Complement Factor H-related Protein Research Articles

Causal association of immune effector proteins with sepsis: A Mendelian randomization study.

Sepsis is an infection-induced systemic inflammatory response syndrome. Immune regulation plays a crucial role in sepsis. We looked into the link between immune effector-related proteins and sepsis in this study by using both univariate and multivariate Mendelian randomization (MR) analyses. We accessed and collected data from the Integrative Epidemiology Unit's Open About Sepsis genome-wide association study database. The 6 immune effector-associated proteins each contained 10,534,735 single-nucleotide polymorphisms from 3301 samples. Using the weighted median, MR-Egger, simplex, inverse-variance weighting, and weighted mode methods, univariate MR then investigated the link between complement factor H-related protein-5 (CFHR5), Fc epsilon receptor II (FCER2), granzyme B (GZMB), major histocompatibility complex, class II, DQ alpha (HLA-DQA2), mannose-binding lectin 2 (MBL2), or myeloperoxidase (MPO) and sepsis. In the inverse-variance weighted results, the P values of all 6 immune effector-related proteins were <0.05, suggesting a possible causal relationship between them and sepsis. MBL2 (odds ratio [OR] = 1.046) was a risk factor for sepsis, while the other proteins (FCER2: OR = 0.922; GZMB: OR = 0.908; CFHR5: OR = 0.858; HLA-DQA2: OR = 0.896; MPO: OR = 0.875) were safety factors. By revealing a causal link between sepsis and CFHR5, FCER2, GZMB, HLA-DQA2, MBL2, or MPO, our study offers an essential resource for additional investigations on the subject.

Immune and regulative characterization of complement-related gene cfhl5 in response to Vibrio harveyi challenge in Cynoglossus semilaevis

Complement factor H-related protein (CFHR) plays an important role in regulating complement activation and defensive responses. The function of CFHR2 (complement factor H related 2), a member of the CFHR family, in fish remains unclear. Here, we report the genetic relationship, expression characteristics and regulatory mechanism of cfhl5 (complement factor H like 5) gene, which encodes CFHR2 in Chinese tongue sole. We observed that the cfhl5 gene was widely expressed in several tissues, such as brain, heart and immune organs, and was most abundantly expressed in liver. After injection with Vibrio harveyi, the expression of cfhl5 was up-regulated significantly in liver, spleen and kidney at 12 or 24 hours post infection (hpi), suggesting an involvement of this gene in the acute immune response. Knockdown of cfhl5 in liver cells significantly up-regulated the expression of the pro-inflammatory cytokines tnf-α (tumor necrosis factor-alpha) and il1β (interleukin-1beta), the immunomodulatory factor il10 (interleukin-10) and the lectin complement pathway gene masp1 (MBL-associated serine protease 1), and down-regulated the expression of complement components c3 (complement 3) and cfi (complement factor I). In our previous work, we found that cfhl5 gene was significantly higher methylated and lower expressed in the resistant family compared with the susceptible family. Therefore, we used dual-luciferase reporter system to determine the effect of DNA methylation on this gene and found that DNA methylation could inhibit the promoter activity to reduce its expression. These results demonstrated that the expression of cfhl5 is regulated by DNA methylation, and this gene might play an important role in the immune response by regulating the expression of cytokines and complement components genes in Chinese tongue sole.

Clinical characteristics and genetic profile of complement system in renal thrombotic microangiopathy in patients with severe forms of arterial hypertension

The spectrum of diseases characterized by the development of renal thrombotic microangiopathy (TMA) encompasses the malignant hypertension (MHT). TMA in MHT has conventionally been regarded as a variation of secondary TMA, the treatment of which is restricted to the stabilization of blood pressure levels, a measure that frequently fails to prevent the rapid progression to end-stage renal disease in patients. Nevertheless, there exists a rationale to suggest that, in certain instances, endothelial damage in MHT might be rooted in the dysregulation of the complement system (CS), thereby presenting potential opportunities for the implementation of complement-blocking therapy. To study clinical manifestations and genetic profile of CS in patients with morphologically confirmed renal TMA combined with severe AH. 28 patients with morphologically verified renal TMA and severe AH were enrolled to the study. Patients with signs of microangiopathic hemolysis and thrombocytopenia were not included in the study due to possible compliance with the criteria for atypical hemolytic uremic syndrome (aHUS). The prevalence of rare genetic defects (GD) of the CS was assessed by molecular genetic analysis (search for mutations in the clinically significant part of the human genome - exome) by next-generation sequencing technology (NGS). GD of CS were detected in a quarter of patients. Rare genetic variants classified as "likely pathogenic" including defects in CFI, C3, CD46, CFHR4, CFHR5 genes were detected in five cases. Two patients were found to have chromosomal deletions containing CFH-related proteins genes (CFHR1, CFHR3). Rare variants of CS genes linked to aHUS were found in 25% of patients with renal TMA, the genesis of which was originally thought to be secondary and attributed to MHT, with partial or complete absence of hematological manifestations of microangiopathic pathology. The key to confirming TMA associated with MHT, particularly in the absence of microangiopathic hemolysis and thrombocytopenia, elucidating its nature, and potentially effective complement-blocking therapy in patients with GD of CS, appears to be a genetic study of CS combined with a morphological study of a renal biopsy.

