Genome-Wide Association Study in Acute Tubulointerstitial Nephritis.

SIGNIFICANCE STATEMENT: Polymorphisms of HLA genes may confer susceptibility to acute tubulointerstitial nephritis (ATIN), but small sample sizes and candidate gene design have hindered their investigation. The first genome-wide association study of ATIN identified two significant loci, risk haplotype DRB1*14-DQA1*0101-DQB1*0503 (DR14 serotype) and protective haplotype DRB1*1501-DQA1*0102-DQB1*0602 (DR15 serotype), with amino acid position 60 in the peptide-binding groove P10 of HLA-DR ß 1 key. Risk alleles were shared among different causes of ATIN and HLA genotypes associated with kidney injury and immune therapy response. HLA alleles showed the strongest association. The findings suggest that a genetically conferred risk of immune dysregulation is part of the pathogenesis of ATIN. BACKGROUND: Acute tubulointerstitial nephritis (ATIN) is a rare immune-related disease, accounting for approximately 10% of patients with unexplained AKI. Previous elucidation of the relationship between genetic factors that contribute to its pathogenesis was hampered because of small sample sizes and candidate gene design. METHODS: We undertook the first two-stage genome-wide association study and meta-analysis involving 544 kidney biopsy-defined patients with ATIN and 2346 controls of Chinese ancestry. We conducted statistical fine-mapping analysis, provided functional annotations of significant variants, estimated single nucleotide polymorphism (SNP)-based heritability, and checked genotype and subphenotype correlations. RESULTS: Two genome-wide significant loci, rs35087390 of HLA-DQA1 ( P =3.01×10 -39 ) on 6p21.32 and rs2417771 of PLEKHA5 on 12p12.3 ( P =2.14×10 -8 ), emerged from the analysis. HLA imputation using two reference panels suggested that HLA-DRB1*14 mainly drives the HLA risk association . HLA-DRB1 residue 60 belonging to pocket P10 was the key amino acid position. The SNP-based heritability estimates with and without the HLA locus were 20.43% and 10.35%, respectively. Different clinical subphenotypes (drug-related or tubulointerstitial nephritis and uveitis syndrome) seemed to share the same risk alleles. However, the HLA risk genotype was associated with disease severity and response rate to immunosuppressive therapy. CONCLUSIONS: We identified two candidate genome regions associated with susceptibility to ATIN. The findings suggest that a genetically conferred risk of immune dysregulation is involved in the pathogenesis of ATIN.

Serum anti-CRP antibodies differentiate etiology and predict relapse in acute tubulointerstitial nephritis.

INTRODUCTION: Acute tubulointerstitial nephritis (ATIN) is a common cause of acute kidney injury with various etiologies. It has been shown that autoimmune-related ATIN (AI-ATIN) has a higher recurrence rate and a greater likelihood of developing into chronic kidney disease compared with drug-induced ATIN, yet misdiagnosis at renal biopsy is not uncommon. METHODS: Patients who were clinicopathologically diagnosed as ATIN from January 2006 to December 2015 in Peking University First Hospital were enrolled. Clinical, pathological and follow-up data were collected. Serum samples on the day of renal biopsy were collected and tested for anti-C-reactive protein (CRP) antibodies. CRP and its linear peptides were used as coating antigens to detect antibodies. Statistical analysis was used to assess the diagnostic value of the antibodies. RESULTS: Altogether 146 patients were enrolled. The receiver operating characteristic-area under the curve of the anti-CRP antibody for the identification of late-onset AI-ATIN was 0.750 (95% confidence interval 0.641-0.860, P < 0.001) and the positivity was associated with ATIN relapse (adjusted hazard ratio = 4.321, 95% confidence interval 2.402-7.775, P < 0.001). Antibodies detected by CRP linear peptide 6 (PT6) were superior with regard to differentiating patients with AI-ATIN, while antibodies detected by peptide 17 (PT17) could predict ATIN relapse. Antibodies detected by these two peptides were positively correlated with the severity of tubular dysfunction and pathological injury. CONCLUSIONS: Serum anti-CRP antibody could be used to differentiate late-onset AI-ATIN and predict relapse of ATIN at the time of renal biopsy. The CRP linear peptides PT6 and PT17 could be used as coating antigens to detect anti-CRP antibodies, which may provide more information for the clinical assessment of ATIN.

