Objective To investigate the relationship between the expression of Toll-like receptor 4 (TLR4) on mononuclear cells in peripheral blood and changes of serum vitamin D level in children with bronchiolitis.Methods The children who were diagnosed as bronchiolitis and received treatment in the Second Affiliated Hospital of Harbin Medical University from October 2013 to January 2014 were chosen as the pre-treatment group,and then divided them into moderate group and severe group according to the clinical symptoms,20 cases for each group.Then the cases in pre-treatment group who recovered after treatment were recruited as the after-treatment group,and the children who were healthy and medical examination in the Second Affiliated Hospital of Harbin Medical University in the same period were recruited as the healthy control group.The expressions of TLR4 on CD14 labeled mononuclear cells in the periphera were measured by flow cytometry.The level of 25 (OH) D in serum was detected by enzyme linked immunosorbent assay (ELISA).Results (1) The expression level in children with bronchiolitis of TLR4:the mode-rate group [(18.98 ±2.29)％] and severe group [(30.13 ±2.13)％] increased significantly (P ＜0.05) compared with control group [(1.17 ± 0.57) ％].And the expression level of moderate group [(2.02 ± 0.48) ％] and severe group [(11.43 ± 1.52) ％] decreased significantly after treatment (P ＜0.05).(2) Serum vitamin D level in children with bronchiolitis of the moderate group[(17.16 ± 3.34) μg/L] and severe group [(6.56 ± 2.28) μg/L] were lower than healthy control group [(53.69 ± 20.18) μg/L] before treatment (P ＜ 0.05),especially the severe group [(6.56 ±2.28) μg/L].The level of moderate group [(9.59 ± 2.31) μg/L] and severe group [(4.70 ± 0.67) μg/L] became lower after treatment (P ＜ 0.05).(3) Both severe group (r =-0.491,P ＜ 0.05) before treatment and moderate group (r =-0.436,P ＜ 0.05) after treatment showed negative correlation between TLR4 on mononuclear cells in peripheral blood and serum 25 (OH)D level in children with bronchiolitis.And no correlation was found among healthy control group,moderate group before treatment and severe group after treatment (P ＞ 0.05).Conclusions The conditions of children with bronchiolitis was positively correlated with the expression level of TLR4,and negatively correlated with the vitamin D level.The serum 25 (OH) D decreased steadily during the treatment.The expression of TLR4 in monocytes has a certain correlation with the level of vitamin D in children with bronchiolitis.

Objective:To examine the expression ofphospholipase A2 receptor (PLA2R) in renal tissues and the level of anti-PLA2R antibody in serum in patients with idiopathic membranous nephropathy (IMN) and secondary membranous nephropathy (SMN),and to evaluate their diagnostic value in IMN.Methods:A total of 73 patients,who were diagnosed between May,2014 and February,2015 in the Department of Nephrology of the Second Xiangya Hospital,Central South University,were divided into three groups:an IMN group (n=48),an SMN group (n=17) and a minimal change disease group (n=8) according to the renal biopsy.PLA2R expression in renal tissues and the level of antiPLA2R antibody in serum were detected by indirect immunofluorescence technique.Results:The positive rate and fluorescence intensity for PLA2R in the renal tissues in the IMN group were higher than those in the SMN group (91.7％ in the IMN group vs 29.4％ in the SMN group,P＜0.05),while the positive rate and serum level for anti-PLA2R antibody in the IMN group were higher than those in the SMN group (85.4％ in the IMN group vs 29.4％ in the SMN group,P＜0.05);the expression of PLA2R in renal tissues and the serum level for anti-PLA2R antibody were not detected in the minimal change disease group,The serum level of anti-PLA2R antibody was positively correlated with 24 h urine protein (r=0.432,P＜0.01) and negatively correlated with serum albumin (r=-0.307,P＜0.05).Conclusion:The expression of PLA2R in renal tissues and the serum level of anti-PLA2R antibody might be potential markers for diagnosis oflMN.

OBJECTIVE: To investigate the new pathological classification of diabetic nephropathy (DN) published by Research Committee of the Renal Pathology Society in 2010. METHODS: Renal biopsy specimens were obtained from 37 patients with type 2 diabetes mellitus with micro-albuminuria (MAU) or clinical albuminuria (CAU). These samples were classified according to new pathological classification for DN and new standard scores for interstitial vascular injury. RESULTS: Before the classification, DN was seen in 26 palients. After re-analysis according to the new pathological classification, the patients diagnosed with DN increased to 32. In these 32 DN patients, 1 was classified as type I, 3 as type IIa, 2 as type IIb, 23 as type III and 3 as type IV; 12 patients had mild interstitial injury, 15 had midrange interstitial injury, while 5 had severe interstitial injury. CONCLUSION The new pathological classification of DN can increase the diagnosis rate and attract more attention to tubular and interstitial damage in DN, contributing to the early diagnosis and treatment of DN.
Adult ; Aged ; Albuminuria ; pathology ; Diabetes Mellitus, Type 2 ; complications ; pathology ; Diabetic Nephropathies ; classification ; pathology ; Female ; Humans ; Male ; Middle Aged ; Reference Standards ; Retrospective Studies

