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AIM: To compare the outcome of C3F8 versus silicone oil tamponade after pars plana vitrectomy(PPV)and inverted internal limiting membrane(ILM)for the treatment of highly myopic macular hole retinal detachment(MHRD).METHODS: Retrospective clinical study. Totally 45 patients(45 eyes)with highly myopic MHRD who visited our hospital between January 2019 and August 2022 were selected as the research subjects. The patients were divided into two groups according to different intraocular tamponade agents: C3F8(22 eyes)and silicone oil(23 eyes)groups. All patients underwent conventional three-incision PPV, ILM was tamped, a venous blood clot was placed on the tamped ILM, and 15% C3F8 and silicone oil were used as tamponade, respectively. The best corrected visual acuity(BCVA), multifocal electroretinogram(mfERG), the closure of the macular hole, retinal reattachment and the complications were observed.RESULTS: The macular hole closure rate was 77% in the C3F8 group and 83% in the silicone oil group, respectively(P>0.05), and retinal reattachment rates were 95% and 96%, respectively(P>0.05). The visual acuity of the two groups significantly improved, which was 0.99±0.34 and 1.22±0.37, respectively, and the C3F8 group was better than that of the silicone oil group(t=-2.156, P=0.037). After operation, the response density of the first ring of P1 wave in the first order kernel in mfERG was 114.27±26.37 nV/deg2 for the C3F8 group and 98.08±24.36 nV/deg2 for the silicone oil group, and the response density of the second ring of P1 wave was 80.45±14.94 nV/deg2 for the C3F8 group and 67.73±15.33 nV/deg2 for the silicone oil group, all of which were significantly higher compared to pre-operation [the response density of the first ring of P1 wave: 58.13±13.96 nV/deg2 for the C3F8 group and 55.30±10.48 nV/deg2 for the silicone oil group, the response density of the second ring of P1 wave: 51.18±8.19 nV/deg2 for the C3F8 group and 47.43±11.97 nV/deg2 for the silicone oil group](all P<0.05). It was found that the response density of the first ring of P1 wave was lower in the silicone oil group than in the C3F8 group(P<0.05). There was no statistically significant difference in the incidence of complications between the two groups(P>0.05).CONCLUSION: Silicone oil tamponade or C3F8 tamponade after PPV combined with ILM can both promote retinal reattachment and macular hole closure in patients with MHRD, and the C3F8 tamponade was superior to silicone oil in visual function recovery.
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Purpose: Outcome of topography?guided excimer laser ablation in conjunction with accelerated, high?fluence cross?linking in corneal ecstatic disease using the NIDEK CXIII equipped with CATz algorithm from the FinalFit software棓Bharat Protocol.� Methods: Retrospective case record review of 30 eyes of 17 patients of stage 1?3 keratoconus who underwent the procedure was performed. Data collected were for visual acuity, distortion?induced eye pain, and keratometry. Pachymetry, lower order and higher order aberrations, spherical aberrations, and topographic cylinder were documented from by Scheimpflug imaging (Pentacam 70700: Oculus, Wetzlar, Germany). Results: At a minimum follow?up of 6 months (range 6.2� months), there was significant improvement in UCVA (P < 0.00001), BCVA (P = 0.0061), decrease in Kmax (P = 0.0349), Ksteep (P < 0.0411), Kflat (P = 0.0099), and pachymetry (P = 0.0001). Significant improvement was also seen in distortion?induced eye pain (27/30 to 2/30; P < 0.00001). A more than two?line improvement in UCVA and BCVA was seen in 23/30 and 17/30 cases, respectively. Ectasia was stabilized in all cases at the last follow?up, and no complications were seen. Conclusions: The 揃harat� Protocol to arrest keratectasia progression and improve corneal regularity is a safe and efficacious alternative as a keratoconus management option. This is the first such study on Nidek Platform for the same.