Influence of birth weight and infancy growth patterns on weight status among first grade primary school pupils / 中国学校卫生

Objective@#To explore the influence of birth weight and growth patterns during infancy on overweight and obesity among first grade primary school pupils, so as to provide a theoretical basis for the formulation of early life prevention and intervention policies.@*Methods@#In 2019, data related to routine physical examinations were collected for primary school pupils in the Minhang District of Shanghai, and information regarding birth and follow ups was collected retrospectively. Physical examination data of 4 434 pupils at 12 months of age were obtained. A multiple linear regression model was used to analyze the relationship between growth patterns during infancy and body mass index (BMI) in the first grade of primary school. A generalized linear model was employed to analyze the relationship between birth weight and growth patterns during infancy and overweight and obesity in the first grade of primary school. A hierarchical analysis was conducted.@*Results@#A linear relationship was observed between growth patterns during infancy and BMI and the BMI Z score of first grade primary school pupils [ β(β 95%CI)=0.30(0.24-0.35),0.12(0.10- 0.15 ), P <0.01]. In addition to subjects classified as small for gestational age (SGA), catch up growth during infancy was identified among subjects who were classified as appropriate for gestational age (AGA) and large for gestational age (LGA). LGA at birth and catch up growth during infancy were independent risk factors for overweight and obesity among first grade primary school children ( RR =1.31-1.55, P <0.05). The hierarchical analysis showed that catch up growth increased the risk of overweight and obesity among first grade primary school pupils classified as AGA [ RR(RR 95%CI )=1.74(1.42-2.14),1.87(1.56-2.26)], and increased the risk of obesity among first grade primary school pupils classified as SGA and LGA [ RR(RR 95%CI )=3.74(1.04-13.49),3.24(1.62-6.46)]( P <0.05). Among those who exhibited catch up growth during infancy, LGA increased the risk of obesity among first grade primary school pupils ( RR= 2.60 , 95%CI=1.35-5.02, P <0.01), but not the risk of being overweight ( P =0.13).@*Conclusion@#Birth weight and growth patterns during infancy have an impact on overweight and obesity among children in the first grade of primary school. It is suggested that attention should be paid to growth and physical development in early life for those classified as LGA and AGA, and catch up growth in children should be closely monitored.

miR-200a-3p Attenuates Coronary Microembolization-Induced Myocardial Injury in Rats by Inhibiting TXNIP/NLRP3-Mediated Cardiomyocyte Pyroptosis.

Coronary microembolization (CME) commonly develops as a complication after percutaneous coronary intervention (PCI), and associated inflammation is a leading driver of myocardial damage. Cardiomyocyte loss in the context of ischemic myocardial disease has been linked to inflammatory pyroptotic cell death. Additionally, miR-200a-3p dysregulation has been linked to myocardial ischemia-reperfusion and many other pathological conditions. However, how miR-200a-3p impacts cardiomyocyte pyroptosis in the context of CME remains to be assessed. Herein, a rat model of CME was established via the injection of microembolic spheres into the left ventricle. When myocardial tissue samples from these rats were analyzed, miR-200a-3p levels were markedly decreased, whereas thioredoxin-interacting protein (TXNIP) levels were increased. The ability of miR-200a-3p to directly target TXNIP and to control its expression was confirmed via dual-luciferase reporter assay. Adeno-associated virus serotype 9-pre-miR-200a-3p (AAV-miR-200a-3p) construct transfection was then employed as a means of upregulating this miRNA in CME model rats. Subsequent assays, including echocardiography, enzyme-linked immunosorbent assays (ELISAs), hematoxylin-eosin (H&E) staining, hematoxylin-basic fuchsin-picric acid (HBFP) staining, TdT-mediated dUTP nick-end labeling (TUNEL) staining, immunofluorescence staining, quantitative real-time polymerase chain reaction (qRT-PCR), and Western blotting revealed that miR-200a-3p overexpression inhibited cardiomyocyte pyroptosis and alleviated CME-induced myocardial injury by inhibiting the TXNIP/NOD-like receptor family pyrin domain-containing 3 (NLRP3) pathway. The ability of miR-200a-3p to protect against CME-induced myocardial injury thus highlights a novel approach to preventing or treating such myocardial damage in clinical settings.

