AS in males and females with a genotype-based approach using data from the Geisinger MyCode DiscovEHR study, an unselected health system-based cohort with exome sequencing and electronic health records. Patients with COL4A5 variants reported as pathogenic (P) or likely pathogenic (LP) in ClinVar, or protein-truncating variants (PTVs), were each matched with up to 5 controls without COL4A3/4/5 variants by sociodemographics, diabetes diagnosis, and year of first outpatient encounter. AS-related phenotypes included dipstick hematuria, bilateral sensorineural hearing loss (BSHL), proteinuria, decreased eGFR, and ESKD. Out of 170,856 patients, there were 29 hemizygous males (mean age 52.0 y [SD 20.0]) and 55 heterozygous females (mean age 59.3 y [SD 18.8]) with a COL4A5 P/LP variant, including 48 with the hypomorphic variant p.Gly624Asp. Overall, penetrance (having any AS phenotypic feature) was highest for non-p.Gly624Asp P/LP variants (males: 94%, females: 85%), intermediate for p.Gly624Asp (males: 77%, females: 69%), compared to controls (males: 32%; females: 50%). The proportion with ESKD was highest for males with P/LP variants (44%), intermediate for males with p.Gly624Asp (15%) and females with P/LP variants (10%), compared to controls (males: 3%, females 2%). Only 47% of individuals with COL4A5 had completed albuminuria screening, and a minority were taking renin-angiotensin aldosterone system (RAAS) inhibitors. Only 38% of males and 16% of females had a known diagnosis of Alport syndrome or thin basement membrane disease. In an unselected cohort, we show increased risks of AS-related phenotypes in men and women compared to matched controls, while showing a wider spectrum of severity than has been described previously and variability by genotype. Future studies are needed to determine whether early genetic diagnosis can improve outcomes in Alport Syndrome.

#234 Clinical spectrum and genetic variants of Alport syndrome and thin basement membrane nephropathy—single center experience

Abstract Background and aims Alport syndrome (AS) and thin basement membrane nephropathy (TBMN) are genetically heterogeneous kidney diseases that manifest with kidney involvement, AS also with hearing and visual impairment. Electron microscopic examination shows characteristic ultrastructural changes in the glomerular basement membrane (GBM) in AS and TBMN, but cannot reliably distinguish between the two entities. Therefore, molecular genetic studies and the identification of individual pathogenic variants in the target genes are crucial to confirm the diagnosis. The aim of our study is to determine the significance of the genetic variants associated with the ultrastructural changes of GBM found in the renal biopsy. Method We retrospectively analyzed records of patients from the University Medical Centre Ljubljana in which ultrastructural GBM changes characteristic for AS and TBMN were found in the renal biopsy, including their clinical, laboratory and histological features. Molecular genetic investigations were made using the new generation sequencing method (NGS) to determine germline pathogenic variants in genes directly related to AS and TBMN (COL4A3, COL4A4 and COL4A5) and in genes that are important in syndromes with a related clinical picture (COL4A6, CD151, LMX1B, MYH9, FN1 and MYO1E). The pathogenicity of variants was determined according to the Clinical Molecular Genetic Society best practice guidelines. Results The study included 73 patients with ultrastructurally confirmed thin GBM. Indications for kidney biopsy were chronic nephritis syndrome in 33 patients (45%), acute worsening of renal function in 16 (22%), nephrotic syndrome in 9 (12%), nephrotic proteinuria and chronic kidney disease in 6 (8%), isolated proteinuria in 5 (7%) and suspected vasculitis in 4 patients (6%). At the time of kidney biopsy average age was 50 ± 16 years, average creatinine 112 ± 70 µmol/L and average estimated glomerular filtration rate (eGFR) 65 ± 24 ml/min/1.73 m2, 7 patients had eGFR below 30 ml/min/1.73 m2. Average daily proteinuria was 2.4 ± 3.3 gr and 88% of patients had hematuria. 38% of them have hearing impairment and 43% had positive familiar history of kidney disease. We recorded 33 patients (45%) with genetic variants associated with AS or TBMN: 22 patients (67%) with pathogenic variants, 2 patients (6%) with likely pathogenic and 9 patients (27%) with variants of uncertain significance. The presence of a positive genetic analysis was associated with a higher level of hematuria (p = 0.003), a lower level of proteinuria (p = 0.004) and a positive family history of kidney disease (p < 0.001). It was also associated with hearing loss, but the association was not statistically significant (p = 0.099). Patients with genetic variants had similar eGFR to those with negative genetics (63.9 ml/min/1.73 m2 (58.3-69.4) vs 66.2 ml/min/1.73 m2 (60.0-72.3)), p = 0.698. We also found no differences in GBM thickness in patients without compared to those with identified gene variants (213 nm (160-315) vs. 209 nm (140-410)), p = 0.761. 32 patients (44%) had only focal chronic changes and ultrastructural changes of thin GBM present on kidney biopsy and 41 patients (56%) had also been diagnosed with other renal pathology. When comparing these two groups, patients without associated pathology had lower daily proteinuria (p = 0.048), higher level of hematuria (p = 0.021), frequent hearing impairment (p = 0.008) and also positive genetics analysis (p = 0.016). Conclusion We have shown that in patients with proven ultrastructural alterations in GBM, the presence of so far recognized genetic variants associated with AS and TBMN is 45%, and corresponds to the frequency of other phenotypic characteristics of AS, including hearing impairment and positive family history. In the future, additional clinical tools will be required to accurately determine the role of coexistent thin GBM on the prognosis of patients with other relevant kidney pathology and other genetic tools beyond NGS should be employed to explore other pathologic genetic variants associated with the AS spectrum.