Genetic variants contribute to modulation of renal function in patients with immune thrombotic thrombocytopenic purpura

AbstractImmune thrombotic thrombocytopenic purpura (iTTP) is a potentially fatal blood disorder, resulting from autoantibodies against ADAMTS13. However, the contribution of genetic variations that may modulate its clinical presentations remains unknown. This study aimed to determine the potential contribution of variants in the genes associated with coagulation, complement activation or regulation, and platelet activation to pathophysiology of iTTP. Multicenter case series, whole-exome sequencing, and bioinformatic approaches were used. We focused on analysis of 20 genes that are involved in regulation of coagulation (eg, ADAMTS13, THBD, MMACHC, INF2, and PLG), complement activation (eg, C3, C3AR1, C5, CFB, CFH, CFI, C4BPA, CD46 [MCP], CD59, and CFHR1-CFHR5), and platelet activation (eg, DGKE) from 40 adult patients with iTTP. Multiple genetic variations were identified in 12 of 20 genes of interest. More than 80% of patients harbored genetic variants in CFI, CFH, C5, and ADAMTS13; 15% to 55% of patients had variants in C3, INF2, CFHR5, and PLG; and <10% of patients had variants in CD46, C3AR1, DGKE, and THBD. Of these, the variants in C5 are associated with a more favorable renal function, whereas the variants in DGKE are associated with more persistently elevated creatinine levels. These results demonstrate that variants in the genes involved in coagulation, complement, and platelet activation are common in patients with iTTP, which may contribute to phenotypical modulations of or predispose to iTTP resulting from severe ADAMTS13 deficiency.

Hearing loss and its association with the proteome of perilymph, cerebrospinal fluid, and tumor tissue in patients with vestibular schwannoma

This study examined the association between hearing loss in sporadic vestibular schwannoma patients and the proteome of perilymph (PL), cerebrospinal fluid (CSF), and vestibular schwannoma. Intraoperative sampling of PL and of CSF, and biopsy of vestibular schwannoma tissue, was performed in 32, 32, and 20 patients with vestibular schwannoma, respectively. Perilymph and CSF in three patients with meningioma and normal hearing were also sampled. The proteomes were identified by liquid chromatography coupled to high-resolution tandem mass spectrometry. Preoperative hearing function of the patients was evaluated with pure tone audiometry, with mean values at frequencies of 500, 1000, 2000, and 4000 Hz (PTA4) in the tumor-affected ear used to delineate three hearing groups. Analysis of the PL samples revealed significant upregulation of complement factor H-related protein 2 (CFHR2) in patients with severe to profound hearing loss after false discovery rate correction. Pathway analysis of biofunctions revealed higher activation scores in the severe/profound hearing loss group of leukocyte migration, viral infection, and migration of cells in PL. Upregulation of CFHR2 and activation of these pathways indicate chronic inflammation in the cochlea of vestibular schwannoma patients with severe to profound hearing loss compared with patients with normal hearing or mild hearing loss.

An unusual case of adult-onset still’s disease complicated with anti-complement factor H antibodies associated atypical haemolytic uraemic syndrome

BackgroundAtypical haemolytic uremic syndrome (aHUS) is an uncommon form of thrombotic microangiopathy (TMA). However, it remains difficult to diagnose the disease early, given its non-specific and overlapping presentation to other conditions such as thrombotic thrombocytopenic purpura and typical HUS. It is also important to identify the underlying causes and to distinguish between primary (due to a genetic abnormality leading to a dysregulated alternative complement pathway) and secondary (often attributed by severe infection or inflammation) forms of the disease, as there is now effective treatment such as monoclonal antibodies against C5 for primary aHUS. However, primary aHUS with severe inflammation are often mistaken as a secondary HUS. We presented an unusual case of adult-onset Still’s disease (AOSD) with macrophage activation syndrome (MAS), which is in fact associated with anti-complement factor H (anti-CFH) antibodies related aHUS. Although the aHUS may be triggered by the severe inflammation from the AOSD, the presence of anti-CFH antibodies suggests an underlying genetic defect in the alternative complement pathway, predisposing to primary aHUS. One should note that anti-CFH antibodies associated aHUS may not always associate with genetic predisposition to complement dysregulation and can be an autoimmune form of aHUS, highlighting the importance of genetic testing.Case presentationA 42 years old man was admitted with suspected adult-onset Still’s disease. Intravenous methylprednisolone was started but patient was complicated with acute encephalopathy and low platelet. ADAMTS13 test returned to be normal and concurrent aHUS was eventually suspected, 26 days after the initial thrombocytopenia was presented. Plasma exchange was started and patient eventually had 2 doses of eculizumab after funding was approved. Concurrent tocilizumab was also used to treat the adult-onset Still’s disease with MAS. The patient was eventually stabilised and long-term tocilizumab maintenance treatment was planned instead of eculizumab following haematology review. Although the aHUS may be a secondary event to MAS according to haematology opinion and the genetic test came back negative for the five major aHUS gene, high titre of anti-CFH antibodies was detected (1242 AU/ml).ConclusionOur case highlighted the importance of prompt anti-CFH antibodies test and genetic testing for aHUS in patients with severe AOSD and features of TMA. Our case also emphasized testing for structural variants within the CFH and CFH-related proteins gene region, as part of the routine genetic analysis in patients with anti-CFH antibodies associated aHUS to improve diagnostic approaches.