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ObjectiveTo clone coumarate-3-hydroxylase gene （C3H） from Angelica sinensis， and analyze the correlation between its bioinformatics， expression patterns and content of ferulic acid， and to explore the functions of ASC3H. MethodReal-time polymerase chain reaction （Real-time PCR） was used to clone the full-length cDNA of ASC3H based on the transcriptome dataset of A. sinensis， and the bioinformatics analysis of the gene sequence was carried out. Real-time PCR and high performance liquid chromatography （HPLC） were used to determine relative expression of ASC3H and content of ferulic acid in different root tissues of A. sinensis （periderm， cortex and stele）. ResultThe open reading frame （ORF） of ASC3H （GenBank accession number： MN2550298） was 1 530 bp， encoding 509 amino acids， with a theoretical molecular weight of 57.86 kDa and an isoelectric point of 8.36. It was a hydrophilic protein that was located in the chloroplast with multiple phosphorylation sites and a transmembrane region， and contained a conserved domain CGYDWPKGYGPIINVW_P450 （383-399 aa） in cytochrome P450. Multiple amino acid sequence alignment analysis showed that ASC3H had high similarity with C3H from other plants， especially Ammi majus in Umbelliferae. The Real-time PCR revealed that ASC3H had different expressions in periderm， cortex and stele tissues of A. sinensis roots. It was found from HPLC that the cortex tissues had the highest content of ferulic acid， and the stele tissues had the lowest. ConclusionASC3H was successfully cloned from A. sinensis， and its sequence characteristics were understood more clearly， suggesting that ASC3H might be involved in the ferulic acid biosynthesis pathway of A. sinensis. This paper provided a basis for further studying the functions of the gene and exploring the biosynthesis and regulation mechanism of ferulic acid in A. sinensis， while laying the foundation for the genetic improvement of A. sinensis．
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ObjectiveTo explore the protective effect and mechanism of Qihong Tongluo prescription on vascular endothelial cells in rats with deep venous thrombosis （DVT）. MethodSixty-six SD rats were randomly divided into a blank group （n=11） and a modeling group （n=55）. The DVT model was induced in rats of the modeling group by slowing down blood flow and damaging vascular endothelium. The model rats were randomly divided into model group， aspirin group （200 mg·kg-1）， and low-，medium-， and high-dose Qihong Tongluo prescription groups （6.5， 13， 26 g·kg-1） according to a random number table. Rats were treated with corresponding drugs by gavage， while those in the model group and the blank group received normal saline， once per day for 7 days. The rats were sacrificed and the abdominal aortic blood was taken. The levels of serum endothelin-1 （ET-1） and interleukin-6 （IL-6） were detected by enzyme-linked immunosorbent assay （ELISA）. Hematoxylin-eosin （HE） staining was used to observe the pathological changes in vascular endothelial tissues. The ultrastructure of vascular endothelial cells was observed by the transmission electron microscope. The viability of vascular endothelial cells was detected by methylthiazolyldiphenyl-tetrazolium bromide （MTT） method，and the release level of lactate dehydrogenase （LDH） was detected by the LDH kit. The messenger ribonucleic acid （mRNA） expression of platelet-activating factor （PAF），nuclear transcription factor κB （NF-κB），Ras-related C3 botulinum toxin substrate 1 （Rac1）， and Ras-related C3 botulinum toxin substrate 2 （Rac2） in vascular endothelial tissues were detected by real-time reverse transcription polymerase chain reaction （Real-time PCR）. The protein expression of PAF，NF-κB，Rac1， and Rac2 in vascular endothelial tissues was detected by Western blot. ResultThe model group showed seriously damaged and swollen vascular endothelial cells with massive shedding， attachment of massive inflammatory cells， nucleus pyknosis and deformation under the electron microscope， highly swollen mitochondria， serious cytoplasmic vacuolation，and exposure of internal elastic membrane. The damage of vascular endothelium and its ultrastructure in Qihong Tongluo prescription groups and the aspirin group was improved in varying degrees. Compared with the blank group，the model group showed increased levels of serum ET-1 and IL-6，potentiated vascular endothelial cell viability， up-regulated mRNA and protein expression of PAF，NF-κB，Rac1， and Rac2 in vascular endothelial tissues，and decreased LDH release level of vascular endothelial cells （P<0.05）. Compared with the model group，the aspirin group and the Qihong Tongluo prescription groups showed decreased levels of serum ET-1 and IL-6，blunted vascular endothelial cell viability，down-regulated mRNA and protein expression of PAF，NF-κB，Rac1， and Rac2 in vascular endothelial tissues，and increased LDH release level of vascular endothelial cells （P<0.05）. The effect of Qihong Tongluo prescription was dose-dependent. ConclusionQihong Tongluo prescription has a protective effect on vascular endothelial cells of DVT rats and can prevent and treat thrombosis，and its therapeutic effect is presumably achieved by inhibiting the expression of PAF，NF-κB，Rac1，and Rac2 and reducing the levels of serum ET-1 and IL-6.
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Objective To study the role of complement C3a receptor (C3aR) in cognitive impairment in rats with sepsis induced by cecal ligation and puncture (CLP), and to explore the possible mechanism. Methods Totally 132 rats were randomly divided into sham operation group (sham group) and sepsis group (CLP group). The initial number of each group was 66 animals, and 22 animals were set at each time point. The SD rat CLP animal model was constructed, and serum and brain (hippocampus and prefrontal cortex) samples were collected at day 1, day 15 and day 30 after operation. The levels of tumor necrosis factor-α(TNF-α), interleukin-1β (IL-1β) and IL-6 in the samples were determined by ELISA. Western blotting detected Tau protein (Tau), phosphorylated Taup-Tauand C3aR expression in brain samples. Totally 66 rats were randomly divided into 3 groups, sham operation group, CLP group and C3aR antagonist(C3aRa) intervention group. On 15th, 17th, and 19th days after CLP, C3aRa intervened in rats, and they received inhibition avoidance test and object recognition test 30 days after CLP. The expressions of C3aR, lonized calcium binding adapter molecule 1(Iba1), GFAP and p-Tau in the hippocampus were detected by immunofluorescence. Results Compared with the sham group, the serum and brain tissue (TNFα, IL-1β and IL-6) of rats in the CLP group first increased and then decreased within 30 days after CLP. In the hippocampus and prefrontal cortex of CLP group, Tau phosphorylation was significantly enhanced at day 30 and day 1 and 15 after surgery, respectively (P<0.05). Compared with the sham group, C3aR protein levels in hippocampus and prefrontal cortex of rats in CLP group increased significantly at day 15 and 30 after operation (P<0.05). Compared with the CLP group, the endogenous C3aR content decreased significantly after C3aRa intervention (P<0.05), and Iba1, GFAP and p-Tau immunostaining were significantly inhibited (P<0.05). The C3aRa intervention in CLP rats reversed the cognitive impairment. Conclusion C3aR plays an important role in the development of biochemical and behavioral changes commonly associated with the onset of sepsis-induced neurodegenerative processes. C3aRa can be injected into the hippocampus to counteract the neurodegenerative process induced by sepsis.