Downregulation of miR-181a-5p alleviates oxidative stress and inflammation in coronary microembolization-induced myocardial damage by directly targeting XIAP.

BACKGROUND: Coronary microembolization (CME) is a complicated problem that commonly arises in the context of coronary angioplasty. MicroRNAs play crucial roles in cardiovascular diseases. However, the role and mechanism of miR-181a-5p in CME-induced myocardial injury remains unclear. METHODS: We established CME rat models. Cardiac function was detected by echocardiography. Haematoxylin-basic fuchsin-picric acid staining was used to measure micro-infarction size. Serum samples and cell culture supernatants were evaluated via enzyme-linked immunosorbent assay. Cellular reactive oxygen species were determined by dichloro-dihydro-fluorescein diacetate assay, and the other oxidative stress related parameters were assayed by spectrophotometry. The dual-luciferase reporter (DLR) assay and RNA pulldown were conducted to validate the association between miR-181a-5p and X-linked inhibitor of apoptosis protein (XIAP). The expression of miR-181a-5p and XIAP mRNA were determined by quantitative reverse transcription polymerase chain reaction. Proteins were evaluated via immunoblotting. The viability of the cell was evaluated via cell counting kit-8 assay. RESULTS: The miR-181a-5p level was significantly increased in CME myocardial tissues. Downregulation of miR-181a-5p improved CME-induced cardiac dysfunction and alleviated myocardial oxidative stress and inflammatory injury, whereas miR-181a-5p exhibited the opposite effects. Then, the DLR assay and RNA pulldown results revealed that miR-181a-5p directly targeting on XIAP. The XIAP level was found to be remarkably decreased after CME. XIAP overexpression attenuated CME-induced myocardial oxidative stress and inflammatory injury. Finally, in vitro rescue experiments revealed that knockdown of XIAP could abolish the protective effects of miR-181a-5p knockdown on hypoxia-induced cardiomyocyte oxidative stress and inflammatory injury. CONCLUSIONS: Downregulation of miR-181a-5p alleviates CME-induced myocardial damage by suppressing myocardial oxidative stress and inflammation through directly targeting XIAP.

Breviscapine Pretreatment Inhibits Myocardial Inflammation and Apoptosis in Rats After Coronary Microembolization by Activating the PI3K/Akt/GSK-3ß Signaling Pathway.