Contributions of Lipid-Related Metabolites and Complement Proteins to Early and Intermediate Age-Related Macular Degeneration

ObjectiveOur objective was to determine the effects of lipids and complement proteins on early and intermediate age-related macular degeneration (AMD) stages using machine learning models by integrating metabolomics and proteomic data. DesignNested case-control study. Subjects, Participants, and/or ControlsThe analyses were performed in a subset of the Singapore Indian Chinese Cohort (SICC) Eye Study. Among the 6,753 participants, we randomly selected 155 AMD Indian and 155 Chinese cases and matched them with 310 controls on age, sex and ethnicity. MethodsWe measured 35 complement proteins and 56 lipid-related metabolites using Mass Spectrometry and Nuclear Magnetic Resonance, respectively. We first selected the most contributing lipids and complement proteins to early and intermediate AMD using random forest models. Then we estimated their effects using a multinomial model adjusted for potential confounders. Main Outcome MeasuresAMD was classified using the Beckman classification system. ResultsAmong the 310 individuals with AMD, 166 (53.5%) had early and 144 (46.5%) intermediate AMD. Firstly, high-density lipoprotein (HDL) particle diameter was positively associated with both early and intermediate AMD (ORearly=1.69 [1.11, 2.55] and ORintermediate=1.72 [1.11, 2.66] per 1-SD increase in HDL diameter). Secondly, five complement proteins: complement protein 2 (C2), complement C1 inhibitor (ICI1), complement protein 6 (C6), complement protein 1QC (C1QC) and complement factor H-related protein 1 (FHR1), were associated with AMD. C2 was positively associated with both early and intermediate AMD (ORearly=1.58 [1.08, 2.30] and ORintermediate=1.56 [1.04, 2.34]). C6 was positively (ORearly=1.41 [1.03, 1.93]) associated with early AMD. However, IC1 was negatively associated with early AMD (ORearly=0.62 [0.38, 0.99]), while C1QC (ORintermediate=0.63 [0.42, 0.93]) and FHR1 (ORintermediate=0.73 [0.54, 0.98) were both negatively associated with intermediate AMD. ConclusionsWhile both HDL diameter and C2 levels show associations with both early and intermediate AMD, dysregulations of IC1, C6, C1QC, and FHR1 are only observed at specific stages of AMD. These findings underscore the complexity of complement system dysregulation in AMD, which appears to vary depending on the disease severity.

The complement factor H-related protein-5 (CFHR5) exacerbates pathological bone formation in ankylosing spondylitis.

Ankylosing spondylitis (AS) is a chronic inflammatory disease, characterized by excessive new bone formation. We previously reported that the complement factor H-related protein-5 (CFHR5), a member of the human factor H protein family, is significantly elevated in patients with AS compared to other rheumatic diseases. However, the pathophysiological mechanism underlying new bone formation by CFHR5 is not fully understood. In this study, we revealed that CFHR5 and proinflammatory cytokines (TNF, IL-6, IL-17A, and IL-23) were elevated in the AS group compared to the HC group. Correlation analysis revealed that CFHR5 levels were not significantly associated with proinflammatory cytokines, while CFHR5 levels in AS were only positively correlated with the high CRP group. Notably, treatment with soluble CFHR5 has no effect on clinical arthritis scores and thickness at hind paw in curdlan-injected SKG, but significantly increased the ectopic bone formation at the calcaneus and tibia bones of the ankle as revealed by micro-CT image and quantification. Basal CFHR5 expression was upregulated in AS-osteoprogenitors compared to control cells. Also, treatment with CFHR5 remarkedly induced bone mineralization status of AS-osteoprogenitors during osteogenic differentiation accompanied by MMP13 expression. We provide the first evidence demonstrating that CFHR5 can exacerbate the pathological bone formation of AS. Therapeutic modulation of CFHR5 could be promising for future treatment of AS. KEY MESSAGES: Serum level of CFHR5 is elevated and positively correlated with high CRP group of AS patients. Recombinant CFHR5 protein contributes to pathological bone formation in in vivo model of AS. CFHR5 is highly expressed in AS-osteoprogenitors compared to disease control. Recombinant CFHR5 protein increased bone mineralization accompanied by MMP13 in vitro model of AS.