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Heterotypic cell-in-cell structures (heCICs) are closely related to tumor development and progression, and have become a new frontier in life science research. Ras-related C3 botulinum toxin substrate 1 (Rac1) belongs to the classic Rho GTPase, which plays a key role in regulating the cytoskeleton and cell movement. To investigate the role and mechanism of Rac1 in the formation of heCICs, tumor cells and immune killer cells were labeled with cell-tracker, respectively, to establish the heCICs model. Upon treatment with the Rac1 inhibitor NSC23766, the formation of heCICs between tumor and immune cells was significantly reduced. The plasmid pQCXIP-Rac1-EGFP constructed by gene cloning was packaged into pseudoviruses that subsequently infect tumor cells to make cell lines stably expressing Rac1. As a result, the formation of heCICs was significantly increased upon Rac1 overexpression. These results demonstrated a promotive role of Rac1 in heCICs formation, which may facilitate treating cell-in-cell related diseases, such as tumors, by targeting Rac1.
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Background Silica nanoparticles (SiNPs) enter the human body through respiratory tract, digestive tract, and skin, causing body damage. Lung is one of the main damaged organs. Objective To observe the expressions of complement activated fragment C3a and its receptor C3aR in the lungs of mice exposed to SiNPs through respiratory tract, and to explore the involvement of C3a/C3aR in lung injury induced by SiNPs exposure. Methods The ultrastructure of SiNPs (particle size 5-20 nm) was determined under a transmission electron microscope, and the hydrodynamic diameter and surface Zeta potential of SiNPs were determined using a nanoparticle size analyzer. A total of 88 SPF C57BL/6J mice were randomly divided into five groups: a blank control group without any treatment (14 mice), a vehicle control group treated with 50 μL stroke-physiological saline solution by intratracheal instillation (14 mice), and three SiNPs exposure groups (low-dose group, medium-dose group, and high-dose group with 20 mice in each group, who were given 50 μL SiNPs suspension of 7, 21, and 35 mg·kg−1 respectively and exposed once every 3 days for 5 times). The mice were anesthetized on day 1 (1-day model group) and day 15 (15-day model group) after exposure, then sacrificed after extraction of bronchoalveolar lavage fluid (BALF), and lung tissues were retained. The morphological changes of lung tissues were observed by HE staining, the expression level of C3a in BALF was detected by enzyme-linked immunosorbent assay, the deposition of C3a and C3aR in lung tissues were observed by immunohistochemistry, the protein expression level of C3aR was determined by Western blotting, and the localization and semi-quantitative detection of C3a and C3aR in lung tissues was observed by immunofluorescence. Results SiNPs agglomerated in stroke-physiological saline solution. The average hydrodynamic diameter was (185.60±7.39) nm and the absolute value of Zeta potential was (43.33±0.76) mV. The condition of mice in the 1-day model group and the 15-day model group was good, while 2 mice died in the medium-dose group of the 1-day model group due to misoperation. The autopsy results of the two mice showed congestion of the lung tissue, emphysema, and no imperfection of trachea integrity. No death was observed in other dose groups. The HE staining results showed pathological damage to the mouse lung, including alveolar wall thickening and inflammatory cell infiltration after SiNPs exposure. The pathological damage became more serious with the increase of dose. Regarding pathological changes, the 15-day model group was slightly relieved compared with the 1-day model group, but there were still pathological changes. The enzyme-linked immunosorbent assay results showed that there was no difference in the expression level of C3a between the blank control group and the vehicle control group (P＞0.05), the expression levels of C3a in the medium-dose group and the high-dose group were significantly higher than that in the vehicle control group (P<0.05). The immunohistochemistry results showed that C3a deposition was consistent with the enzyme-linked immunosorbent assay results. The Western blotting and the immunohistochemistry results showed that C3aR expression was low in the blank control group and the vehicle control group, while the expression in each dose group tended to increase with the increase of dose. The immunofluorescence results showed that the fluorescence signals of C3a and C3aR were weak in the blank control group and the vehicle control group in the 1-day model group and the 15-day model group, while the fluorescence signals in the lung tissues of mice in the SiNPs exposure groups tended to increase with the increase of dose. Conclusion The increased expressions of C3a and C3aR in complement activation may be related to lung injury induced by intratracheal instillation of SiNPs, suggesting that C3a/C3aR may be involved in lung injury induced by SiNPs exposure.
