The transcription factor HMGB2 indirectly regulates APRIL expression and Gd-IgA1 production in patients with IgA nephropathy.

BACKGROUND: IgA nephropathy (IgAN) is the most common primary glomerulonephritis worldwide. Proliferation-inducing ligand (APRIL) was identified as an important cause of glycosylation deficiency of IgA1 (Gd-IgA1), which can 'trigger' IgAN. Our previous study indicated that high migration group protein B2 (HMGB2) in peripheral blood mononuclear cells from patients with IgAN was associated with disease severity, but the underlying mechanism remains unclear. MATERIALS AND METHODS: The location of HMGB2 was identified by immunofluorescence. qRT-PCR and Western blotting were used to measure HMGB2, HMGA1, and APRIL expression. Gd-IgA1 levels were detected by enzyme-linked immunosorbent assay (ELISA). In addition, we used DNA pull-down, protein profiling, and transcription factor prediction software to identify proteins bound to the promoter region of the APRIL gene. RNA interference and coimmunoprecipitation (Co-IP) were used to verify the relationships among HMGB2, high mobility group AT-hook protein 1 (HMGA1), and APRIL. RESULTS: HMGB2 expression was greater in IgAN patients than in HCs and was positively associated with APRIL expression in B cells. DNA pull-down and protein profiling revealed that HMGB2 and HMGA1 bound to the promoter region of the APRIL gene. The expression levels of HMGA1, APRIL, and Gd-IgA1 were downregulated after HMGB2 knockdown. Co-IP indicated that HMGB2 binds to HMGA1. The Gd-IgA1 concentration in the supernatant was reduced after HMGA1 knockdown. HMGA1 binding sites were predicted in the promoter region of the APRIL gene. CONCLUSION: HMGB2 expression is greater in IgAN patients than in healthy controls; it promotes APRIL expression by interacting with HMGA1, thereby inducing Gd-IgA1 overexpression and leading to IgAN.

Tacrolimus treatment after short-term intravenous methylprednisolone in incipient minimal change disease for adults: A retrospective analysis.

The present study aimed to investigate the efficacy and safety of tacrolimus for treating incipient minimal change disease in adults. The clinical data of 52 adult patients with minimal change disease of nephrotic syndrome diagnosed by renal biopsy in the First affiliated hospital of Zhengzhou University between August 2013 and August 2015 were retrospectively analyzed. According to the treatment plan, the patients were divided into a tacrolimus group and a glucocorticoid group. The efficacy and safety of tacrolimus in the treatment of minimal change disease in adult patients was analyzed and compared with that of glucocorticoids. The results revealed that the baseline characteristics of the two groups were similar (P>0.05). At 24 weeks, there was a significant difference in serum albumin between the two groups (P<0.01). The serum albumin levels of tacrolimus group was higher compared with the glucocorticoid group. In addition, the complete remission rates in the tacrolimus and glucocorticoid groups were 93.75 and 77.8%, respectively (P=0.095), and the mean complete remission time was 6.33±4.21 and 5.14±2.45 weeks, respectively (P=0.175). The relapse rate was 12.5 and 22.2% in the tacrolimus and glucocorticoid groups, respectively (P=0.368). During the follow-up, in tacrolimus group, the incidence of new onset diabetes or impaired glucose tolerance, osteoporosis, infection, abnormal liver function, Cushing's syndrome, acne and gastrointestinal symptoms were significantly less than those of glucocorticoids (P<0.05). In conclusion, tacrolimus treatment after short-time intravenous methylprednisolone is an effective treatment option with fewer adverse effects in adult onset minimal change disease.

The comprehensive analysis of clinical trials registration for IgA nephropathy therapy on ClinicalTrials.gov.