Objective:To establish a method for iohexolquantification based on high performance liquid chromatography (HPLC) to measure the concentration of blood iohexolafterlow dose and contrast dose injection.Methods:Weperformed the method establishment and evaluation in this study. A HPLC-UV system (high performance liquid chromatography plus ultraviolet detector) was used to establish the method. The linearity, imprecision, recovery rate, limit of detection, lower limit of measuring interval and carryover of the method were evaluated. The stability of iohexol under different storage conditions, the differences of iohexolbetween serum and plasma concentrations, and the drug′s interference with the method were evaluated preliminarily. The single sample t test was used for the stability test of iohexolin samples, and the Wilcoxon symbol rank sum test was used for the comparison of iohexol concentrations between serum and plasma.Results:The linearity of iohexol ranging from 5 to 250 μg/ml ( R2=0.999 9) and from 250 to 4 000 μg/ml ( R2=0.999 8); when the concentration of iohexol was 20-3 000 μg/ml, the intra-and inter-assay coefficient of variation were 1.63% to 3.31% and 2.10% to 4.09%, respectively. The recovery rate was 94.17% to 106.13%; the limit of detection was 1 μg/ml and the lower limit of measuring interval was 5 μg/ml; it shows no carryover at the concentration of iohexol 4 000 μg/ml; after 48 hours at room temperature storage, the relative deviation of the concentration was -5.55% to +5.58%, after repeated freeze-thaw cycles 6 times at -80 ℃, the relative deviation of the concentration was -1.28% to+6.68%; there was no statistic difference between the measurement results between serum and plasma; valsartan and other drugs did not interfere with this methodsignificantly. Conclusion:Awide-range HPLC method for iohexolquantification has been established, which can stably and accurately detect the blood concentration of iohexol at low and contrast doses.

Objective:This work aims to assess the distribution of peripheral blood monocyte subsets, the expression level of the functional markers in rheumatoid arthritis (RA) patients, and analyze the correlation between the above indexes and the onset of RA.Methods:Peripheral blood mononuclear cells were collected and isolated from 62 RA patients, 52 healthy control (HC) and 12 disease control group′s patients via density centrifugation. The enrolled patients were attended or underwent physical examination in East Hospital, Tongji University from June 2020 to December 2021. Monocytes could be classified into classical (CM), intermediate (IM) and non-classical (NCM). Then, the flow cytometry was performed to examine the distribution of monocyte subsets and the measure the expression level of human leukocyte antigen DR (HLA-DR), intracellular tumor necrosis factor α (TNF-α) in peripheral blood monocytes. The statistical methods in this study mainly include: Kruskal-Wallis H test, Chi-Square test, Mann-Whitney U test, Wilcoxon matched-pairs signed ranks test, Spearman correlation coefficient test and Logistic regression analysis. The diagnostic value of IM proportion in RA was analyzed by ROC curve. Results:The monocytes number and monocytes proportion in white blood cells were much higher in RA [0.40 (0.40, 0.50), 7.60% (5.97%, 8.53%)] and disease control [0.40 (0.40, 0.68), 8.20% (5.85%, 10.28%)] compared with HC [0.30 (0.30, 0.40), 5.80% (5.03%, 6.38%)] ( H=24.733, P<0.001; H=27.469, P<0.001). A statistic-significant difference was detected among the proportion of CM[85.49%(76.91%,89.21%),88.94%(86.36%,91.72%),90.26%(80.25%, 92.56%)],IM[11.65%(8.47%,17.89%),7.89%(5.36%,10.75%), 5.56%(4.17%, 8.27%)], NCM[2.22%(1.39%, 3.74%), 2.49%(1.74%, 4.66%), 5.13%(3.39%, 9.85%)] in RA group, HC group and disease control group ( H=11.389, P=0.003; H=20.815, P<0.001; H=10.640, P=0.005). The proportion of CM was lower in RA and the IM proportion was increased in RA( P=0.003; P=0.003). The intracellular TNF-α level of monocytes in all three groups revealed the trend that IM>NCM>CM. The intracellular TNF-α in IM of RA was positively associated with serum TNF-α ( r=0.376, P=0.041). The HLA-DR expression in IM subsets were higher than CM and NCM subsets in all RA,HC and disease control groups. The expression of HLA-DR of IM in RA group and disease control was higher than HC group [8 611.50 (6201.3, 9890.8), 10 295.0 (7 899.0, 13632.0), 6 278.00(4 057.8, 9522.0), H=10.495, P=0.005]. There were no correlations between the proportion of peripheral blood IM and clinical characteristics CRP ( r=0.119, P=0.359), RF ( r=0.204, P=0.112) and ESR ( r=0.153, P=0.236). Logistic regression analysis showed that the proportion of IM ( OR=1.169, 95% CI 1.003-1.363, P=0.046), CRP ( OR=1.277, 95% CI 1.000-1.631, P=0.050), RF ( OR=1.179, 95% CI 1.080-1.287, P<0.001) are positively correlated with RA onset. The area under ROC curve for diagnosis of RA with IM proportion was 0.687, and the 95% confidence interval was 0.590-0.784, P<0.001. Conclusions:The distribution of monocyte subsets in peripheral blood of RA patients is abnormal. The increase in the proportion of IM, the enhanced antigen-presenting ability, and the increased level of TNF-α secretion in RA patients may play an important role in the pathogenesis of RA.