PURPOSE: Coronary microembolization (CME) can cause myocardial inflammation, apoptosis and progressive cardiac dysfunction. On the other hand, breviscapine exerts a significant cardioprotective effect in many cardiac diseases although its role and the potential mechanisms in CME remain unclear. Therefore, the present study aimed to ascertain whether pretreatment with breviscapine could improve CME-induced myocardial injury by alleviating myocardial inflammation and apoptosis. The possible underlying mechanisms were also explored. METHODS: In this study, 48 Sprague-Dawley (SD) rats were randomly assigned to the CME, CME + breviscapine (CME + BE), CME + breviscapine + LY294002 (CME + BE + LY) and sham groups (12 rats per group). In addition, the CME model was successfully established by injecting 42 µm inert plastic microspheres into the left ventricle of rats. Rats in the CME + BE and CME + BE + LY groups received 40 mg/kg/d of breviscapine for 7 days before inducing CME. Moreover, rats in the CME + BE + LY group were intraperitoneally injected with the phosphoinositide 3-kinase (PI3K) specific inhibitor, LY294002 (10 mg/kg) 30 minutes before CME modeling. 12 h after surgery, the study measured cardiac function, the serum levels of markers of myocardial injury, myocardial inflammation-associated mRNAs and proteins, myocardial apoptosis-associated mRNAs and proteins and conducted myocardial histopathology. RESULTS: The findings demonstrated that pretreatment with breviscapine alleviated myocardial injury following CME by improving cardiac dysfunction, decreasing the serum levels of markers of myocardial injury, reducing the size of myocardial microinfarct and lowering the cardiomyocyte apoptotic index. More importantly, pretreatment with breviscapine resulted to a decrease in the levels of inflammatory and pro-apoptotic mRNAs and proteins in myocardial tissues and there was an increase in the levels of anti-apoptotic mRNAs and proteins. However, these protective effects were eliminated when breviscapine was combined with LY294002. CONCLUSION: The findings from this study indicated that breviscapine may inhibit myocardial inflammation and apoptosis by regulating the PI3K/protein kinase B (Akt)/glycogen synthase kinase-3ß (GSK-3ß) pathway, thereby ameliorating CME-induced cardiac dysfunction and reducing myocardial injury.

Puerarin pretreatment attenuates cardiomyocyte apoptosis induced by coronary microembolization in rats by activating the PI3K/Akt/GSK-3ß signaling pathway.

Coronary microembolization (CME) is associated with cardiomyocyte apoptosis and cardiac dysfunction. Puerarin confers protection against multiple cardiovascular diseases, but its effects and specific mechanisms on CME are not fully known. Hence, our study investigated whether puerarin pretreatment could alleviate cardiomyocyte apoptosis and improve cardiac function following CME. The molecular mechanism associated was also explored. A total of 48 Sprague-Dawley rats were randomly divided into CME, CME + Puerarin (CME + Pue), sham, and sham + Puerarin (sham + Pue) groups (with 12 rats per group). A CME model was established in CME and CME + Pue groups by injecting 42 µm microspheres into the left ventricle of rats. Rats in the CME + Pue and sham + Pue groups were intraperitoneally injected with puerarin at 120 mg/kg daily for 7 days before operation. Cardiac function, myocardial histopathology, and cardiomyocyte apoptosis index were determined via cardiac ultrasound, hematoxylin-eosin (H&E) and hematoxylin-basic fuchsin-picric acid (HBFP) stainings, and TdT-mediated dUTP nick-end labeling (TUNEL) staining, respectively. Western blotting was used to measure protein expression related to the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt)/glycogen synthase kinase-3ß (GSK-3ß) pathway. We found that, puerarin significantly ameliorated cardiac dysfunction after CME, attenuated myocardial infarct size, and reduced myocardial apoptotic index. Besides, puerarin inhibited cardiomyocyte apoptosis, as revealed by decreased Bax and cleaved caspase-3, and up-regulated Bcl-2 and PI3K/Akt/GSK-3ß pathway related proteins. Collectively, puerarin can inhibit cardiomyocyte apoptosis and thus attenuate myocardial injury caused by CME. Mechanistically, these effects may be achieved through activation of the PI3K/Akt/GSK-3ß pathway.

Elevation of erythrocyte sedimentation rate and C-reactive protein levels reflects renal interstitial inflammation in drug-induced acute tubulointerstitial nephritis.