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ObjectiveTo investigate the value of serum complement C3 level in determining the stage of liver fibrosis in primary biliary cholangitis （PBC）. MethodsClinical data were collected from 108 patients with PBC who attended Tianjin Second People’s Hospital and underwent liver biopsy from January 2012 to October 2022. The degree of liver fibrosis （S0-4） was assessed according to the Scheuer scoring system， with ≥S2 defined as significant liver fibrosis， ≥S3 defined as progressive liver fibrosis， and S4 defined as liver cirrhosis. The independent samples t-test was used for comparison of normally distributed continuous data between two groups， and a one-way analysis of variance was used for comparison between multiple groups； the Mann-Whitney U test was used for comparison of non-normally distributed continuous data between two groups， and the Kruskal-Wallis H test was used for comparison between multiple groups； the chi-square test or the Fisher’s exact test was used for comparison of categorical data between groups. The area under the ROC curve （AUC） was used to evaluate the efficacy of complement C3 in the diagnosis of liver fibrosis in patients with PBC. The Spearman correlation analysis was used to investigate the correlation between complement C3 and liver fibrosis stage. ResultsAmong the 108 patients with PBC， there were 87 （80.6%） female patients and 102 patients （94.4%） with positive autoantibody. As for the stage of liver fibrosis， there were 5 patients （4.6%） in S0 stage， 41 （38.0%） in S1 stage， 23 （21.3%） in S2 stage， 25 （23.1%） in S3 stage， and 14 （13.0%） in S4 stage. There was a significant difference in the level of complement C3 between the patients with different liver fibrosis stages （H=42.891， P<0.001）. The level of complement C3 gradually decreased with the aggravation of liver fibrosis， with a negative correlation between them （r=-0.565， P<0.001）. Liver stiffness measurement （LSM）， aspartate aminotransferase/alanine aminotransferase ratio， aspartate aminotransferase-to-platelet ratio index， and fibrosis-4 were negatively correlated with complement C3， with a correlation coefficient of -0.439 （P<0.001）， -0.323 （P=0.001）， -0.206 （P=0.033）， and -0.291 （P=0.002）， respectively. The multivariate logistic regression analysis showed that complement C3 level was an independent predictive factor for significant liver fibrosis， progressive liver fibrosis， and liver cirrhosis， while LSM was an independent predictive factor for significant liver fibrosis and progressive liver fibrosis. The ROC curve analysis showed that complement C3 had an AUC of 0.731， 0.832， and 0.968， respectively， in the diagnosis of significant liver fibrosis， progressive liver fibrosis， and liver cirrhosis， with a corresponding cut-off value of 1.445， 1.235， and 1.005， respectively， and complement C3 combined with LSM had an AUC of 0.811， 0.941， and 0.976， respectively， in the diagnosis of significant liver fibrosis， progressive liver fibrosis， and liver cirrhosis. There was a significant difference in AUC between complement C3 combined with LSM and complement C3 alone in the diagnosis of significant liver fibrosis （Z=2.604， P=0.009）， and there was also a significant difference in AUC between complement C3 combined with LSM and complement C3 alone in the diagnosis of progressive liver fibrosis （Z=3.033， P=0.002）； there was no significant difference in AUC between complement C3 combined with LSM and complement C3 alone in the diagnosis of liver cirrhosis （Z=1.050， P=0.294）， while There was a significant difference in AUC between complement C3 combined with LSM and LSM alone in the diagnosis of liver cirrhosis （Z=2.326， P=0.020）. ConclusionSerum complement C3 level has a certain clinical value in assessing the degree of liver fibrosis in patients with PBC， and complement C3 combined with LSM can further improve the efficacy of complement C3 or LSM in the diagnosis of liver fibrosis in PBC.
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Objective:To observe the clinical effects of pars plana vitrectomy (PPV) combined with internal limiting membrane (ILM) peeling and C 3F 8 tamponade for patients with highly myopic macular hole (HM-MH) with and without foveoschisis. Methods:A retrospective case controlled study. From January 2017 to February 2022, 23 eyes of 23 patients with highly myopic macular hole with and without foveoschisis diagnosed in the Shandong Eye Hospital were included in the study. Among them, 5 males had 5 eyes, and 18 females had 18 eyes, the age was (54.43±12.96) years old. The patients with or without foveoschisis were 12 eyes in 12 cases and 11 eyes in 11 cases. Studies were divided into two groups, depending on the presence of a concomitant myopic foveoschisis or not. The groups are high myopia macular hole with foveoschisis (group A) and high myopia macular hole without foveoschisis (group B). Best-corrected visual acuity (BCVA), B-scan ultrasonography, optical coherence tomography and axial length (AL) measurement were performed in all eyes. Snellen chart was used for BCVA examination, and the visual acuity was converted into logarithm of minimum angle of resolution (logMAR) during statistics. The age of the two groups, sex, macular hole (MH) diameter, logMAR BCVA, AL, posterior scleral staphyloma, there was no significant difference ( P>0.05). PPV combined with ILM peeling and C 3F 8 filling were performed in all eyes. Follow-up was at least 3 months after the last operation. BCVA changes and MH closure were compared between the two groups after surgery. Wilcoxon test was used to compare BCVA before and after operation. Mann-whiteny U test was used to compare preoperative and postoperative BCVA between groups. Results:After initial surgery, MH was closed in 17 of 23 eyes (74%, 17/23). MH was closed in 8 eyes in group A (66.7%, 8/12). Four eyes were not closed (33.3%, 4/12); MH closed in 9 eyes in group B (81.8%, 9/11). There was no significant difference between the two groups after initial operation ( P>0.05). At 1 and 3 months after surgery, the logMAR BCVA of patients in group A and group B were 1.00±0.46, 1.03±0.83 and 0.53±0.63, 0.55±0.41, respectively. Compared with before operation, there was no significant difference at 1 month ( P=0.783, 0.358), but the difference was statistically significant at 3 months ( P=0.012, 0.007). There was no significant difference in logMAR BCVA between group A and group B at 1 and 3 months after operation ( P=0.687, 0.950). Conclusion:PPV combined with ILM peeling and C 3F 8 tamponade can promote MH closure and improve visual acuity in most affected eyes with HM-MH with and without foveoschisis.