ObjectiveTo observe the effects of Guben Fangxiao decoction-containing serum on the expression of transcription factors and cytokines associated with calcitonin gene-related peptide （CGRP）， γ-aminobutyric acid （GABA）， and type Ⅱ innate lymphoid cells （ILC2） in human bronchial epithelial cells （BEAS-2B） stimulated with interleukin-4 （IL-4） combined with interleukin-13 （IL-13）， and explore the possible mechanism of Guben Fangxiao decoction-containing serum in alleviating type Ⅱ inflammation of bronchial epithelial cells. MethodsBEAS-2B cells stimulated with IL-4 combined with IL-13 were treated with the sera containing high （20%）， medium （15%）， and low （10%） doses of Guben Fangxiao decoction and the montelukast-containing serum （10%）. Real-time PCR was used to measure the mRNA levels of CGRP， GABA， mucin 5AC （MUC5AC）， thymic stromal lymphopoietin （TSLP）， interleukin （IL）-25， IL-33， IL-5， GATA-binding protein 3 （GATA3）， and B cell lymphoma/leukemia 11B （Bcl-11B） in each group. Western blot was employed to determine the relative protein levels of CGRP and MUC5AC. The immunofluorescence assay was employed to observe the relative expression of CGRP and GABA in the cells. Enzyme-linked immunosorbent assay was used to quantify the protein levels of IL-33， MUC5AC， and IL-5 in the culture and cell supernatants. Periodic acid-Schiff staining was performed to assess mucin secretion. ResultsCompared with the control group， the model group showed up-regulated mRNA levels of CGRP， GABA， MUC5AC， TSLP， IL-25， and IL-33 （P<0.05， P<0.01）. Compared with the normal group， the Guben Fangxiao decoction-containing serum down-regulated the mRNA levels of CGRP， GABA， MUC5AC， IL-5， GATA3， and Bcl-11B （P<0.05， P<0.01） in a dose-dependent manner. Compared with the normal group， the model group showed up-regulated protein levels of CGRP， GABA， MUC5AC， IL-33， and IL-5 （P<0.05， P<0.01）， which were down-regulated by the Guben Fangxiao decoction-containing serum （P<0.05， P<0.01）. Compared with the normal group， the model group showed enhanced average fluorescence intensity of CGRP and GABA （P<0.01）， which was weakened by the Guben Fangxiao decoction-containing serum （P<0.01）. Compared with the normal group， the model group showed increased secretion of mucin， which was reduced by the Guben Fangxiao decoction-containing serum. ConclusionGuben Fangxiao decoction can treat bronchial asthma by alleviating type Ⅱ immune airway inflammation and reducing airway mucus secretion， which is achieved by regulating the expression of CGRP， GABA， MUC5AC， and ILC2-related transcription factors.

OBJECTIVE: To investigate the mechanism of mitochondrial oxidative injury induced by high glucose peritoneal dialysis solution (PDS) and the protective effect of peroxisome proliferator activated receptor gamma coactivator 1-alpha (PGC-1α) in the mitochondria of human peritoneal mesothelial cells (HPMC) in the high glucose ambience. METHODS: HPMC was cultured in a PDS containing 1.5%, 2.5% and 4.25% glucose for 24 hours. Western blot analysis was used to detect PGC-1α expression. MitoSOX? Red staining, respiratory chain complexes and antioxidant enzyme activities were determined. RESULTS: The activities of respiratory chain complex III and antioxidant enzymes decreased significantly in a concentration- and time-dependent manner, along with the increased production of mitochondrial reactive oxygen species (ROS) and cellular apoptosis. In addition, protein expression of PGC-1α was also decreased in the high glucose PDS ambience. CONCLUSION High glucose PDS might inhibit PGC-1α expression, resulting in the inhibition of mitochondrial function and increase of mitochondrial ROS and cellular apoptosis.
Apoptosis ; Dialysis Solutions ; adverse effects ; Epithelial Cells ; pathology ; Glucose ; adverse effects ; Humans ; Mitochondria ; metabolism ; pathology ; Oxidative Stress ; Peritoneal Dialysis ; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha ; Reactive Oxygen Species ; Transcription Factors ; metabolism