OBJECTIVES: IgA Nephropathy (IgAN) is common chronic kidney disease with a high incidence. This study aims to analyze comprehensively therapeutic clinical trials for IgAN registered on ClinicalTrials.gov. METHODS: Therapeutic trials for IgAN registered on ClinicalTrials.gov. up to 15 August 2021 were obtained. The general characteristics, features of experimental design, treatment strategies, and some main inclusion criteria and outcome measures were accessed. RESULTS: A total of 104 therapeutic clinical trials for IgAN were extracted on ClinicalTrials.gov up to 15 August 2021. Most of these trials explored the treatment for primary IgAN confirmed by renal biopsy in adults. Only 9% of all selected trials had results. Forty-five percent of trials recruited 50 or fewer participants, and 73% were adults or older adults. 99% of trials were interventional studies, and of all the interventional trials, 70% of trials were randomized, and 68% exercised a parallel assignment of intervention model. Immunosuppression was the most studied for the treatment of IgAN. Moreover, many novel agents had been increasingly studied in recent years. Furthermore, the inclusion criteria and primary outcome measures in these trials were diverse, and the level of proteinuria and change of proteinuria levels were the most used as inclusion criteria and primary outcome, respectively. CONCLUSIONS: The majority of therapeutic trials for IgAN were randomized, none masking and parallel-assignment interventional studies, primarily recruiting adult patients as research subjects. These trials had relatively small sample sizes and short observation. Thus, more large-scale, multicenter, and randomized controlled trials are still needed to improve the management for IgAN.

Lupus susceptibility region containing CTLA4 rs17268364 functionally reduces CTLA4 expression by binding EWSR1 and correlates IFN-α signature.

BACKGROUND: Dysregulation of T cells mediated immune responses is a hallmark in the development of systemic lupus erythematosus (SLE). Recent genome wide association study (GWAS) revealed the genetic contribution of variants located in the cytotoxic T lymphocyte-associated protein-4 (CTLA4)-inducible T cell co-stimulator (ICOS) intergenic region to SLE susceptibility. Our aim is to find a functional variant in this region. METHODS: The genetic association results in the CTLA4-ICOS region from previous GWAS were adopted to select the potential variant which was further replicated in two independent cohorts (Henan cohort 2053 SLE patients and 1845 healthy controls, Beijing cohort 2303 SLE patients and 19,262 healthy). In order to explore the functional significance in SLE, bioinformatics with validation experiments (including electrophoretic mobility shift assay and luciferase reporter assay) and mRNA expression analysis were also performed. RESULTS: A variant located in the CTLA4-ICOS intergenic region, rs17268364, was associated with susceptibility to SLE patients in Chinese populations (risk allele, pmeta = 7.02×10-11, OR 1.19, 95%CI 1.13-1.26). The bioinformatics suggested that rs17268364 might affect the expression of CTLA4, not ICOS. The rs17268364 risk G allele containing sequence reduced the expression of the reporter gene by binding transcriptional repressor Ewing sarcoma breakpoint region 1 (EWSR1). Following genotype-mRNA expression, the analysis also showed the risk allele of rs17268364 was associated with low CTLA4 expression in lupus nephritis (LN) patients. Healthy individuals carrying rs17268364 risk G allele was significantly correlated with higher levels of IFN-α signature including increased lymphocyte antigen 6E (LY6E) (p=0.031), interferon-stimulated gene 15 (ISG15) (p=0.038), interferon regulatory factor 9 (IRF9) (p=0.028), and interferon regulatory factor 5 (IRF5) (p=0.040) mRNA expression. CONCLUSIONS: The present study confirmed the functional role of rs17268364 in the CTLA4-ICOS intergenic region that increased SLE susceptibility in the Chinese population.

MicroRNAs in IgA nephropathy.

IgA nephropathy (IgAN) is the most common primary glomerulonephritis worldwide. It is considered that the pathogenesis of IgAN involves the 'multiple hit theory' and the immune-inflammatory mechanism; however, these theories have certain limitations. The gold standard for diagnosing IgAN is still renal biopsy. Although renal biopsy is accurate, it is traumatic and is associated with some risks and limitations, so there is a need for non-invasive diagnostic methods. According to recent studies, microRNAs (miRNAs) play important roles in the occurrence and development of IgAN; thus, they provide the possibility of the noninvasive diagnosis of IgAN and also have some value in predicting prognosis. This review summarizes the current research status of miRNAs in the occurrence, development, diagnosis, and prognosis of IgAN. We also highlight some interesting and challenging points that require further study.

Association of the PINX1 Variant rs6984094, Which Lengthens Telomeres, with Systemic Lupus Erythematosus Susceptibility in Chinese Populations.