Atypical Haemolytic Uraemic Syndrome in an Infant with Ventricular Septal Defect: A Case Report

Atypical Haemolytic Uraemic Syndrome (aHUS) is a rare type of thrombotic microangiopathy that occurs without Shiga toxin producing bacteria. It is a condition related to complement regulation, which may be genetic or acquired. The complement system’s alternative pathway is commonly implicated, with around 6-10% of cases being caused by autoantibodies directed against factor H. While it typically affects children between 9 to 13-years-old, it can also occur in adults. Many patients do not have circulating Complement Factor H-related proteins 1 and 3 due to a homozygous deletion involving CFHR1 and CFHR3. Authors hereby report a case of a six-month-old female child who was diagnosed with Ventricular Septal Defect (VSD) at one and a half months of age. She presented with pneumonia and subsequently developed haemolytic anaemia with thrombocytopenia, oliguria, and acute kidney failure. She was diagnosed with Antifactor H antibody-mediated HUS. She was treated with plasma therapy, but the patient succumbed due to multiorgan dysfunction.

Biallelic mutations in the CFHR genes underlying atypical hemolytic uremic syndrome in a patient with catastrophic adult-onset Still's disease and recurrent macrophage activation syndrome: A case report

We report the fatal case of a 20-year-old woman with refractory adult-onset Still's disease (AOSD) accompanied by fulminant macrophage activation syndrome (MAS) and atypical hemolytic uremic syndrome (aHUS). Anakinra and tocilizumab temporarily controlled AOSD. In 2021, she presented to ICU with generalized tonic-clonic seizure, lymphocytic aseptic meningitis, and acute kidney injury. Despite hemodialysis and methylprednisolone, she developed another seizure, MAS, and disseminated intravascular coagulation (DIC). Following brief control, MAS flares —reflected by increased plasma CXCL9 and CXCL10— re-emerged and were controlled through dexamethasone, etoposide, cyclosporin and tofacitinib. No mutations were detected in haemophagocytic lymphohistiocytosis (HLH)-associated genes, nor in genes associated with periodic fever syndromes. Post-mortem genetic testing revealed loss-of-function biallelic deletions in complement factor H-related proteins (CFHR) genes, predisposing aHUS. This case underscores the importance of prompt genetic assessment of complement-encoding alleles, in addition to HLH-related genes, in patients with severe AOSD with recurrent MAS and features of thrombotic microangiopathy (TMA).

Mesangial C3 Deposition, Complement-Associated Variant, and Disease Progression in IgA Nephropathy.

IgA nephropathy is the most common primary GN worldwide, with dominant deposition of IgA and co-deposits of complement component 3 (C3). Phenotypes and progression of IgA nephropathy varies among different ethnic populations, while patients with IgA nephropathy from Asia showed more severe clinical phenotypes, active kidney lesions, and rapid progression. Our previous genome-wide association study identified complement factor H ( CFH ) variant rs6677604, tightly linked with the deletion of CFH -related protein 3 and CFH -related protein 1 genes ( ΔCFHR3-1 ), as IgA nephropathy susceptible variant, and additionally revealed its effect on complement regulation in IgA nephropathy. To further explore the effect of rs6677604 on IgA nephropathy progression, here we enrolled a Chinese IgA nephropathy cohort of 1781 patients with regular follow-up for analysis. The rs6677604 genotype was measured, and the genotype-phenotype correlation was analyzed using the t test, the chi-squared test, or the nonparametric test, and the association between rs6677604 genotype or mesangial C3 deposition and IgA nephropathy prognosis was analyzed using Kaplan-Meier analysis and Cox regression. We found that patients with rs6677604-GG genotype had a stronger intensity of mesangial C3 deposition than those with the rs6677604-AA/AG genotype. Patients with IgA nephropathy who had stronger intensity of C3 deposition manifested with more severe clinical and pathological manifestations, including lower eGFR and higher Oxford-M/S/T/C (mesangial hypercellularity, endocapillary cellularity, segmental sclerosis, interstitial fibrosis/tubular atrophy, and crescent) scores. In the survival analysis, stronger intensity of mesangial C3 deposition, but not rs6677604-GG genotypes, was associated with poor long-term kidney outcome in IgA nephropathy. We found that in Chinese patients with IgA nephropathy, variant rs6677604 was associated with mesangial C3 deposition, and mesangial C3 deposition, but not rs6677604, was associated with IgA nephropathy severity and progression.