#2438 ADPKD and collagen genes (COL4A3, COL4A4, COL4A5)

Abstract Background and Aims Familial hematuria diseases are a heterogeneous group of monogenic conditions caused by mutations in one of the collagen IV genes: COL4A3 (2q36.3), COL4A4 (2q36.3), and COL4A5 (Xq22.3) that are expressed in the glomerular basement membranes (GBM) and are responsible for the most frequent forms of microscopic hematuria (MH), Alport syndrome, and thin basement membrane nephropathy (TBMN). Recent data suggest that about 1% (1 in 106 individuals) of the world population may have heterozygous predicted pathogenic COL4A3 or COL4A4 variants, a frequency that leads to the occasional superimposition of TBMN with other glomerulopathies. Autosomal Dominant Polycystic Kidney Disease (ADPKD) is a major genetic disorder affecting up to 12.5 million individuals worldwide and it is the fourth most common global cause of renal replacement therapy. Two are the principal causative genes: PKD1(16p13.3) and PKD2 (4q22.1). The aim of the study was to describe a cohort of clinically ADPKD patients studying collagen IV genes COL4A3, COL4A4, and COL4A5. Method We performed NGS with Sophia Genetic “Nephropathies Solution” Panel on clinically ADPKD patients (pts). This panel includes 44 genes (target region 105.8 kb) involved in different types of nephropathies. We considered variants that are categorized as Variant of Uncertain Significance (Vus), likely pathogenic, or pathogenic (Class 3,4, or 5 of American College of Medical Genetics (ACMG)) in COL4A3, COL4A4, and COL4A5 genes. Results We analyzed 250 consecutive clinically ADPKD pts (consecutive in the outpatient clinic). We found 13 (6.3%) clinically potentially interesting (Class 3,4 or 5 of ACMG) variants in collagen genes. We found 2 pts that were negative for PKD1, PKD2, PKHD1 analysis: 1 pt carrier COL4A4 NM_000092.5: c.2908C>T (p.Gln970Ter), Pathogenic and c.2756A>G (p.Glu919Gly) Vus; 1 male pt carrier COL4A3 NM_000091.5: c.609+3_609+6del, Vus and COL4A5 NM_000495:c.169G>A p.(Gly57Arg), likely pathogenic. We also found 4 pts with 4 Vus variants in collagens genes combined with at least 1 pathogenic or likely pathogenic variant in PKD1 or PKD2 genes. We found 3 pts with likely pathogenic variants in COL4A4 or COL4A3 genes combined with Vus or likely benign variants in PKD1 or PKD2 genes and 1 patient with 1 likely pathogenic variant in COL4A3 and 1 likely pathogenic variant in PKD1. Conclusion Variants in collagen genes are very frequent and the associated phenotype is varied, sometimes very mild such as microhematuria. Furthermore, penetrance has to be considered. The study outlines the importance of considering also collagen genes, even if we have a clear clinical manifestation of ADPKD. An additional variant in a kidney already compromised can have an impact on the phenotype. Analysis of genes involved in these 2 pathologies could lead to a better understanding of the phenotype-genotype correlation at least in patients with both conditions.

#370 Macroscopic hematuria-associated severe acute kidney injury from thin basement membrane disease

Abstract Background Macroscopic hematuria-associated acute kidney injury (MH-AKI) is a rare condition reported in IgA nephropathy and others, and the mechanism of renal tubular damage caused by red blood cells in the tubules appears to be similar to that of anticoagulant nephropathy. Case presentation A 66-year-old Japanese woman with no previous medical history was referred to our hospital with anuric severe kidney injury. Seven days earlier, she had transient acute lumbar back pain, which was followed by gradual worsening of malaise and loss of appetite. On admission, her serum BUN was 147.8 mg/dL, serum Cr 18.57 mg/dL, hemoglobin 11.2 g/dL, urinary red blood cells> 100/HPF, urinary protein 28.8 g/gCr, no hydronephrosis in either kidney, but a small stone in the right kidney. After admission, while starting hemodialysis, she was treated with methylprednisone 1000 mg for 3 days followed by prednisone 40 mg/day because of suspected rapidly progressive glomerulonephritis, but both MPO-/PR3-ANCA and anti-GBM antibodies were negative. Renal biopsy from left kidney showed acute tubular injury with large amounts of red blood cells and red blood cell casts filling the distal tubules (Figure), and prednisone was discontinued. Subsequently, urine flowed out with gross hematuria, and her hematuria and renal function improved with increased urine output, and she was weaned from hemodialysis. Electron microscopy showed glomerular thin basement membrane (TBMD) (Figure). Her renal function and hematuria slowly improved and the right kidney stone disappeared. Microscopic hematuria eventually disappeared, and her serum Cr level was 1.1 mg/dL one year after onset. Conclusions This is the first report of MH-AKI in TBMD without a background of anticoagulation, severe thrombocytopaenia, or iron overload. In one report, compared to IgA nephropathy, TBMD is associated with increased urinary Ca and uric acid excretion, which is associated with gross hematuria and back pain [1]. In this case of MH-AKI from TBMD, both red blood cells and crystal formation in the tubular lumens may have transiently and synergistically caused renal tubular obstruction and heme iron-induced tubular damage.