Objective:To establish a rat model of neurogenic bladder and analyze the changes in kidney morphology and function and the expression of proteins in AngiotensinⅡ(AngⅡ)/transforming growth factor β1 (TGF-β1)/Smads pathway.Methods:Sprague-Dawley rats were randomly divided into experimental group (spinal nerve amputation, n=36) and control group (sham operation, n=12). At 6, 12, and 24 weeks, the bladder compliance was measured by cystometry, the kidney morphology was detected by B-ultrasound, blood urea nitrogen (BUN) and serum creatinine (Scr) in blood samples were examined, the kidney pathological changes were detected by Masson and HE staining, the distribution of AngⅡ/TGF-β1/Smads pathway proteins was analyzed by immunohistochemisty, and the protein expressions in kidney were detected by Western blotting. Results:Urodynamics showed that the basic bladder pressure in experimental group was higher than that in control group. B-ultrasound showed that compared with the control group, the diameter of the renal pelvis of the rats with nerve dissection gradually increased ( P<0.05), and the hydronephrosis was gradually obvious. Compared with the control group, the BUN and Scr in experimental group gradually increased (both P<0.01). Masson and HE staining showed that compared with the control group, the collagen expression and renal tubulointerstitial scores in experimental group were gradually increased (both P<0.01). Immunohistochemisty showed that compared with the control group, in experimental group the expression of angiotensinⅡ receptor type 1 (AT1), TGF-β receptor 1(TGF-βR1), phosphorylated Smad2 gradually increased (all P<0.01), the pathway inhibitor Smad6 gradually decreased ( P<0.01), and the distribution of each protein in kidney was consistent. Western blotting showed a corresponding expression trend with immunohistochemisty. Conclusions:In neurogenic bladder caused by bilateral spinal nerve amputation, due to bladder dysfunction, increased bladder pressure induces hydronephrosis, destruction of the nephron structure, activation of AngⅡ/TGF-β1/Smads pathway, and renal fibrosis. This method is effective and has clinical similarities, laying a foundation for exploring neurogenic bladder treatment.

Objective:With the in-depth study of complement dysregulation,glomerulonephritis with dominant C3 has received increasing attention,with a variety of pathologic types and large differences in symptoms and prognosis between pathologic types.This study analyzes the clinical,pathological,and prognostic characteristics of different pathological types of glomerulonephritis with dominant C3,aiming to avoid misdiagnosis and missed diagnoses. Methods:The clinical,pathological,and follow-up data of 52 patients diagnosed as glomerulonephritis with dominant C3 by renal biopsy from June 2013 to October 2022 were retrospectively analyzed.According to the clinical feature and results of pathology,15 patients with post-infectious glomerulonephritis(PIGN)and 37 patients with of non-infectious glomerulonephritis(N-PIGN)were classified.N-PIGN subgroup analysis was performed,and 16 patients were assigned into a C3-alone-deposition group and 21 in a C3-dominant-deposition group,or 27 in a C3 glomerulopathy(C3G)group and 10 in a non-C3 nephropathy(N-C3G)group. Results:The PIGN group had lower creatinine values(84.60 μmol/L vs 179.62 μmol/L,P= 0.001),lower complement C3 values(0.36 g/L vs 0.74 g/L,P<0.001)at biopsy,and less severe pathological chronic lesions compared with the N-PIGN group.In the N-PIGN subgroup analysis,the C3-dominant-deposition group had higher creatinine values(235.30 μmol/L vs 106.70 μmol/L,P=0.004)and higher 24-hour urine protein values(4 025.62 mg vs 1 981.11 mg,P=0.037)than the C3-alone-deposition group.The prognosis of kidney in the PIGN group(P=0.049),the C3-alone-deposition group(P=0.017),and the C3G group(P=0.018)was better than that in the N-PIGN group,the C3-dominant-deposition group,and the N-C3G group,respectively. Conclusion:Glomerulonephritis with dominant C3 covers a variety of pathological types,and PIGN needs to be excluded before diagnosing C3G because of considerable overlap with atypical PIGN and C3G;in addition,the deposition of C1q complement under fluorescence microscope may indicate poor renal prognosis,and relevant diagnosis,treatment,and follow-up should be strengthened.