Predictive value of serum proteomic biomarkers for noise-induced hearing loss.

Early detection of noise-induced hearing loss (NIHL) in patients with long-term noise exposure is vital for improving public health and reducing social burden. However, at present, the diagnosis of NIHL mainly depends on audiometric testing, and the primary test is pure-tone audiometry. Moreover, testing requires professional operators and complex equipment; thus, NIHL is often diagnosed at a later disease stage. Using a liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomic approach, we identified 9 differentially expressed proteins (DEPs), namely, 6 upregulated serum proteins and 3 downregulated serum proteins, in samples from 50 ground crew personnel working at an air force station. Then, according to the results, we predicted that caldesmon (CALD1), myocilin (MYOC), zyxin (ZYX), creatine kinase M-type (CKM), insulin-like growth factor-binding protein 2 (IGFBP2), complement factor H-related protein 4 (CFHR4), prenylcysteine oxidase 1 (PCYOX1), heat shock cognate 71 kDa protein (HSPA8), and immunoglobulin lambda variable 3-21 (IGLV3-21) were associated with NIHL. We selected these DEPs as variables to perform logistic regression. Finally, a logistic regression model was constructed based on IGFBP2, ZYX, CKM, and CFHR4. The area under the curve was 0.894 (95% CI = 0.812 to 0.977). These findings suggested that IGFBP2, ZYX, CKM, and CFHR4 in serum are differentially expressed in NIHL patients and have the potential to be biomarkers for predicting the risk for NIHL. Further experiments in mice showed that ZYX and IGFBP2 in the cochlear were increased after noise exposure. ZYX and IGFBP2 may be involved in the occurrence and development of NIHL.

Complement Dysregulation in Obese Versus Nonobese Polycystic Ovary Syndrome Patients.

Upregulation of complement system factors are reported to be increased in polycystic ovary syndrome (PCOS) and may be due to obesity and insulin resistance rather than inherently due to PCOS. We directly compared complement factors from an obese, insulin-resistant PCOS population to a nonobese, non-insulin-resistant PCOS population in a proteomic analysis to investigate this. Plasma was collected from 234 women (137 with PCOS and 97 controls) from a biobank cohort and compared to a nonobese, non-insulin-resistant population (24 with PCOS and 24 controls). Slow off-rate modified aptamer (SOMA) scan plasma protein measurement was undertaken for the following complement system proteins: C1q, C1r, C2, C3, C3a, iC3b, C3b, C3d, C3adesArg, C4, C4a, C4b, C5, C5a, C5b-6 complex, C8, properdin, factor B, factor D, factor H, factor I, Mannose-binding protein C (MBL), complement decay-accelerating factor (DAF) and complement factor H-related protein 5 (CFHR5). The alternative pathway of the complement system was overexpressed in both obese and nonobese PCOS, with increased C3 (p < 0.05) and properdin (p < 0.01); additionally, factor B increased in obese PCOS (p < 0.01). For inhibitors of this pathway, factor I was increased (p < 0.01) in both slim and obese PCOS, with an increase in CFHR5 and factor H in obese PCOS (p < 0.01). Complement factors iC3b, C3d and C5a, associated with an enhanced B cell response and inflammatory cytokine release, were increased in both slim and obese PCOS (p < 0.05). C3a and its product, C3adesArg, were both significantly elevated in nonobese PCOS (<0.01) but not altered in obese PCOS. Hyperandrogenemia correlated positively with properdin and iC3b in obese PCOS (p < 0.05) but not in nonobese PCOS. There was no association with insulin resistance. BMI correlated positively in both groups with factor B, factor H and C5a. Additionally, in obese PCOS, BMI correlated with C3d, factor D, factor I, CFHR5 and C5a (p < 0.05), and in nonobese PCOS, BMI correlated with properdin, iC3b, C3, C3adesArg, C3a, C4, C5, C5a and C1q. In obese controls, BMI correlated with C3, C3desArg, C3a, C3d, C4, factor I, factor B, C5a and C5, whilst in nonobese controls, BMI only correlated negatively with C1q. Comparison of nonobese and obese PCOS showed that properdin, C3b, iC3b, C4A, factor D, factor H and MBL differed. The upregulation of the alternative complement pathway was seen in nonobese PCOS and was further exacerbated in obese PCOS, indicating that this is an inherent feature of the pathophysiology of PCOS that is worsened by obesity and is reflected in the differences between the nonobese and obese PCOS phenotypes. However, the increase in the complement proteins associated with activation was counterbalanced by upregulation of complement inhibitors; this was evident in both PCOS groups, suggesting that insults, such as a cardiovascular event or infection, that cause activation of complement pathways may be amplified in PCOS.