BACKGROUND: A renal biopsy is needed to define active inflammatory infiltration and guide therapeutic management in drug-induced acute tubulointerstitial nephritis (D-ATIN). However, factors such as various contraindications, refusal of informed consent and limited technical support may stop the biopsy process. It is thus of great importance to explore approaches that could deduce probable pathologic changes. METHODS: A total of 81 biopsy-proven D-ATIN patients were enrolled from a prospective cohort of ATIN patients at Peking University First Hospital. The systemic inflammation score (SIS) was developed based on the CRP and ESR levels at biopsy, and patients were divided into high-SIS, median-SIS, and low-SIS groups. The demographic data, clinicopathologic features, and renal outcomes were compared. RESULTS: The SIS was positively correlated with inflammatory cell infiltration and was inversely correlated with interstitial fibrosis. The number of interstitial inflammatory cells increased significantly with increasing SISs. The proportions of neutrophils and plasma cells were the highest in the high-SIS group compared with the other two groups. Prednisone (30-40 mg/day) was prescribed in all patients. The high-SIS group tended to have more favorable renal restoration than the other two groups. By 12 months postbiopsy, a decreased eGFR (< 60 mL/min/1.73 m2) was observed in 66.7% of medium-SIS patients, 32.4% of high-SIS patients, and 30.4% of low-SIS patients. CONCLUSION: The SIS was positively correlated with active tubulointerstitial inflammation and therefore could help to aid therapeutic decisions in D-ATIN.

Diagnostic roles of urinary kidney injury molecule 1 and soluble C5b-9 in acute tubulointerstitial nephritis.

Acute tubulointerstitial nephritis (ATIN) is a common cause of acute kidney injury characterized by inflammatory cells infiltrating in the interstitium. The present study aimed to explore noninvasive biomarkers that might indicate activity of pathological injuries and help direct treatment. Fifty-four patients with clinical-pathologically diagnosed ATIN from January 1, 2014, to June 30, 2016, at Peking University First Hospital were enrolled. Urine samples were collected on the morning of renal biopsy and assessed for urinary kidney injury molecule-1 (KIM-1) and urinary soluble C5b-9 (sC5b-9). Immunofluorescence staining for KIM-1 and C5b-9 was performed in biopsied kidney sections from ATIN cases. The clinical and pathological relevance of the two urinary biomarkers was analyzed. Both urinary KIM-1 and sC5b-9 values were significantly elevated in patients with ATIN compared with healthy controls. The urinary KIM-1 level positively correlated with urinary N-acetyl-ß-d-glucosaminidase (r = 0. 542, P = 0.001) and the pathological tubular injury score (r = 0.469, P < 0.001), whereas the urinary sC5b-9 level was related to pathological activity scores for tubular injury (r = 0.413, P = 0.002), interstitial inflammation (r = 0.388, P = 0.004), and treatment response (r = 0.564, P < 0.001). Urinary KIM-1 tended to have better diagnostic value for tubular injury than urinary sC5b-9, whereas only urinary sC5b-9 was able to demonstrate severe interstitial inflammation. A combination of urinary KIM-1 and sC5b-9 had an area under the receiver-operating characteristic curve of 0.864 (95% confidence interval: 0.766-0.963, P < 0.001, sensitivity: 75%, specificity: 88%) for acute tissue injury in ATIN. KIM-1 expression was markedly increased in renal tubular cells in both ATIN and acute tubular necrosis conditions, whereas a significant upregulation of C5b-9 was only detected in the tubular cells and interstitial cells in ATIN cases. Urinary KIM-1 is a specific biomarker for renal tubular injury in ATIN, whereas urinary sC5b-9 is valuable in demonstrating severe interstitial inflammation. The combination of these two biomarkers helps identify patients at an acute injury stage and, therefore, might facilitate clinical evaluation and guide immunosuppressive therapy.

In vivo MRI detection of atherosclerosis in ApoE-deficient mice by using tenascin-C-targeted USPIO.