Remission induced by renal protective therapy in nephrotic syndrome with thin basement membrane in an older patient: a case report

BackgroundAdult nephrotic syndrome is a well-known kidney disease that causes heavy proteinuria, hypoalbuminemia, hypercholesterolemia, edema, and hypertension. The treatment varies according to its underlying cause but often faces medication resistance or adverse drug effects.Case presentationA Japanese woman in her 80s presented with nephrotic syndrome after a 3 year latent period of urinary protein and occult blood. She did not have any secondary causes of nephrotic syndrome. Renal biopsy revealed thin glomerular basement membrane, partial foot process fusion on electron microscopy with minor glomerular change on light microscopy, and slight coarse immunoglobulin M deposition in the mesangium on immunofluorescence microscopy, which was inconsistent with any other glomerular diseases. Without steroid treatment, she dramatically remitted from proteinuria after the administration of the renal protective agents enalapril, ezetimibe, rosuvastatin, and dapagliflozin. Recurrence after 8 months of follow-up subsided with the administration of additional doses of the agents.ConclusionsThis case illustrated the novel outcomes of combining medical treatment without steroid use for nephrotic syndrome with thin glomerular basement membrane disease. At the time of writing this report, the patient’s renal function was stable and she was free of edema, although moderate proteinuria and occult hematuria persisted. The final diagnosis was uncertain because of the lack of genetic investigation; however, the response to the aforementioned medical treatment suggests the effectiveness of the supportive therapy.

Cystic phenotype and chronic kidney disease in autosomal dominant Alport syndrome.

Autosomal dominant Alport Syndrome (ADAS), also known as thin basement membrane disease (TBMD), is caused by pathogenic variants in the COL4A3 and COL4A4 genes. A cystic phenotype has been described in some patients with TBMD, but no genetic studies have been performed. We conducted a genetic and radiologic investigation in a cohort of ADAS patients to analyze the prevalence of multicystic kidney disease (MKD) and its association with chronic kidney disease (CKD). This was a retrospective single-center cohort study. Thirty-one patients showing pathogenic or likely pathogenic variants in COL4A3 or COL4A4 from a cohort of 79 patients with persistent microscopic hematuria were included. Mean follow-up was 9.4±9.6years. The primary objective of the study was to determine the prevalence of MKD in the cohort of ADAS patients. Secondary objectives were to determine risk factors associated with an estimated glomerular filtration rate (eGFR) <45mL/min/1.73 m2 at the time of genetic and radiologic evaluation and to investigate the coexistence of other genetic abnormalities associated with familial hematuria and cystic kidney disease. MKD was found in 16 patients (52%). Mean number of cysts per kidney was 12.7±5.5. No genetic abnormalities were found in a panel of 101 other genes related to familial hematuria, focal segmental glomerulosclerosis and cystic kidney disease. A greater number of patients with MKD had an eGFR <45mL/min/1.73 m2 (63% vs 7%, P=.006) and more advanced CKD than patients without MKD. The annual rate of eGFR decline was greater in patients with MKD: -1.8 vs 0.06mL/min/1.73 m2/year (P=.009). By multivariable linear regression analysis, the main determinants of eGFR change per year were time-averaged proteinuria (P=.002) and MKD (P=.02). MKD is commonly found in ADAS and is associated with a worse kidney outcome. No pathogenic variants were found in genes other than COL4A3/COL4A4.

Clinicopathological Features of Hereditary Nephritis in the Iranian Population: Analysis of a 14-Year Survey in Kidney Biopsies From a Large Referral Center.

Hereditary nephritis (HN), including Alport syndrome (AS) and thin basement membrane nephropathy (TBMN), is a rare genetic cause of hematuria. A definitive diagnosis requires electron microscopy (EM). Therefore, the clinical characteristics of these conditions are less known. This study aimed to determine the percentage and clinicopathological features of HN in patients from a referral center in Iran. We checked kidney biopsy reports from 2007 to 2021 and extracted cases with HN. Fresh specimens of the cases diagnosed in the last two years were stained by immunofluorescence (IF) for collagen type IV alpha chains. EM findings in these cases were re-evaluated and categorized as diffuse glomerular basement membrane (GBM) thinning, definite, and suspicious features of AS. We analyzed 3884 pathology reports of kidney biopsies from 2007 to 2021 and identified 210 patients (5.4%) with HN, with a mean age of 13.78±12.42 years old. Hematuria with proteinuria (53.3%), isolated hematuria (44.2%), and proteinuria with hematuria and increased creatinine (2.5%) were found in these patients. The re-evaluation of EM findings revealed GBM thinning, definite, and suspicious findings of AS in 37.5%, 43.8%, and 18.8% cases, respectively. The most common diagnosis in 32 cases after the IF study was X-linked AS (71.9%), and 6.2% of cases were autosomal recessive AS. TBMN and autosomal dominant AS remained the differential diagnoses in 21.9%. It was found that EM is helpful for the primary diagnosis of patients with definite AS. Immunostaining improves the diagnostic sensitivity for the differentiation of those with suspicious EM findings and determines the inheritance pattern. However, a multidisciplinary approach for a subset of cases is required for the best diagnosis and management.