Ultrasensitive Electrochemiluminescence Immunosensor for Bladder Marker Human Complement Factor H-Related Protein Detection.

The development of noninvasive and sensitive detection methods for the early diagnosis and monitoring of bladder cancer is critical but challenging. Herein, an ultrasensitive electrochemiluminescence (ECL) immunosensor that uses Ru(bpy)32+-metal-organic framework (Ru-MOF) nanospheres and a DNA tetrahedral (TDN) probe was established for bladder cancer marker complement factor H-related protein (CFHR1) detection. The synthesized Ru(bpy)32+-metal-organic frameworks (Ru-MOFs) served as a linked substrate for immobilization of AuNPs and antibody (Ab2) to prepare the ECL signal probe (Ru-MOF@AuNPs-Ab2), exhibiting a stable and strengthened ECL emission. At the same time, the inherent advantages of TDN probes on the electrode as the capture probe (TDN-Ab1) improve the accessibility of targets to probes. In the presence of CFHR1, the signal probe Ru-MOF@AuNPs-Ab2 was modified on the electrode through immune binding, thereby obtaining an outstanding ECL signal. As expected, the developed ECL immunosensor exhibited splendid performance for CFHR1 detection in the range of 0.1 fg/mL to 10 pg/mL with a quite low detection limit of 0.069 fg/mL. By using the proposed strategy to detect CFHR1 from urine, it showed acceptable accuracy, which can effectively distinguish between bladder cancer patients and healthy samples. This work contributes to a novel, noninvasive, and accurate method for early clinical diagnosis of bladder cancer.

Several potential serum proteomic biomarkers for diagnosis of osteoarticular tuberculosis based on mass spectrometry

BackgroundOsteoarticular tuberculosis is one of the extrapulmonary tuberculosis (EPTB) diseases, which is mainly caused by infection of Mycobacterium tuberculosis (MTB) in bone and joints. The limitation of current clinical test methods is leading to a high misdiagnosis rate and affecting the treatment and prognosis. This study aims to search serum biomarkers that can assist in the diagnosis of osteoarticular tuberculosis. MethodsProteomics can serve as an important method in the discovery of disease biomarkers. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to analyze proteins in 90 serum samples, which were collected from June 2020 to December 2021, then evaluated by statistical analysis to screen potential biomarkers. After that, potential biomarkers were validated by enzyme-linked immunosorbent assay (ELISA) and diagnostic models were also established for observation of multi-index diagnostic efficacy. Results118 differential expressed proteins (DEPs) were obtained in serum after statistical analysis. After the diagnostic efficacy evaluation and clinical verification, inter-alpha-trypsin inhibitor heavy chain H2 (ITIH2), complement factor H-related protein 2 (CFHR2), complement factor H-related protein 3 (CFHR3), and complement factor H-related protein 5 (CFHR5) were found as potential biomarkers, with 0.7167 (95 %CI: 0.5846–0.8487), 0.8600 (95 %CI: 0.7701–0.9499), 0.8150 (95 %CI: 0.6998–0.9302), and 0.9978 (95 %CI: 0.9918–1.0040) AUC value, respectively. The remaining DEPs except CFHR5 were constructed as diagnostic models, the diagnostic model contained CFHR2 and CFHR3 had good diagnostic efficacy with 0.942 (95 %CI: 0.872–0.980) AUC value compared to other models. ConclusionThis study provides a reference for the discovery of serum protein markers for osteoarticular tuberculosis diagnosis, and the screened DEPs can also provide directions for subsequent pathogenesis research.

#6120 MIRNA PROFILING OF URINARY EXTRACELLULAR VESICLES (EVS) IN TBMN AND CFHR5 NEPHROPATHY FOR THE IDENTIFICATION OF NOVEL DISEASE BIOMARKERS