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Objective:To explore the relationship between the levels of serum complement C3 and C4 and the degree of renal pathological injury in patients with IgA nephropathy (IgAN).Methods:It was a retrospective study. The clinical and pathological data of patients with primary IgAN diagnosed by renal biopsy in the Department of Nephrology of the Second People's Hospital of Qujing City, Yunnan Province from December 1, 2019 to December 31, 2022 were collected. According to the IgAN Oxford classification criteria, the patients were divided into mild renal pathological injury group (mild group, <3 pathologic types) and severe renal pathological injury group (severe group, ≥3 pathological types). The levels of serum C3 and C4 and other clinical data were compared between the two groups. Spearman correlation method was used to analyze the correlation between serum C3, C4 levels and estimated glomerular filtration rate (eGFR) during renal biopsy.Multivariate logistic regression model was used to analyze the influencing factors of the pathological injury degree in IgAN patients and the forest map depicted the effect of risk factors.Results:A total of 164 IgAN patients were included in the study, including 77 males (47.0%), aged (35.5±12.9) years old. There were 60 patients in the mild group and 104 patients in the severe group. Compared with the mild group, the patients in the severe group were older, had higher levels of serum C4, serum uric acid, low density lipoprotein cholesterol and 24 h urinary protein, higher proportions of hypertension, glucocorticoids/immunosuppressant therapy, C3 deposition in renal tissues and microscopic hematuria, and had lower hemoglobin and serum C3 level (all P<0.05). The results of Spearman correlation analysis showed that the level of serum C3 was positively correlated with eGFR ( r=0.303, P<0.001), and the level of serum C4 was negatively correlated with eGFR ( r=-0.238, P=0.002). Multivariate logistic regression analysis results showed that serum C3 (every 0.01 g/L increase, OR=0.976, 95% CI 0.957-0.996, P=0.018), serum C4 (every 0.01 g/L increase, OR=1.091, 95% CI 1.020-1.166, P=0.011), hemoglobin ( OR=0.969, 95% CI 0.950-0.988, P=0.002), and serum uric acid ( OR=1.005, 95% CI 1.001-1.009, P=0.012) were independent related factors of renal pathological damage (severe injury /mild injury) in IgAN patients. Conclusions:Serum C3 and C4 are independent related factors of the severity of renal pathological injury in IgAN patients.
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Objective:To explore the expression of type 2 complement receptor (CR2) in mesangial cells of the renal tissue in IgA nephropathy (IgAN) and its possible mechanism involved in complement C3 deposition.Methods:The demographic data, samples of plasma and renal tissues of primary IgAN patients diagnosed by renal biopsy in the Guangdong Provincial People's Hospital from August 2021 to May 2022 were collected. According to the fluorescent intensity of mesangial complement C3 deposition, the patients were divided into complement C3 deposition ≥2+ group and complement C3 deposition <2+ group. The circulating IgA and complement C3 levels were detected by enzyme linked immunosorbent assay (ELISA). The influencing factors of kidney prognosis, plasma IgA and complement C3 levels were compared between the two groups. Immunofluorescence was used to detect the expression of IgA, complement C3 and CR2 in the renal mesangial cells of IgAN patients and normal renal tissues around renal carcinoma. Human mesangial cells were cultured in vitro and randomly divided into control group and experimental group. The experimental group was incubated with IgA protein (2 g/L) for 8 hours. The expressions of CR2 protein and mRNA were measured by Western blotting and real-time fluorescence quantitative PCR. The biological function of differential genes was analyzed by gene ontology (GO) and Kyto encyclopedia of genes and genomes (KEGG) enrichment analysis. Results:A total of 75 patients with IgAN were included in this study, including 50 patients in the complement C3 deposition ≥2+ group and 25 patients in the complement C3 deposition <2+ group. The proportions of patients with urine red blood cell count negative, 1+, 2+ and 3+-4+ in the complement C3 deposition ≥2+ group were 2.0%, 8.0%, 18.0% 72.0%, respectively, which were more serious than those in the complement C3 deposition <2+ group (4.0%, 4.0%, 52.0%, 40.0%) ( Z=-2.320, P=0.020). Meanwhile, the proportion of S1 in Oxford pathological classification in the complement C3 deposition ≥2+ group was higher than that in the complement C3 deposition <2+ group (68.0% vs. 40.0%, χ2=5.389, P=0.020), and there were no statistically significant differences in gender, age, 24-hour urinary protein, serum creatinine, other indicators of Oxford pathological classification between the two groups. ELISA results showed that plasma IgA concentration in the complement C3 deposition ≥2+ group was higher than that in the complement C3 deposition <2+ group [3.62 (2.95, 5.53) g/L vs. 2.72 (2.15, 4.24) g/L, Z=2.405, P=0.016], and the plasma complement C3 concentration was lower than that in the complement C3 deposition <2+ group [199.6 (116.0, 328.0) mg/L vs. 319.2 (158.3, 