scholarly journals Function of miR-24 and miR-27 in Pediatric Patients With Idiopathic Nephrotic Syndrome

2021 ◽  
Author(s):  
Fen-fen Ni ◽  
Guang-lei Liu ◽  
shi-lei Jia ◽  
Ran-ran Chen ◽  
Li-bing Liu ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Nephrotic Syndrome ◽  
Idiopathic Nephrotic Syndrome ◽  
Gradient Centrifugation ◽  
Th2 Cells ◽  
Rt Pcr ◽  
Critical Function ◽  
Significant Difference ◽  
Ficoll Density Gradient Centrifugation ◽  
Ficoll Density Gradient

Abstract Background: The specific etiology and mechanism of idiopathic nephrotic syndrome (INS) in children remain unclear, so we investigated the pathogenesis of INS by measuring the effects two specific miRNAs on Th2 cells in children with this disease. Methods: Flow cytometry was used to measure the levels of Th2 cells and a cytometric bead array was used to measure the levels of IgE, interleukin (IL) -4, and IL-13. RT-PCR was used to measure the levels of miR-24 and miR-27 in CD4+TCD25− cells. PBMCs were isolated using Ficoll density gradient centrifugation, and transfected with different mimic or inhibitor miRNAs. RT-PCR was used to measure the expression of different RNAs, and flow cytometry was used to determine the percentages of Th2 cells. Results: Children with active INS had higher percentages of Th2 cells than healthy controls (P<0.05), but there was no significant difference for children in remission. The plasma levels of IgE, IL-4, and IL-13 were significantly increased in children with active INS (P<0.05). miR-24 and miR-27 were at lower levels in children with active non-atopic INS (P<0.05). Transfection experiments indicated that upregulation of each miRNA decreased the percentage of Th2 cells and the level of IL-4 (P<0.05), and down-regulation of each miRNA had the opposite effects (P<0.05). Conclusion: Children with active INS, with or without atopy, had higher levels of IgE, possibly related to their higher levels of IL-13 and IL-4 due to drift toward Th2 cells. miR-24 and miR-27 suppress the expression of Th2 cells and have a critical function in Th2 expression in INS.

scholarly journals Function of miR-24 and miR-27 in Pediatric Patients With Idiopathic Nephrotic Syndrome

2021 ◽  
Vol 9 ◽  
Author(s):  
Fen-fen Ni ◽  
Guang-lei Liu ◽  
Shi-lei Jia ◽  
Ran-ran Chen ◽  
Li-bing Liu ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Nephrotic Syndrome ◽  
Idiopathic Nephrotic Syndrome ◽  
Gradient Centrifugation ◽  
Th2 Cells ◽  
Healthy Controls ◽  
Rt Pcr ◽  
Critical Function ◽  
Significant Difference ◽  
Ficoll Density Gradient Centrifugation

Purpose: We investigated the pathogenesis of idiopathic nephrotic syndrome (INS) by measuring the effects two specific miRNAs on Th2 cells in children with this disease.Methods: After informed consent, we enrolled 20 children with active INS before steroid initiation, 20 children with INS in remission after steroid therapy, and 20 age-matched healthy controls. Flow cytometry was used to measure the levels of Th2 cells and a cytometric bead array was used to measure the levels of IgE, interleukin (IL)−4, and IL-13. RT-PCR was used to measure the levels of miR-24 and miR-27 in CD4+TCD25− cells. PBMCs were isolated using Ficoll density gradient centrifugation, and transfected with different mimic or inhibitor miRNAs. RT-PCR was used to measure the expression of different RNAs, and flow cytometry was used to determine the percentage of Th2 cells.Results: Relative to healthy controls, children with active INS had higher percentages of Th2 cells (P &lt; 0.05), but there was no significant difference in controls and children in remission. The plasma levels of IgE, IL-4, and IL-13 were significantly increased in children with active INS (P &lt; 0.05). There were lower levels of miR-24 and miR-27 in children with active non-atopic INS (P &lt; 0.05). Transfection experiments indicated that upregulation of each miRNA decreased the percentage of Th2 cells and the level of IL-4 (P &lt; 0.05), and down-regulation of each miRNA had the opposite effects (P &lt; 0.05).Conclusion: Children with active INS, with or without atopy, had higher levels of IgE, possibly related to their higher levels of IL-13 and IL-4 due to a drift toward Th2 cells. miR-24 and miR-27 suppressed the expression of Th2 cells and have a critical function regulating Th2 cell expression in INS.