Abstract Background and Aims Thin basement membrane nephropathy (TBMN) and complement factor-H related protein 5 (CFHR5) nephropathy are two of the most common renal monogenic diseases with high phenotypic heterogeneity that is only partly accounted for by locus and allelic factors. Intra-familial variability in the age of ESKD onset for some patients particularly exemplifies such highly variable phenotypes, suggesting a potential modifying role of additional factors such as non-coding regulatory RNAs. Given that miRNAs are key regulators of gene expression, miRNAs from urine-derived extracellular vesicles (EVs) were investigated for their potential use as biomarkers in TBMN and CFHR5 diagnosis and prognosis. Method MiRNA profiling of human urinary EVs was carried out by small RNA-sequencing in an initial discovery cohort of 54 patients with TBMN and 44 patients with CFHR5 nephropathy, each classified further into three subgroups based on disease severity (Grey-zone: eGFR &amp;gt;60 ml/min/1.73m2 and age &amp;lt;50 y/o; Mild: eGFR &amp;gt;60 ml/min/1.73m2 and age &amp;lt;50 y/o; Severe: eGFR &amp;lt;60 ml/min/1.73m2 independently of age). Differential expression of miRNAs from urine-derived EVs was compared with 30 age and sex matched healthy controls, whilst bioinformatic analysis was performed to delineate their associated mechanisms. Results Kidney-enriched candidate miRNA families were identified in urine-derived EVs for both TBMN and CFHR5 patients and further verified by RT-qPCR in the respective validation cohorts. Distinct miRNAs with significantly differentiated expression levels, compared to that of healthy control cohort, showed significant correlations with some of the evaluated clinical characteristics for both TBMN and CFHR5 patients suggesting a possible diagnostic and prognostic value on long follow-up. Finally, inverse correlations of miRNA expression with their predicted targeted genes identified dysregulated pathways and transcriptional networks for further investigation. Conclusion Collectively, distinct profiles of miRNAs from urine-derived EVs were identified in TBMN and CFHR5 nephropathy suggesting that a subset of differentially expressed miRNAs could serve as novel non-invasive biomarkers of disease progression and/or targets for the development of novel therapeutic approaches.

#5990 COMPLEMENT GENES PROFILE OF PATIENTS WITH SEVERE HYPERTENSION AND RENAL THROMBOTIC MICROANGIOPATHY

Abstract Background and Aims Hereditary or acquired complement system dysregulation found to be driving cause of endothelial damage in atypical haemolytic uremic syndrome (aHUS) – ultrarare life-threatening disease, that always manifests with renal thrombotic microangiopathy (TMA), resulting in poor outcome in absence of appropriate treatment. Nevertheless, several studies show high incidence of rare genetic variants of complement system in other TMA’s, including hypertension-induced TMA. Here we report results of complement genes screening in patients (pts) with histologically identified renal TMA and severe hypertension in Russia, at first diagnosed as hypertension-induced TMA. Method A total of 28 Caucasian pts diagnosed with renal TMA (confirmed by kidney biopsy in all cases) and severe hypertension enrolled in the study. The main characteristic of the clinical picture was the absence of microangiopathic hemolysis and thrombocytopenia in prevailing of the pts, thus most of the primary and secondary causes of TMA clinically excluded. Examination of DNA samples was performed in all pts using next generation sequencing (NGS) technology. For statistical analysis continuous variables were presented as Mean (±Standart deviation) or Median (interquartile range) as appropriate. Results Baseline demographic and clinical characteristics of group are listed in Table 1. 13 pts (46%) had malignant form of hypertension (MHT) confirmed by eye examination. Mean systolic arterial pressure (SAP) observed at the time of presentation was 210±29 mmHg, mean diastolic arterial pressure (DAP) was 122±19 mmHg. Kidney morphology revealed chronic TMA lesions in 12 pts (43%), acute and chronic alterations simultaneously detected in 13 pts (46%). Predominantly acute lesions with glomerular and extraglomerular thrombi found in 3 (11%) pts only. Complement genes abnormalities revealed in 7 (25%) out of 28 pts. Complement factor I (CFI) rare variants were found in 2 pts (CFI c. 772&amp;lt;A, CFI c.1217G&amp;gt;A), one of this pts also had additional variant in complement factor H related protein 4 (CFHR4 c.766G&amp;gt;A). Complement factor C3 rare variant was observed in 1 pt (C3 c.2203C&amp;gt;T), membrane cofactor protein (MCP or CD46) rare variant in 1 pt (CD46 c. 198A&amp;gt;T), complement factor H related protein 5 (CFHR5) variant detected in 1 pt (CFHR5 c.136C&amp;gt;T), homozygous deletion of CFHR1 and CFHR3 detected in 1 pt and heterozygous deletion of the same complement factor H related protein genes found in another pt. Conclusion A quarter of pts with TMA and severe hypertension may have genetic defects in the complement system. These results confirmed that complement-mediated HUS may be misdiagnosed as hypertension-induced TMA. Lack of convincing hematological signs of TMA is common in this group, making aHUS diagnosis more sophisticated.