Beyond COL4A: Combined Renal Histopathology and Genetic Testing Reveals the Genetic Complexity of Thin Glomerular Basement Membrane Disease

Beyond COL4A: Combined Renal Histopathology and Genetic Testing Reveals the Genetic Complexity of Thin Glomerular Basement Membrane Disease

Glomerulocystic Kidney Disease with Concurrent Thin Basement Membrane Disease in an Adult Male

Glomerulocystic Kidney Disease with Concurrent Thin Basement Membrane Disease in an Adult Male

Genetic Modifiers of Mendelian Monogenic Collagen IV Nephropathies in Humans and Mice.

Familial hematuria is a clinical sign of a genetically heterogeneous group of conditions, accompanied by broad inter- and intrafamilial variable expressivity. The most frequent condition is caused by pathogenic (or likely pathogenic) variants in the collagen-IV genes, COL4A3/A4/A5. Pathogenic variants in COL4A5 are responsible for the severe X-linked glomerulopathy, Alport syndrome (AS), while homozygous or compound heterozygous variants in the COL4A3 or the COL4A4 gene cause autosomal recessive AS. AS usually leads to progressive kidney failure before the age of 40-years when left untreated. People who inherit heterozygous COL4A3/A4 variants are at-risk of a slowly progressive form of the disease, starting with microscopic hematuria in early childhood, developing Alport spectrum nephropathy. Sometimes, they are diagnosed with benign familial hematuria, and sometimes with autosomal dominant AS. At diagnosis, they often show thin basement membrane nephropathy, reflecting the uniform thin glomerular basement membrane lesion, inherited as an autosomal dominant condition. On a long follow-up, most patients will retain normal or mildly affected kidney function, while a substantial proportion will develop chronic kidney disease (CKD), even kidney failure at an average age of 55-years. A question that remains unanswered is how to distinguish those patients with AS or with heterozygous COL4A3/A4 variants who will manifest a more aggressive kidney function decline, requiring prompt medical intervention. The hypothesis that a subgroup of patients coinherit additional genetic modifiers that exacerbate their clinical course has been investigated by several researchers. Here, we review all publications that describe the potential role of candidate genetic modifiers in patients and include a summary of studies in AS mouse models.

#3618 CLINICAL SIGNIFICANCE OF MILD ABNORMALITIES IN GLOMERULAR FILTRATION BARRIER (GBM) DIAGNOSED BY RENAL BIOPSY

Abstract Background and Aims The well-known GBM abnormalities without immunological reaction are Alport syndrome (ALPORT) and thin basement membrane disease (TMD). Two diseases occasionally share common genetic abnormalities, although, clinical outcomes are worse in ALPORT. Sometimes, there are also mild abnormalities of GBM unable to be classified into ALPROT or TMD without any immunological or hemodynamical abnormalities in the kidney. We compared clinical and pathologic findings among ALPORT, TMD, and GBM abnormalities not classified (GBM NOS) diagnosed by renal biopsy. Method Among 18,134 adult patients who underwent kidney biopsy from 1979 to 2018 in 17 hospitals in Korea (number of IRB: B1707/408-106), we selected patients with ALPORT, TMD, or GBM NOS diagnosed by renal biopsy. GBM NOS was defined as any abnormality of GBM structure of lamellation, thickness, and integrity by electron microscopic examination, which are not able to classify into specific pathologic diagnosis. We excluded ALPORT, TMD, and GBM NOS combined to the other pathologic diagnosis or showing electron dense deposit. We collected clinicopathologic findings at renal biopsy and the last visit to clinics and incidences of mortality and end stage renal disease during follow-up period. Results There were 179 (0.99%) TMDs, 33 (0.18%) ALPORTs, and 8 (0.04%) GBM NOSs among all patients. The clinico-pathologic findings were presented in Table 1. Age and gender proportion at renal biopsy were not different among groups. Patients with ALPORT showed the lowest level of estimated glomerular filtration rate by MDRD equation (eGFR) (p&amp;lt;0.001) and the highest level of urine protein to creatinine ratio (UPCR) (p&amp;lt;0.001). Patients with GBM NOS had similar levels of eGFR and UPCR compared to patients with TMD at renal biopsy. Under microscopic examinations, segmental sclerosis in glomeruli was most prevalent in patients with ALPORT compared to patients with the other disease. Global sclerosis in glomeruli was prevalent in patients with ALPORT and GBM NOS. Other glomerular changes, such as cellular infiltration and matrix change in mesangium, and ischemic glomerular change, were not different among groups. Patients with GBM NOS showed more severe change of interstitial fibrosis and tubular atrophy compared to patients with TMD but similar to patients with ALPORT. Patients with GBM NOS did not show vascular changes. Podocyte foot process effacement was present more frequently in patients with GBM NOS or ALPORT compared to patient with TMD (p&amp;lt;0.001). During 97.9 ± 61.8 months of follow-up period, one (3.0%) patient was dead and 10 (30.3%) patients must have renal replacement therapy (RRT) among patients with ALPORT. The other patients with GBM NOS or TMD had survived and did not need RRT. Conclusion Clinical findings of GBM NOS were similar to TMD, although pathologic changes of glomerular global sclerosis and tubulo-interstitial changes are more severe in GBM NOS compared to TMD. GBM changes in GBM NOS might be related to tubulo-interstitial damages which needs to be defined with more studies.