A recent genome-wide association study (GWAS) of Asian ancestry reported that single nucleotide polymorphism (SNP) in TERT (telomerase reverse transcriptase) was associated with systemic lupus erythematosus (SLE). TERT has a critical role in maintaining the chromosomal stability and the length of telomere. Given that only a small portion of the genetic heritability of SLE has been explained so far, we aimed to identify novel loci in telomere-related genes responsible for SLE susceptibility in Chinese populations. We performed a comprehensive genetic association analysis of SLE with telomere-related genes. To identify functional significance, we analyzed the publicly available HaploReg v4.1 and RegulomeDB databases. Differential gene expression analysis was also performed using ArrayExpress. A novel signal of PINX1 rs6984094 was identified (P discovery = 4.13 × 10-2, OR = 0.58, 95% CI 0.35-0.98) and successfully replicated (P replication = 5.73 × 10-3, OR = 0.45, 95% CI 0.26-0.81). Multiple layers of functional analysis suggested that the PINX1 rs6984094 risk T allele exhibited increased nuclear protein binding. We also observed an increased expression of PINX1 mRNA in peripheral blood mononuclear cells from SLE patients compared with healthy controls. Overall, we observed a novel genetic association between PINX1 (encodes the PinX1 protein, an inhibitory telomerase enzyme that lengthens telomeres) and SLE susceptibility in Chinese populations.

A botanical medicine dragon's blood exhibited clinical antithrombosis efficacy similar to low molecular weight heparin.

Deep vein thrombosis (DVT) is a common complication following traumatic fracture with a 0.5%-1% annual incidence. Low molecular weight heparin (LMWH) is the most commonly used anticoagulation drug for DVT prevention, but treatment with LMWH is invasive. Our aim is to compare the antithrombotic effect of dragon's blood, an oral botanical anticoagulant medicine approved by the Chinese FDA, with LMWH in patients undergoing hip fracture surgery and to explore the molecular mechanisms of anticoagulation treatment. Our study recruited patients and divided them into LMWH and dragon's blood treatment group. Coagulation index tests, Doppler ultrasound and mRNA sequencing were performed before and after anticoagulation therapy. There was no significant difference in postoperative DVT incidence between the two groups (23.1% versus 15.4%, P=0.694). D-dimer (D-D) and fibrinogen degradation product (FDP) showed significant reductions in both groups after anticoagulation treatments. We identified SLC4A1, PROS1, PRKAR2B and seven other genes as being differentially expressed during anticoagulation therapy in both groups. Genes correlated with coagulation indexes were also identified. Dragon's blood and LMWH showed similar effects on DVT and produced similar gene expression changes in patients undergoing hip fracture surgery, indicating that dragon's blood is a more convenient antithrombosis medicine (oral) than LMWH (hypodermic injection).

Elevated Endostatin Expression Is Regulated by the pIgA Immune Complex and Associated with Disease Severity of IgA Nephropathy.

BACKGROUND/AIMS: Renal vascular injury accounts for the poor outcomes of patients with IgA nephropathy (IgAN). In this study, we investigated whether endostatin, a potent inhibitor of angiogenesis, is associated with IgAN. METHODS: Serum endostatin levels were detected in patients with IgAN, disease controls, and healthy controls, and the correlation among endostatin and clinicopathologic manifestations, as well as prognosis in patients with IgAN, was analyzed. In addition, serum endostatin levels were compared in patients "before" and "after" treatment. Data on endostatin expression in the renal interstitium of patients with IgAN were downloaded and analyzed from the GSE35489 array in the GEO database. The poly-IgA1 (pIgA) immune complex is widely recognized as the "trigger" of IgAN initiation. pIgA in the plasma of patients was extracted and used to stimulate human glomerular endothelial cells (GECs). Endostatin, IL-6, and CXCL1 in the cell supernatant were detected by ELISA kits. RESULTS: We found that serum endostatin levels were significantly increased in patients with IgAN, as was endostatin expression in the renal interstitium. Patients with IgAN were divided into 2 groups according to the median value. The high endostatin expression group had significantly higher levels of serum creatinine and BUN and more severe tubular/interstitial damage. Moreover, patients with arteriolar injury and endothelial cell proliferation had higher serum endostatin levels. Patients with high serum endostatin levels had poor prognosis. According to the in vitro experiment, the GEC apoptosis rate and the supernatant levels of endostatin, IL-6, and CXCL1 were significantly increased following pIgA stimulation. CONCLUSION: Our study found that elevated endostatin expression was associated with disease severity and poor prognosis in patients with IgAN and can be upregulated by pIgA, but how it participates in the pathogenesis of IgAN deserves further exploration.