RNA expression as a prognostic tool in idiopathic nephrotic syndrome

2007 ◽  
Vol 148 (23) ◽  
pp. 1067-1075
Author(s):  
Krisztina Fischer ◽  
Orsolya Galamb ◽  
Béla Molnár ◽  
Zsolt Tulassay ◽  
András Szabó
Keyword(s):  
Nephrotic Syndrome ◽  
Northern Blot ◽  
Idiopathic Nephrotic Syndrome ◽  
Minimal Change ◽  
Ribonuclease Protection Assay ◽  
Rna Expression ◽  
Rt Pcr ◽  
Protection Assay ◽  
Ribonuclease Protection

A gyermekkori nephrosis 90%-a idiopathiás nephrosis szindróma. Az idetartozó három kórkép, a minimal change betegség, a mesangialis proliferatio és a focalis sclerosis hasonló klinikai képpel jelentkező, eltérő prognózisú és terápiás válaszú betegség. Dolgozatunk célja az idiopathiás nephrosis szindrómába tartozó kórképek kialakulásával, progressziójával összefüggő genetikai ismeretek, génexpressziós változások áttekintése és funkcionális csoportosítása. A génexpressziós változások meghatározásának eszközeként, dolgozatunk röviden összefoglalja a northern blot, a ribonuclease protection assay, az in situ RNS-hibridizáció, a kvantitatív RT-PCR és a microarray módszerek lényegét. Az eddig elvégzett vizsgálatok a DNS-szintézis és repair gének, növekedési faktorok, extracelluláris mátrix, extracelluláris ligandreceptorok, extracelluláris jelátvitel zavarai mellett kiemelik a metabolikus és transzporter gének, illetve az immunszabályozó gének molekuláris eltéréseit, amelyek összefüggésben vannak az idiopathiás nephrosis szindróma eddig megismert molekuláris hátterével. A chiptechnológia fejlődésével és elterjedésével ezek a markerek és a hagyományos vizsgálati módszerek párhuzamos alkalmazása rutindiagnosztikai szempontból is fontossá válhat.

scholarly journals A Flow Cytometry-Based Assay for the Measurement of Total Complement Activity in the Serum and Clinical Practice

2022 ◽  
Vol 2022 ◽  
pp. 1-12
Author(s):  
Xuewei Ding ◽  
Shijun Li ◽  
Hui Liu
Keyword(s):  
Flow Cytometry ◽  
Nephrotic Syndrome ◽  
Linear Relationship ◽  
High Volume ◽  
Healthy Individuals ◽  
Serum Samples ◽  
Complement Activity ◽  
Significant Difference ◽  
Cell Groups ◽  
Total Complement

Objective. To develop a novel sensitive and accurate assay suitable for high-volume testing of the total complement activity in the serum for clinical laboratories. Methods. The total complement activity (TCA) to be measured was quantified by detecting the number of fragments produced by erythrocyte lysis and the erythrocyte fragmentation index (EFI), indicating TCA. EFI = M × M 2 / M 1 + M 2 , where M is the number of erythrocyte fragments (removed from the background), M 1 is the number of unagglutinated red cells, M 2 is the number of agglutinated red cell groups, and M 2 / M 1 + M 2 is the agglutination coefficient indicating the degree of erythrocyte agglutination. Mild changes in hemolysin and erythrocyte concentrations were made to optimize the testing conditions. The same serum samples were tested for 10 consecutive days to determine the stability of the experimental results. Serum EFI was detected in both nephrotic syndrome patients and healthy subjects. Results. There was a linear relationship between hemolysin and erythrocyte agglutination ( r = 0.999 , P < 0.001 ). A good linear relationship existed between EFI and TCA ( r = 0.991 , P < 0.001 ). The results were not affected by slight fluctuations in the concentrations of hemolysin or erythrocytes. The interbatch CV = 8.6 % of the test results showed good stability. There was a significant difference in the EFI between nephrotic syndrome patients and healthy individuals, P < 0.001 , and EFI was reduced in nephrotic syndrome patients compared to healthy individuals. Conclusion. The flow cytometry-based assay for TCA was sensitive and accurate and had potential value for clinical application.

scholarly journals Gamma-H2AX biodosimetry for use in large scale radiation incidents: comparison of a rapid lyse/fix protocol with a routine method