#4113 CLINICAL EXOME SEQUENCING AS A TOOL FOR GENETIC DIAGNOSIS OF ALPORT SYNDROME AND THIN BASEMENT MEMBRANE DISEASE

Abstract Background and Aims Alport syndrome (AS) is an inherited nephropathy caused by pathogenic variants in COL4A3 (autosomal dominant -AD- and autosomal recessive -AR- inheritance), COL4A4 (AR) and COL4A5 (X-linked dominant -XLD). It is characterized by glomerular nephropathy with hematuria progressing to end-stage renal disease, frequently associated with sensorineural deafness and ocular anomalies. In thin basement membrane disease (TBMD) most patients are asymptomatic and are incidentally noted to have microhematuria, mild proteinuria, and occasionally gross hematuria, with normal renal function. This condition is caused by pathogenic variants in COL4A3 and COL4A4 (AD). We have systematically applied genetic testing to pediatric and adult patients with clinical features suggestive of AS (Table 1). Method Clinical exome sequencing (CES) was performed on a cohort of 95 patients referred to our center from 2019 to 2022. Analyses were performed on an in-silico designed panel including 523 renal genes. Whenever possible, identified variants were segregated by Sanger sequencing. Results We identified causative (C4/C5) variants in 43 (45.3%) patients. Among them, genetic diagnosis of AS was obtained in 25 patients (58.1%): one (4.0%) with biallelic variants in COL4A4, 13 (52.0%) with COL4A5 variants (6 females and 7 males), 5 (20.0%) with COL4A3 variants (4 heterozygous and 1 homozygous) and 6 (24.0%) cases with coexisting variants in one or more other collagen genes. In 16 (37.2%) patients, we found causative monoallelic variants compatible with TBMD: 12 (75.0%) patients with variants in COL4A4 and 4 (25.0%) in COL4A3. Lastly, in 2 cases, we found variants in COL4A1 (#120130) and MYH9 (#160775), both genes associated with other renal diseases with a clinical presentation partially overlapping with AS. In additional 16 cases (16.8%), we identified C3 variants in collagen genes. In the remaining patients, genetic testing was negative (14 patients, 14.7%) or inconclusive (22, 23.2%). Of note, we identified 6 families with digenic AS. In 4 of them - 3 with COL4A5/COL4A4 variants and one with COL4A3/COL4A4 variants – the co-existence of two variants was associated to earlier renal failure, compared to family members bearing a single variant. In contrast, in the remaining 2 families (33.3%), members with COL4A5/COL4A3 missense variants had a milder phenotype compared to male family members with the single variant COL4A5 variant. In families with digenic AS, at least one variant involved the substitution of a glycine with another amino acid in the triple helix domain, frameshift indel variants in the triple helix domain or inframe indel in the collagen IV domain non-collagenous. To better understand the impact of multiple variants on collagen structure, computational studies are currently ongoing. Conclusion CES is a powerful tool to clearly define the diagnosis when AS or TBMD are suspected, as witnessed by a detection rate of 43.2%. Genetic diagnosis allows to identify the causative gene offering the possibility of extending diagnosis to other family members, also in the prenatal setting. In addition, it offers the possibility to identify multiple variants in collagen genes associated with digenic AS, as well as to find variants in other genes implicated in kidney disease. Notably, the presence of digenic variants is not necessarily predictive of a worse disease outcome. For all these reasons, molecular diagnosis can be useful to improve clinical management, to calculate recurrence risk and to better define the prognosis of the patient.

#2909 DOUBLE GLOMERULOPATHY OF THIN BASEMENT MEMBRANE DISEASE AND IGA NEPHROPATHY PRESENTING WITH NEPHROTIC SYNDROME IN AN ADULT WOMAN

Abstract Background and Aims We report a case of a 44-year-old woman with no known comorbid conditions who, eventually was diagnosed to have simultaneous occurrence of IgA nephropathy and thin basement membrane disease atypically presenting with heavy proteinuria, hypoalbuminemia, bipedal edema, ascites, bilateral pleural effusion, and hemodynamic instability, but with a surprisingly normal light microscopic findings and an Oxford MEST C score of 0 on renal biopsy. who, clinically improved with immunosuppression and angiotensin II receptor blocker therapy. Physical exam revealed an overweight female, with facial swelling, no rashes, periorbital and a Grade II bipedal pitting oedema. The presence of decreased breath sounds and dullness on percussion on both lower lung fields were also observed. Shifting dullness and fluid wave on abdominal exam. The patient was afebrile, no desaturation at room air and a palpatory blood pressure of 70 mmHg. Secondary causes of glomerulonephritis such as Hepatitis B and Hepatitis C infection were ruled out. Antinuclear antibody (ANA) test, compliment (C3 and C4) with anti-dsDNA determination to rule out SLE were unremarkable as well. Serum creatinine was elevated at 2.63 with estimated GFR of 22 mL/min. Further work-up with 24-hour total urine protein was 22,026.2 grams/day The patient underwent percutaneous kidney biopsy which showed IgA Nephropathy with a MEST C score of 0 with simultaneous occurrence of thin basement membrane disease. Method/ Treatment Pulse steroid therapy was started with Methylprednisone 1-gram intravenous infusion once daily for 3 days followed with Prednisone (1mg/kg/day) 60 mg a day and a low-dose of Angiotensin receptor blocker, Candesartan, 4mg once daily for the proteinuria. Results After 8 weeks of corticosteroid and ARB therapy, patient was seen at the clinic with resolution of bipedal edema, no demonstrable ascites and clear breath sounds on chest and lungs auscultation. Blood pressure was 130/80 with no proteinuria on dipstick, random urine protein: creatinine ratio (UPCR) of 0.07, creatinine of 0.97 mg/dl and BUN of 17 mg/dl. Prednisone was then reduced with the goal to taper the dose for a period of 6 months. Conclusion/ Significance Both IgA nephropathy and thin basement membrane disease usually presents with hematuria, hypertension, and varying degrees of proteinuria. However, nephrotic syndrome as its presentation has not been well characterized to date, and whether this adds to the mortality is not yet established. Although the most widely accepted system, the Oxford MEST-C scores is utilized in predicting renal outcomes, its role in double glomerulopathies has not been validated. Even though many investigators have indicated that the presence of heavy proteinuria at the initial evaluations was almost always associated with progressive renal failure and that there is no effective medical treatment aside from supportive care, in this study, a trial of immunosuppression yielded improved clinical outcomes.