BACKGROUND: Atherosclerosis is the main cause of cardiovascular and cerebrovascular diseases. Non-invasive molecular imaging to detect and characterize the plaques is essential for reducing life-threatening cardiovascular events. PURPOSE: To investigate the possibility of the anti-tenascin-C-USPIO specific probe as a molecular marker of atherosclerotic plaques detected by 7.0-T magnetic resonance imaging (MRI). MATERIAL AND METHODS: Twenty ApoE-/- mice fed with a high fat diet were used for detecting the aorta arch atherosclerotic plaques by 7.0-T MRI at 16 and 24 weeks. Ten mice in the targeted group were injected with anti-tenascin-C-USPIO and another ten in the control group were injected with pure USPIO (n = 5 each time point in each group). Histopathologic examination was used to evaluate the plaques and immunohistochemistry analysis was used to compare tenascin-C expression. RESULTS: The relative signal intensity (rSI) changes of the targeted group decreased more than those of the control group (16 weeks: -15.65 ± 0.78% vs. -3.43 ± 2.57%; 24 weeks: -26.38 ± 1.54% vs. -11.12 ± 1.60%, respectively; P < 0.05). Histopathological analyses demonstrated visible atherosclerotic plaques formation and development over time from 16 weeks to 24 weeks. Tenascin-C expression of the plaques at 24 weeks was higher than that at 16 weeks (0.22 ± 0.04 vs. 0.13 ± 0.02, P < 0.05). The MR images correlated well with the progression of atherosclerotic plaques. CONCLUSION: Tenascin-C expression increased with the progression of atherosclerosis. Anti-tenascin-C-USPIO could provide a useful molecular imaging tool for detecting and monitoring atherosclerotic plaques by MRI.

A novel three-dimensional tumorsphere culture system for the efficient and low-cost enrichment of cancer stem cells with natural polymers.

Cancer stem cells (CSCs) are considered to serve a key role in tumor progression, recurrence and metastasis. Tumorsphere culture is the most important method for enriching CSCs and is widely used in basic research and drug screening. However, the traditional suspension cell culture system has several disadvantages, including low efficiency, high cost and difficult procedure, making it difficult to produce tumorspheres on a large scale. In the present study, two biomaterials, methylcellulose (MC) and gellan gum (GG), were used to construct a novel culture system based on the traditional system. Subsequently, the characteristics of the novel three-dimensional (3D) culture system were evaluated, the design scheme was optimized, and the morphological and biological features of the tumorspheres cultured in this 3D system were compared with the traditional system. The results revealed that the tumorspheres cultured in the novel 3D system presented a higher seeding density and improved morphology, while maintaining stem-like properties. This evidence suggests that a simple, efficient and low-cost culture system that produces tumorspheres on a large scale was successfully constructed, which can be widely used in various aspects of stem cell research.

Characteristics and comparison between diabetes mellitus and non-diabetes mellitus among chronic kidney disease patients: A cross-sectional study of the Chinese Cohort Study of Chronic Kidney Disease (C-STRIDE).

Although the prevalence of chronic kidney disease (CKD) and diabetes mellitus (DM) is increasing globally, information on Chinese CKD patients with DM is lacking. A total of 3499 pre-dialysis CKD patients from across China were enrolled in the Chinese Cohort Study of Chronic Kidney Disease (C-STRIDE) between November 2011 and April 2016. We divided the C-STRIDE patients into CKD with DM and CKD without DM groups and compared their clinical, demographic, and laboratory data in this cross-sectional study. CKD patients with DM were older, had a higher male-to-female ratio, and had more complications than CKD patients without DM. Age, smoking, and 24-h urinary protein levels were associated with co-occurrence of CKD and DM. Less than 50% of patients in either group took antilipemic, cardiovascular, cerebrovascular, or anti-anemic drugs. In addition, only 18.38% of CKD patients with DM had undergone a renal biopsy, and diabetic nephropathy was confirmed in 35.4% of them. Our findings suggest that several types of medication and renal biopsies should be used more frequently in the treatment of Chinese CKD patients with DM.