2013 ◽  
Author(s):  
Jayne Moquet ◽  
Stephen G R Barnard ◽  
Kai Rothkamm
Keyword(s):  
Large Scale ◽  
Processing Time ◽  
Clinical Symptoms ◽  
Gradient Centrifugation ◽  
Routine Method ◽  
Sample Processing ◽  
Ficoll Density Gradient Centrifugation ◽  
One Step ◽  
The Cost ◽  
Ficoll Density Gradient

Following a radiation incident, preliminary dose estimates made by γ-H2AX foci analysis can supplement the early triage of casualties based on clinical symptoms. Sample processing time is important when many individuals need to be rapidly assessed. A protocol was therefore developed for high sample throughput that requires less than 0.1 ml blood, thus enabling finger prick sampling. The technique combines red blood cell lysis and leukocyte fixation in one step on a 96 well plate, in contrast to the routine protocol, where lymphocytes are separated by Ficoll density gradient centrifugation with subsequent washing and fixation steps. The rapid lyse/fix method reduced the estimated sample processing time for 96 samples to about 4 h compared to 15 h using the routine protocol. However, scoring 20 cells in 96 samples prepared by the rapid protocol took longer than for the routine method (3.1 versus 1.5 h at zero dose; 7.0 versus 6.1 h for irradiated samples). Similar foci yields were scored for both protocols and reliable dose estimates were obtained for coded samples, with mean absolute differences from the actual doses of 0.26 and 0.27 Gy for the routine and lyse/fix method, respectively. The lyse/fix protocol can therefore facilitate high throughput processing for γ-H2AX biodosimetry for use in large scale radiation incidents, at the cost of somewhat longer foci scoring times.

scholarly journals MiR-24 and miR-27 negatively regulate the expression of Th2 cells in children with idiopathic nephrotic syndrome

Authorea ◽  
2020 ◽  
Author(s):  
Cheng rong Li ◽  
Fen Ni ◽  
Guang lei Liu ◽  
Jun Yang ◽  
Shi lei Jia ◽  
...  
Keyword(s):  
Nephrotic Syndrome ◽  
Idiopathic Nephrotic Syndrome ◽  
Th2 Cells

scholarly journals Effect of calcium and vitamin D supplementation on serum calcium level in children with idiopathic nephrotic syndrome

2014 ◽  
Vol 54 (3) ◽  
pp. 162 ◽  
Author(s):  
Vaya Dasitania ◽  
Alex Chairulfatah ◽  
Dedi Rachmadi
Keyword(s):  
Vitamin D ◽  
Nephrotic Syndrome ◽  
Serum Calcium ◽  
Idiopathic Nephrotic Syndrome ◽  
Controlled Trial ◽  
Vitamin D Supplementation ◽  
Significant Difference ◽  
Low Levels ◽  
Single Blind ◽  
Elemental Calcium

Background Patients with idiopathic nephrotic syndrome (NS) maydevelop hypocalcemia caused by low levels of albumin and vitaminD -binding protein, which subsequently decreases calcium absorptionin the intestine. Hypocalcemia may result in neuromuscularmanifestations, such as Chvostek's and Trosseau's signs.Objectives To evaluate the effect of calcium and vitamin Dsupplementation on hypocalcemia and its clinical manifestationsin idiopathic NS children.Methods A randomized, single-blind, controlled trial wasperformed in idiopathic NS patients aged 1-14 years. Subjectswere divided into treatment and placebo groups. Subjects inthe treatment group received 800 mg elemental calcium and400 IU vitamin D supplementation, while they in control groupreceived placebo syrup, both for 8 weeks. Serum calcium andmanifestations of hypocalcemia were examined before and aftersupplementation.Results Thirty subjects completed the study (15 in each group).Seventeen subjects experienced hypocalcemia. Chvostek's andTrosseau's signs were observed in 6 subjects in the treatment groupand 2 subjects in the placebo group (P= 0.427). After 8 weeks ofintervention, Chvostek's and Trosseau's signs disappeared in bothgroups, and calcium levels were significantly increased in bothgroups compared to the levels before intervention. However, therewas no significant difference in serum calcium levels after 8 weeksbetween the treatment and placebo groups (P =0.707).Conclusion Normalization of serum calcium levels and improvedclinical manifestations ofhypocalcemia occur both in NS patientswho receive calcium and vitamin D supplementation and thosewho do not.

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