454.5) mg/L, Z=-2.383, P=0.017]. Spearman correlation analysis showed that the complement C3 deposition intensity was positively correlated with IgA deposition intensity in mesangial area ( rs=0.441, P<0.001). Immunofluorescence results showed that there was colocalization of IgA and complement C3 in the glomeruli of IgAN patients. The expression of CR2 in the kidney was consistent with complement C3 deposition, and CR2 was colocalization with complement C3. In vitro experiments, the expression of CR2 in IgA protein group was higher than that in the control group ( P<0.05). GO and KEGG enrichment analysis found that IgA protein induced active changes in various pathways of mesangial cells. Conclusion:IgA protein induces mesangial cells to express CR2 and participates in complement C3 deposition, which may be an important mechanism of complement C3 activation in IgAN.
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The study was a retrospective study. The clinical data of 866 patients with IgA nephropathy (IgAN) in Beijing Anzhen Hospital, Capital Medical University from March 2010 to March 2021 were analyzed, to investigate the clinical pathology and renal prognosis of IgAN patients with intrarenal arteriolosclerosis, and to preliminarily explore whether abnormal activation of complement system is involved in the injury of arteriolosclerosis. The patients were divided into renal arteriolar lesions group and non-renal arteriolar lesions group according to the renal histopathology, and the differences of clinical pathological manifestations, prognosis between the two groups were compared. The results showed that, compared with the non-renal arteriolar lesions group ( n=236), IgAN patients in the renal arteriolar lesions group ( n=630) had higher proportions of hypertension and malignant hypertension, higher levels of urinary albumin-creatinine ratio, 24-hour urine protein quantification and serum uric acid, lower estimated glomerular filtration rate, and more severe MEST-C lesions of the Oxford classification (all P < 0.05). Cox regression analysis results showed that intrarenal arteriolosclerosis was the independent risk factor affecting the progression of IgAN to ESRD ( HR=6.437, 95% CI 2.013-20.585, P=0.002). Renal histopathology showed that the deposition of complement C3c on the wall of intrarenal arterioles in the renal arteriolar lesions group ( n=98) was stronger than that in non-renal arteriolar lesions group ( n=18, P < 0.05). IgAN patients with renal arteriolosclerosis present with serious clinical and pathological manifestations, and renal prognosis. Abnormal activation of complement system may be involved in the pathogenesis of intrarenal arteriolosclerosis.
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Objective:To investigate the clinicopathological characteristics and prognosis of idiopathic membranous nephropathy (IMN) with or without C3 deposition.Methods:Clinical and pathological data of 576 patients with IMN diagnosed in Affiliated Hospital of Qingdao University from January 2017 to January 2021 were retrospectively analyzed. The patients were divided into C3 deposition group and non-C3 deposition group according to the immunofluorescence staining of C3. The clinical and pathological characteristics were compared between the two groups. Kaplan-Meier survival curve was used to compare the prognosis of the two groups.Results:A total of 576 IMN patients (male 364 (63.20%)) were enrolled, including 400 patients (69.44%) with C3 deposition and 176 patients (30.56%) without C3 deposition. Compared with the non-C3 deposition group, the levels of total blood cholesterol ( t=0.94, P=0.002) and the proportion of phospholipase A2 receptor ( χ2=9.99, P=0.002), IgG ( χ2=10.67, P=0.001), IgM ( χ2=7.00, P=0.008), IgA ( χ2=7.87, P=0.005) and C1q ( χ2=8.28, P=0.004) depositions in renal tissues was higher in C3 deposition group, while the levels of serum C3 ( t=2.87, P=0.004), albumin ( t=3.57, P<0.001) and IgG ( Z=3.55, P<0.001) were lower in C3 deposition group. There were no significant differences in other clinicopathological indicators between the two groups. The survival analysis was performed in 460 patients who were followed for>6 months, including 319 cases (69.35%) of C3 deposition and 141 cases (30.65%) of non-C3 deposition. The end point event was defined as an eGFR decline>30% or entry into end stage renal disease (ESRD). There was no statistically significant difference in treatment method between the two groups ( P>0.05). The median follow-up time was 22 (13,32) months, 327 (71.09%) patients achieved remission, and 22 patients had renal end-point events. Compared with the non-C3 deposition group, the proportion of urinary protein remission was lower ( χ2=10.85, P<0.05), the incidence of renal end-point events was higher ( χ2=5.05, P<0.05). Kaplan-Meier survival analysis showed that patients with C3 deposition had a lower cumulative remission rate (Log-rank χ2=6.68, P=0.010), and a lower cumulative renal survival than those without C3 deposition had ( χ2=5.42, P=0.020). Conclusions:Compared with patients without C3 deposition, IMN patients with C3 deposition have more severe clinical and pathological changes, lower renal cumulative remission rate, and are more likely to have poor prognosis.