Comparative proteomic analysis of glomerular proteins in IgA nephropathy and IgA vasculitis with nephritis

BackgroundIgA nephropathy (IgAN) and IgA vasculitis with nephritis (IgAVN) are related glomerular diseases characterized by marked similarities in immunological and histological findings. We herein performed a comparative proteomic analysis of glomerular proteins in IgAN and IgAVN.MethodsWe used renal biopsy specimens from 6 IgAN patients without nephrotic syndrome (NS) (IgAN-I subgroup), 6 IgAN patients with NS (IgAN-II subgroup), 6 IgAVN patients with 0–8.0% of glomeruli with crescent formation (IgAVN-I subgroup), 6 IgAVN patients with 21.2–44.8% of glomeruli with crescent formation (IgAVN-II subgroup), 9 IgAVN patients without NS (IgAVN-III subgroup), 3 IgAVN patients with NS (IgAN-IV subgroup), and 5 control cases. Proteins were extracted from laser microdissected glomeruli and analyzed using mass spectrometry. The relative abundance of proteins was compared between groups. An immunohistochemical validation study was also performed.ResultsMore than 850 proteins with high confidence were identified. A principal component analysis revealed a clear separation between IgAN and IgAVN patients and control cases. In further analyses, 546 proteins that were matched with ≥ 2 peptides were selected. The levels of immunoglobulins (IgA, IgG, and IgM), complements (C3, C4A, C5, and C9), complement factor H-related proteins (CFHR) 1 and 5, vitronectin, fibrinogen chains, and transforming growth factor-β inducible gene-h3 were higher (> 2.6 fold) in the IgAN and IgAVN subgroups than in the control group, whereas hornerin levels were lower (< 0.3 fold). Furthermore, C9 and CFHR1 levels were significantly higher in the IgAN group than in the IgAVN group. The abundance of some podocyte-associated proteins and glomerular basement membrane (GBM) proteins was significantly less in the IgAN-II subgroup than in the IgAN-I subgroup as well as in the IgAVN-IV subgroup than in the IgAVN-III subgroup. Among the IgAN and IgAVN subgroups, talin 1 was not detected in the IgAN-II subgroup. This result was supported by immunohistochemical findings.ConclusionsThe present results suggest shared molecular mechanisms for glomerular injury in IgAN and IgAVN, except for enhanced glomerular complement activation in IgAN. Differences in the protein abundance of podocyte-associated and GBM proteins between IgAN and IgAVN patients with and without NS may be associated with the severity of proteinuria.

Characteristics and management of thrombotic microangiopathy in kidney transplantation.

Thrombotic microangiopathy is not a rare complication of kidney transplantation and is characterized by microangiopathic hemolytic anemia, thrombocytopenia, and acute kidney injury with extensive thrombosis of the arterioles and capillaries. Various factors can cause thrombotic microangiopathy after kidney transplantation, including surgery, warm and cold ischemia-reperfusion injury, exposure to immunosuppressants, infection, and rejection. Many recent studies on atypical hemolytic uremic syndrome have described genetic abnormalities related to excessive activation of the alternative complement pathway. The affected patients' genetic backgrounds revealed significant genetic heterogeneity in several genes involved in complement regulation, including the complement factor H, complement factor H-related proteins, complement factor I, complement factor B, complement component 3, and CD46 genes in the alternative complement pathway. Although clinical studies have provided a better understanding of the pathogenesis of diseases, the diverse triggers present in the transplant environment can lead to thrombotic microangiopathy, along with various genetic predispositions, and it is difficult to identify the genetic background in various clinical conditions. Given the poor prognosis of posttransplant thrombotic microangiopathy, further research is necessary to improve the diagnosis and treatment protocols based on risk factors or genetic predisposition, and to develop new therapeutic agents.

Protein-centric omics analysis reveals circulating complements linked to non-viral liver diseases as potential therapeutic targets.

To evaluate the causal correlation between complement components and non-viral liver diseases and their potential use as druggable targets. We conducted Mendelian randomization (MR) to assess the causal role of circulating complements in the risk of non-viral liver diseases. A complement-centric protein interaction network was constructed to explore biological functions and identify potential therapeutic options. In the MR analysis, genetically predicted levels of complement C1q C chain (C1QC) were positively associated with the risk of autoimmune hepatitis (odds ratio 1.125, 95% confidence interval 1.018-1.244), while complement factor H-related protein 5 (CFHR5) was positively associated with the risk of primary sclerosing cholangitis (PSC;1.193, 1.048- 1.357). On the other hand, CFHR1 (0.621, 0.497-0.776) and CFHR2 (0.824, 0.703-0.965) were inversely associated with the risk of alcohol-related cirrhosis. There were also significant inverse associations between C8 gamma chain (C8G) and PSC (0.832, 0.707-0.979), as well as the risk of metabolic dysfunction-associated steatotic liver disease (1.167, 1.036-1.314). Additionally, C1S (0.111, 0.018-0.672), C7 (1.631, 1.190-2.236), and CFHR2 (1.279, 1.059-1.546) were significantly associated with the risk of hepatocellular carcinoma. Proteins from the complement regulatory networks and various liver diseaserelated proteins share common biological processes. Furthermore, potential therapeutic drugs for various liver diseases were identified through drug repurposing based on the complement regulatory network. Our study suggests that certain complement components, including C1S, C1QC, CFHR1, CFHR2, CFHR5, C7, and C8G, might play a role in non-viral liver diseases and could be potential targets for drug development.