Elevated Serum Chloride Levels Contribute to a Poor Prognosis in Patients with IgA Nephropathy.

INTRODUCTION: The identification of reliable prognostic factors is a crucial requirement for patients with IgA nephropathy (IgAN). Here, we explored the relationship between serum chloride levels and prognosis in patients with IgAN. METHODS: We recruited all patients with primary IgAN, as diagnosed by renal biopsy, between 1st January 2015 and 1st April 2019. Patients were divided two groups (high chloride group and low chloride group) based on the best cut-off values from survival receiver operating characteristic (ROC) curves. The baseline clinicopathological characteristics of two groups were then compared. Cox proportional hazard models were used to determine the prognostic value of serum chloride levels in patients with IgAN. Finally, we screened reliable prognostic indicators and built a clinical prediction model and validated the performance of the model. RESULTS: Compared with patients in the high chloride group, patients in the low chloride group had significantly lower levels of 24-hour urinary total protein (24 h-UTP), serum creatinine (sCr), and higher levels of hemoglobin (Hb), albumin (all p < 0.05), and less proportion of Oxford classification grade E1 (endothelial cell proliferation) and T2 (renal tubule atrophy or renal interstitial fibrosis). Cox analysis revealed that serum chloride level ≥ 105.4 mmol/L was a significant and independent risk factor for prognosis in patients with IgAN (p < 0.05). Serum chloride, sCr, T, hypertension, and Hb were used to generate a predictive model for prognosis. Thec-indices of our predictive model were 0.80, 0.86, and 0.78, for 1, 2, and 3 years, respectively; Brier scores were 0.06, 0.09, and 0.16, respectively. CONCLUSIONS: A serum chloride level ≥ 105.4 mmol/l was identified as a significant and independent risk factor for the prognosis of patients with IgAN. A predictive prognosis model was generated using serum chloride, sCr, T, hypertension, and Hb; this model exhibited a good predictive effect.

The soluble VEGF receptor sFlt-1 contributes to endothelial dysfunction in IgA nephropathy.

Endothelial injury is a common manifestation in IgA nephropathy (IgAN). After the previous identification of the upregulated soluble fms-like tyrosine kinase-1 (sFlt-1) correlated with endothelial injury in IgAN, in the present study, we further explored the role of sFlt-1 in endothelial injury in IgAN. We enrolled 72 patients with IgAN and detected the sFlt-1 levels. The polymeric IgA1 (pIgA1) complexes were isolated from the pooled plasma samples of another 10 patients with IgAN. Apoptosis proteins were detected in cultured human umbilical vein endothelial cells (HUVECs) with the stimulation of recombinant sFlt-1 or the caspase-9 inhibitor Z-LEHD-FMK. We identified there were positive correlations between sFlt-1 and IgA-IgG complex as well as vWF levels in patients with IgAN. The sFlt-1 levels in HUVECs were significantly upregulated by pIgA1 complex derived from IgAN patients in a concentration-dependent manner. The proliferation ability of HUVECs was damaged when stimulated with sFlt-1 protein in a time- and dose- dependent manner. And the apoptosis rate was up-regulated significantly as the stimulation concentrations of sFlt-1 increased. We found sFlt-1 challenge could significantly increase the expression of vWF. In addition, sFlt-1 increased the levels of caspase-9, caspase-3, Bax and mitochondrial membrane potential; facilitated the release of cytochrome C from mitochondria to cytoplasma. In contrast, Z-LEHD-FMK attenuated high sFlt-1-induced HUVECs apoptosis. In conclusion, our study demonstrated that sFlt-1 expression was up-regulated by the challenge of pIgA1 complex derived from patients with IgAN. Furthermore, increased sFlt-1 facilitated human umbilical vein endothelial cells apoptosis via the mitochondrial-dependent pathway.