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OBJECTIVE To investigate the role of the complement C3/C3aR signaling pathway in the prefrontal cortex and colon neuroglia cell interactions during meth-amphetamine(METH)addiction,to observe the effects of TLR4 inhibitors as well as complement C3 elimination on METH reward and relapse behavior,and to explore the neuroinflammatory mechanisms of complement C3 acti-vation in METH addiction.METHODS ①A 14 d and 28 d rat METH addiction model was established to observe the effects of TLR4 antagonist ibudilast 3 mg·kg-1 and 10 mg·kg-1 on self-administration,reward motivation,relapse,and natural reward behavior in METH-trained 14 d rats and the effects of 0.02 mg·kg-1 complement C3 antago-nist on self-administration behavior in METH-trained 28 d rats.② Differences in the expression of TLR4,NF-κB,GRP94,C3,cathepsin L,CD68,and GFAP in the pre-frontal cortex of each group were examined using West-ern blotting.③ In addition,the expression of ATF6 in the prefrontal cortex of each group and the effects on neuro-nal and microglia/macrophage INOS,CD206 GRP94,and complement C3/C3aR.RESULTS ① Endoplasmic reticulum stress occurred in neurons and microglia after METH exposure depending on GRP94 and unfolded pro-tein responses to the ATF6 pathway.In addition,it acti-vates the TLR4-NF-κB pathway.② Microglia with high complement C3/C3aR expression in the prefrontal cortex were recruited to synaptic pruning and phagocytic responses around neurons with high GRP94,comple-ment C3/C3aR expression and these effects were blocked by complement C3 antagonists.③ In the rec-tum,GRP94 functions as a molecular chaperone for com-plement C3 and cathepsin L.Crosstalk occurs between enteric neurons high in GRP94,complement C3,and macrophages high in C3aR,located in the submucosa,lamina propria,and muscular,respectively,and all of these effects are blocked by complement C3 antago-nists.④ Treatment with the TLR4 antagonist ibudilast inhibits self-administration,reward motivation,and cue-or METH-priming in METH-trained 14 d rats,but fails to affect natural reward behavior.Ibudilast treatment attenu-ates the TLR4-NF-κB inflammatory pathway and comple-ments C3/C3aR pathway in the prefrontal cortex.CON-CLUSION Activation of the complement C3/C3aR signal-ing pathway by TLR4-NF-κB inflammatory signaling in the prefrontal cortex mediates the METH addiction pro-cess,providing an experimental basis for the clinical treatment of METH addiction,and targeting TLR4/NF-κB inflammatory signaling and complement C3/C3aR may be a new way to intervene in METH addiction.
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Objective:To determine the expression of circular RNA-SEC31A(circSEC31A) in pancreatic cancer and investigate the effects on the invasion and migration of pancreatic cancer cells and the underlying molecular mechanism.Methods:Differentially expressed circRNAs between pancreatic cancer cells (BXPC-3, PANC1, CaPan-2, SW1990) and human normal pancreatic cells (HPDE) were identified by qRT-PCR. Then, two cell lines with high circSEC31A expression were selected to conduct next experiments. According to the sequence of the back-splicing site in circSEC31A, siRNAs for downregulation of circSEC31A were designed and transfected by liposome to silence circSEC31A in pancreatic cancer cells, and grouped as followed siR-circSEC31A#1 and siR-circSEC31A#2. Meanwhile, siR-NC group transfected with non-specific siRNA served as control. Transwell assays and wound healing assays were operated to evaluate the functional role of circSEC31A on the invasion and migration of pancreatic cancer cells. RNA Pull-down assay with circSEC31A probe and oligo control probe was used to screen the miRNA combining with circSEC31A and the effects of miRNA on cell invasion and migration of pancreatic cancer cells were validated. The effect of miR-200c-3p and circSEC31A silencing on the expression of PDK1 mRNA was identified by qRT-PCR. The protein expression of PDK1, downstream Akt and p-Akt after circSEC31A silencing was verified by Western blotting assays.Results:The relative expression level of circSEC31A in HPDE (1.000±0.120) was obviously lower than that in BXPC-3 (1.920±0.130), SW1990 (2.93±0.528), PANC1 (4.557±0.692) and CaPan-2 (5.247±0.194), and all the differences were statistically significant ( P<0.001). Compared with the PANC1 siR-NC group (1301.3±94.6) and CaPan-2 siR-NC group (1835.0±70.1) per 100 high power field, transwell assays showed that the numbers of invasive pancreatic cancer cells was highly decreased in PANC1 siR-circSEC31A#1 group (727.3±92.9), siR-circSEC31A#2 group (792.0±18.1), CaPan-2 