#3067 ALPORT SYNDROME NATURAL HISTORY FROM THE RADAR REGISTRY: ASSOCIATIONS WITH GENE, VARIANT TYPE AND SEX

Abstract Background and Aims Alport Syndrome (AS) is caused by pathogenic variants in COL4A3, COL4A4 &amp; COL4A5 genes. The clinical course of AS can be highly variable, depending on gene affected, mutation type and Male (M) or Female (F) sex. Proteinuria is associated with worse prognosis. Previous genotype-phenotype correlation studies have shown protein length altering variants are associated with a more severe phenotype in males with COL4A5 variants, however studies in females have shown contradicting results. This study aims to describe demographics and investigate renal outcomes associated with pathogenic variant type in M vs F AS patients, using longitudinal data from the National Registry of Rare Kidney Diseases (RaDaR) which recruits patients at 108 UK renal clinics. Method RaDaR is linked to Regional Genetics hubs for clinical genetic reports, renal IT systems for routine test results and the UK Renal Registry for Renal Replacement Therapy (RRT) initiation data. eGFR was calculated using CKD-EPI Cr equation (2021) or Schwartz equation for those ≤ 16 yrs. eGFR slope was calculated over the last 3 years (or 3 years prior to End Stage Kidney Disease (ESKD)/death for patients who reached those outcomes), with patients required to have a minimum of 4 values over 2 years. For genotype data, variants classified in clinically issued reports as “Pathogenic” or “Likely Pathogenic” were included and classed as: 1. Protein Length Altering and 2. Missense variants. Where both protein length altering and missense variants in the same gene were identified in an individual, the missense variant was used. Data are presented as percentages for categorical variables and mean ± SD for continuous variables. Kaplan Meier analysis and the log rank statistic were used to compare age at RRT start, stratified by gender and variant types. Lower quartile (LQ) (25%) estimates are presented where too few events have occurred to calculate a median (50%) estimate. Results Between Jan 2013–Jul 2022, 920 AS patients were recruited; 53% M vs. 47% F. Genetic test reports were available for 343/920 (37%) patients. 294/343 (86%) reported a pathogenic or likely pathogenic variant, 140 protein length altering and 154 missense. Individuals with 2x COL4A3/COL4A4 variants were youngest at RRT start, whilst females with COL4A5 variants were oldest (LQ 20.1 vs 60.9 yrs, log-rank p&amp;lt;0.0001) (Figure 1a). COL4A5 variants Males with protein length altering COL4A5 variants were younger at ESKD compared with missense variants (median age 30.2 vs 52.1 yrs, p = 0.01). Conversely 0/36 females with protein length altering COL4A5 variants had started RRT, compared with 6/37 with missense variants (p = 0.01) (Figure 1bi-ii). COL4A3 or COL4A4 variants Whilst females with heterozygous protein length altering COL4A3/4 variants were younger at RRT start vs. those with missense variants (LQ 47.1 vs 64.9 yrs, p = 0.05), age at RRT start did not differ by mutation type for males with COL4A3/4 heterozygous variants (Figure 1b iii-iv). However, males with homozygous or 2x protein length altering COL4A3/4 variants were younger at RRT start than those with missense variants (median age 20.1 vs 24.3 yrs, P = 0.05), whilst no difference was observed by variant type for females with 2x COL4A3/4 variants (Figure 1b v-vi). Proteinuria and eGFR slope were generally correlated with age at ESKD. Conclusion The observed effect of pathogenic variant type on renal outcomes varied by gene affected, number of mutations and sex. The relatively reduced severity among females harbouring a protein length altering COL4A5 variant may represent an effect of skewed X-inactivation or a missense gain-of-function mechanism. RaDaR recruits patients with a clinical diagnosis of Alport syndrome or Thin Basement Membrane Nephropathy; ascertainment of individuals with a single COL4A3 or COL4A4 variant is therefore likely to favour those with more severe kidney disease. Linkage of the RaDaR AS cohort with genetic report data is ongoing; further correlations may be observed with larger numbers.