Increased APRIL Expression Induces IgA1 Aberrant Glycosylation in IgA Nephropathy.

Aberrant glycosylated IgA1 molecules, mainly galactose-deficient IgA1 (Gd-IgA1), are important causal factors in IgA nephropathy; however, the underlying mechanism for the production of aberrantly glycosylated IgA1 is unknown. A recent genome-wide association study identified a novel IgAN susceptibility gene, TNFSF13, which encoded a proliferation-inducing ligand (APRIL) that promotes lymphocyte proliferation and IgA class switching. We aimed to explore the mechanism of APRIL's involvement in IgAN. We enrolled 166 patients with IgAN and 77 healthy controls and detected the plasma APRIL levels by the ELISA method, identified the mRNA expression of APRIL and its receptors by relative quantitative PCR, and confirmed by in vitro experiment. We identified increased plasma APRIL levels in IgAN, which was further proved by upregulated mRNA expression in B-lymphocytes from 27 IgAN patients. Analysis of the clinical characteristics of patients with IgAN showed that higher plasma APRIL level was associated with more severe clinical presentations (high proteinuria and low eGFR). The plasma APRIL level was positively correlated with Gd-IgA1 levels. Furthermore, exogenous APRIL could induce more production of Gd-IgA1 in cultured lymphocytes from patients with IgAN, compared with that from healthy controls. And, the relative higher expression of receptors of APRIL, that is, BCMA and TACI, in B-lymphocytes from IgAN patients were observed. Our findings implied that in patients with IgAN, increased APRIL is accompanied elevated expression of its receptors in B-lymphocytes, which induces overproduction of Gd-IgA1, ultimately contributing to the pathogenesis of IgAN.

Elevated soluble VEGF receptor sFlt-1 correlates with endothelial injury in IgA nephropathy.

BACKGROUND: Endothelial injury, which may present clinically as hypertension, proteinuria and increased von Willebrand Factor (vWF) level, is a common manifestation in IgA nephropathy (IgAN). However, causal factors for endothelial injury in IgAN are not completely understood. An imbalance of vascular endothelial growth factor/Soluble fms-like tyrosine kinase-1 (VEGF/sFlt-1) has been observed in many diseases with endothelial dysfunction, including pre-eclampsia and diabetic retinopathy, but whether it contributes to endothelial injury in IgAN requires further exploration. METHODS: Initially, 96 IgAN patients and 22 healthy volunteers were enrolled as a discovery cohort. VEGF/sFlt-1, sFlt-1 and VEGF levels were compared between patients with IgAN and healthy volunteers to explore the underlying factors that contribute to endothelial injury in IgAN. The identified contributor (sFlt-1) was further confirmed in a replication cohort, which included 109 IgAN patients and 30 healthy volunteers. Correlations of sFlt-1 with hypertension, proteinuria, Oxford-E score and plasma vWF were further evaluated in the combined 205 patients with IgAN. RESULTS: VEGF/sFlt-1 levels were significantly lower in IgAN patients than healthy volunteers (0.33±0.27 vs. 0.43±0.22, p = 0.02) in the discovery cohort. Within the ratio, plasma sFlt-1 levels were significantly elevated (101.18±25.19 vs. 79.73±18.85 pg/ml, p<0.001), but plasma VEGF levels showed no significant differences. Elevated sFlt-1 levels in the replication cohort were confirmed in IgAN patients (93.40±39.78 vs. 71.92±15.78 pg/ml, p<0.001). Plasma sFlt-1 levels in IgAN patients correlated with proteinuria (severe (>3.5 g/d) vs. moderate (1-3.5 g/d) vs. mild (<1 g/d) proteinuria: 115.95±39.09 vs. 99.89±28.55 vs. 83.24±33.92 pg/ml; severe vs. mild: p<0.001, moderate vs. mild p = 0.001, severe vs. moderate: p = 0.014), hypertension (with vs. without hypertension: 107.87±31.94 vs. 87.32±32.76 pg/ml, p = 0.015) and vWF levels (r = 0.161, p = 0.021). CONCLUSIONS: The present study found elevated sFlt-1 in IgAN patients and further identified its correlation with proteinuria, hypertension and vWF levels. These results suggested that elevated sFlt-1 contributes to endothelial injury in IgAN.