#5400 CLINICAL PROFILE, MICROSCOPIC FEATURES AND OUTCOMES OF PATIENTS WITH MINOR GLOMERULAR ABNORMALITIES AT NKTI: A 10-YEAR DESCRIPTIVE CROSS-SECTIONAL STUDY

Abstract Background and Aims Minor Glomerular Abnormality (MGA) is a kidney biopsy diagnosis characterized by the presence of minor structural abnormalities which, are not sufficient to be classified under the other known glomerular diseases. Although usually considered a benign renal problem, few literatures mentioned it may cause deterioration of kidney function in the long term. The objective of this study is to describe the clinical profile, microscopy findings, treatment outcome and renal survival of patients diagnosed with Minor Glomerular Abnormalities in NKTI from 2005 to 2015. Method This is a Retrospective, Descriptive, Cross-sectional study which included 190 patients who were diagnosed with MGA. Demographic, clinical profile, CKD classification and treatment outcomes were presented using descriptive statistics while renal survival was analysed using Fisher's Exact Test. Based on the level of proteinuria and serum creatinine, treatment outcomes of the patients were classified as either with (1) complete response [if with reduction in proteinuria to ≤ 300 mg/day (&amp;lt;/ = 0.3 g/day) and a creatinine clearance ≥ 80 ml/min with no reduction from baseline] (2) partial response [proteinuria between 300 mg and 3 g/day OR by at least 50% from baseline level and with no deterioration in creatinine clearance], (3) relapse[patients whom (a) after a complete response to treatment (baseline proteinuria ≤ 300 mg/day), there is an increase in proteinuria to ≥ 1.0 g/day or (b) after a partial response (baseline proteinuria between 300 mg and 3 g/day), there is an increase in proteinuria to ≥ 3 g/day or increase in proteinuria by ≥ 50%, or (c) after a partial response (baseline proteinuria &amp;gt; 3 g/day), there is an increase in proteinuria by ≥ 50%], or (4) unsatisfactory response to treatment if complete or partial response is not met. Renal survival was assessed by classifying the renal status of the patients as either with (1) Normal or Mildly Decreased GFR [CKD 1-2] or (2) with moderately decreased kidney function to renal failure [CKD 3-5]. Results The mean age of the MGA patients at diagnosis was 37 years old. Of the 190 subjects of this study, 53.68% were female and 73.16% has no comorbidities. Proteinuria was the most common reason for consult (66.32%), followed by hematuria (38.42%) and worsening renal function (14.21%). Most of the patients had Nephrotic Syndrome (39%) before biopsy. The top three Electron Microscopy findings were Minimal Change Disease (24.4%) Segmental Podocyte Foot Processes Effacement (True MGA) at 27.4% and Thin Basement Membrane Disease (22.6%). At one year and five years, 13.68% and 4.74% respectively have complete response to treatment while 22.10% and 13.15% respectively have either partial or unsatisfactory response to treatment. Eighty patients (86.02%) at one year and 33 patients (78.57%) at 5 years remain at CKD 1-2. When the renal survival of the True MGA and Non-MGA patients were compared using Fisher Exact Test, there was no significant difference between the two groups (p values of 0.080 at 1 year and 0.99 at 5 years). Conclusion Minimal Change Disease is the most common Electron Microscopy diagnosis. Majority have good renal survival, but few progressed to CKD 3-5. Survival between MGA and Non-MGA patients are similar.

#4220 NOT JUST AIN? OTHER IMMUNE-CHECKPOINT INHIBITOR LESIONS: A SINGLE INSTITUTION'S EXPERIENCE OVER 12 YEARS

Abstract Background and Aims Immune-checkpoint inhibitors (ICI) are antineoplastic therapies that unleash the immune-system and have revolutionized cancer outcomes. Though they are a drug class with great potential, their effects are not always limited to the malignant cells and can cause a spectrum of immune-related adverse events (irAEs). Roughly 5% of patients will experience acute kidney injury (AKI) with acute interstitial nephritis (AIN) being the predominant pathologic lesion. Glomerular and vascular lesions are thought to be rare and, as a result, not well-defined. We aim to document glomerular and vascular pathology in this context at our institution. Method We performed a retrospective review of internal native renal biopsies from 2010-2022 that had known ICI treatment. Standard processing of renal biopsies included light microscopy (LM), immunofluorescence (IF), and electron microscopy (EM). We reviewed the biopsies and pathology reports, as well as clinical data from patient electronic medical records. Results Our search yielded 32 kidney biopsies; of those, most had AIN (27/32), 8 cases had glomerular pathology, and 1 case had arterial endotheliitis. Of the cases with glomerular pathology, 4 had subacute glomerular thrombotic microangiopathy (TMA). Among the cases with TMA, 2 were also treated with vascular endothelial growth factor inhibitor (VEGFi),1 with gemcitabine, and 1 with BRAF-inhibitor therapy. Further, only one of these patients had complement testing, which did not reveal definitive deficiency. One patient had proliferative glomerulonephritis with monoclonal immunoglobulin deposits (PGNMID), IgG-κ. One patient had IgA nephropathy (IgAN) and another had membranous nephropathy (MN). Most patients were biopsied for rising serum creatinine (SCr) (median: 1.6 mg/dL; range: 0.55-5.5); 1 had nephrotic-range proteinuria (median urine protein/creatinine ratio (UPCR): 0.3; range: 0.12-5.64). One, with thin basement membrane disease, had significant hematuria (&amp;gt;100 RBC/hpf). Five patients had irAEs in other organs. 7/9 patients died or entered hospice care (median: 14 months post-biopsy; range: 2-48 months) due to malignancy progression. Of the deceased patients, 2 were dialysis-dependent. Of the living patients, 1 has dialysis-independent chronic kidney disease (CKD). Conclusion Overall, glomerular and vascular pathology due to ICI-use appears rare though difficult to isolate since other drugs and neoplasia can cause similar lesions. In our cohort, TMA was the most common glomerular lesion, though direct causation due to ICI is difficult to prove. In all patients with TMA, we identified concurrent use of other known causative drugs. IgAN and MN have been reported in association with neoplasia. PGNMID has been reported in a variety of malignant and nonmalignant conditions, and ICI-use could be a triggering factor. In conclusion, the patterns of glomerular injury seen in our cohort may be secondary to other concurrent risk factors other than ICI use. Arterial endotheliitis appears to be a rare but significant secondary effect to ICI-use.

The same heterozygous Col4A4 mutation triggered different renal pathological changes in Chinese family members.

Background: Mutations in the collagen components of the glomerular basement membrane (GBM) often lead to hereditary glomerulonephritis. Previous studies have identified that autosomal dominant mutations of Col4A3, Col4A4 or Col4A5 are associated with thin basement membrane nephropathy (TBMN), Alport syndrome and other hereditary kidney diseases. However, the genetic mutations underlying other glomerulonephritis types have not been elucidated. Methods: In this study, we investigated a Chinese family with hereditary nephritis using the methods of genetic sequencing and renal biopsy. Genomic DNA was extracted from peripheral blood of the proband and her sister, and subsequently was performed genetic sequencing. They were found to have the similar mutation sites. Other family members were then validated using Sanger sequencing. The proband and her sister underwent renal puncture biopsies, and experienced pathologists performed PAS, Masson, immunofluorescence, and immunoelectron microscopic staining of the kidney tissue sections. Results: Through genetic sequencing analysis, we detected a novel heterozygous frameshift mutation c.1826delC in the COL4A4 (NM_000092.4) gene coding region, and 1 hybrid missense variation c.86G>A (p. R29Q) was also detected in the TNXB (NM_019105.6) gene coding region in several members of this Chinese family. Interestingly, we found that the same mutations caused different clinical features and distinct pathological changes in individual family members, which confirmed that pathological and genetic testing are crucial for the diagnosis and treatment of hereditary kidney diseases. Conclusion: In this study, we found a novel heterozygous mutation in Col4A4 and co-mutations of the TNXB gene in this Chinese family. Our study indicated that the same Col4A4 mutated variants produced different pathological and clinical changes in different family members. This discovery may provide novel insights into the study of hereditary kidney disease. In addition, new genetic biology techniques and renal biopsy of individual family members are essential.

Direct observation of epoxy resin blocks for renal biopsy by low-vacuum scanning electron microscopy

To improve the resolution of low-vacuum scanning electron microscopy (LVSEM), the epoxy resin block for the transmission electron microscopy (TEM) was observed directly with LVSEM. After observing ultrathin sections from renal biopsies of IgA nephropathy, membranous nephropathy, lupus nephritis, diabetic nephropathy (DM), thin basement membrane disease (TBMD), Alport’s syndrome, Fabry’s disease, and renal amyloidosis, the epoxy resin blocks of the same sites were observed by LVSEM and compared. The LVSEM image of the epoxy resin block corresponds to the negative of the TEM image, and when the gradation is reversed, the LVSEM image was comparable to the TEM image. At a low magnification of 100 ×, the entire specimen, including the glomerulus, was obtained. LVSEM at 5000 × magnification was sufficient to identify paramesangial deposits in IgA nephropathy and subepithelial electron-dense deposits (EDD) and spikes in membranous nephropathy. Glomerular basement membrane thickening in DM and thinning in TBMD could be sufficiently diagnosed with LVSEM at 6000 ×. Accumulation of ceramide in Fabry's disease was easily identified, but amyloid fibril could not be identified by LVSEM. LVSEM of renal biopsy epoxy resin blocks can replace TEM up to moderate magnification.

Pediatric glomerulopathy after COVID-19 vaccination: A case series and review of the literature

Cases of glomerulopathy after COVID-19 vaccination have been reported in the adult population, while only a few cases have been reported in children and adolescents. For better understanding of this association in pediatric population, we aimed to describe clinical course of patients with glomerulopathy within 60 days of COVID-19 vaccination who were under followed up in the pediatric nephrology department of National Taiwan University Children's Hospital. We reviewed the clinical characteristics, vaccine types, and outcomes of patients with newly diagnosed glomerular diseases or relapse of underlying glomerulopathy within 60 days after COVID-19 vaccination at our facility between January 2021 and July 2022. Thirteen pediatric patients were found to have newly diagnosed glomerular diseases or relapse from their underlying glomerulopathy after receiving their first, second, or third COVID-19 vaccines in our facility. Of the five pediatric patients with newly diagnosed glomerulopathy after vaccination, thin basement membrane nephropathy, idiopathic nephrotic syndrome, and hematuria have been identified. Seven patients had relapse episodes of underlying nephrotic syndrome and one patient with underlying isolated microscopic hematuria developed subnephrotic proteinuria after COVID-19 vaccination. All patients experienced remission or improvement with either immunosuppressive or conservative treatment during the follow-up period. This is the largest case series to date of pediatric glomerulopathy after COVID-19 vaccination. From our report, patients with either newly diagnosed or relapse of glomerulopathy after vaccination had good outcomes, and receiving vaccination to prevent COVID-19 infection or complications should be encouraged in pandemic era under close monitoring kidney manifestations.