siR-circSEC31A#1 group (718.0±90.6), siR-circSEC31A#2 group (692.7±84.8). Wound healing assays showed that silencing circSEC31A decreased the wound healing rate of pancreatic cancer cells in PANC1 siR-circSEC31A#1 group (20.667±3.215)%, siR-circSEC31A#2 group (20.000±4.583)%, CaPan-2 siR-circSEC31A#1 group (28.000±8.185)%, siR-circSEC31A#2 group (29.667±5.686)%, compared with the PANC1 siR-NC group (55.000±4.359)% and CaPan-2 siR-NC group (69.000±3.606)%. RNA Pull-down assays showed that compared with PANC1 oligo probe group (1.000±0.091) and CaPan-2 oligo probe group (1.000±0.153), miR-200c-3p was significantly enriched in the PANC1 circSEC31A probe group (2.237±0.175) and CaPan-2 circSEC31A probe group (2.166±0.156). Compared with PANC1 siR-NC group (939.3±57.0) and CaPan-2 siR-NC group (786.7±51.5) per 100 high power field, the numbers of invasive pancreatic cancer cells were up-regulated in PANC1 siR-miR-200c-3p group (1206.0±99.1) and CaPan-2 siR-miR-200c-3p group (1838.0±105.7), while the low numbers of invasive pancreatic cancer cells were observed in PANC1 siR-miR-200c-3p+ siR-circSEC31A group (932.7±116.4) and CaPan-2 siR-miR-200c-3p+ siR-circSEC31A group (785.3±58.8). Compared with PANC1 siR-NC group (1.000±0.103) and CaPan-2 siR-NC group (1.000±0.107), the relative expression of PDK1 mRNA in PANC1 siR-miR-200c-3p group (1.898±0.159) and CaPan-2 siR-miR-200c-3p group (2.102±0.337) was upregulated. Furthermore, the expression of PDK1 mRNA was decreased in the siR-miR-200c-3p+ siR-circSEC31A group (0.980±0.070, 1.015±0.079). Western blot assays showed that the expression of PDK1 protein in PANC1 siR-NC group, siR-circSEC31A#1 group, siR-circSEC31A#2 group was 0.767±0.086, 0.281±0.191, 0.333±0.062 and in CaPan-2 siR-NC group, siR-circSEC31A#1 group, siR-circSEC31A#2 group was 0.712±0.038, 0.353±0.061, 0.308±0.018. The expression of p-Akt protein in PANC1 siR-NC group and siR-circSEC31A group was 0.741±0.050, 0.114±0.027, 0.139±0.041. In addition, p-Akt protein expression in CaPan-2 siR-NC group and siR-circSEC31A group was 0.823±0.052, 0.141±0.045, 0.280±0.089. PDK1 and p Akt expression in siR circSEC31A group was obviously lower than those in sir NC group. All the differences between either groups above were statistically significant ( P<0.05). Conclusions:circSEC31A is upregulated in pancreatic cancer cells, which facilitates the invasion and metastasis of pancreatic cancer cells via miR-200c-3p/PDK1/Akt signaling pathway, supporting that circSEC31A may function as a new diagnostic and therapeutic target for pancreatic cancer patients.
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El síndrome de Alport es una entidad hereditaria que se produce principalmente por una mutación en los genes que codifican el colágeno tipo IV. Por otro lado, la glomerulonefritis C3 es una entidad rara que presenta un patrón de glomerulonefritis membranoproliferativa y su etiología se basa en un control anormal de la activación de la vía alternativa del complemento. A continuación se describe un caso de un paciente, de sexo masculino, que cursa con un síndrome nefrótico corticorresistente en el que se documenta un patrón de glomerulonefritis membranoproliferativa en la biopsia renal con depósitos de C3 en la inmunofluorescencia, asociada a una deleción heterocigota en el gen CFHR1 en el estudio genético de las proteínas reguladoras del complemento. Además, en el panel genético realizado por corticorresistencia se encuentra una variante COL4A5 asociada al síndrome de Alport ligado al X. Estas entidades pueden presentarse con un curso clínico diverso, pero al estar asociadas pueden acelerar la progresión a enfermedad renal crónica, por lo que se hace necesario hacer un seguimiento clínico más estricto.
Alport Syndrome is a hereditary entity that occurs mainly due to a mutation in the genes that encode type IV collagen. C3 glomerulonephritis is a rare entity with a pattern of membranoproliferative glomerulonephritis and its etiology is based on abnormal control of the activation of the alternative complement pathway. We describe a case of a male patient who presents with a corticosteroid nephrotic syndrome in which a pattern of membranoproliferative glomerulonephritis is documented in the renal biopsy with C3 deposits in the immunofluorescence, associated with a heterozygous deletion in the gene CFHR1 in the genetic study of complement regulatory proteins. Furthermore, a variant COL4A5 associated with X-linked Alport syndrome is found in the genetic panel for corticosteroid resistance. These entities can present with a diverse clinical course, but when associated they can accelerate progression to chronic kidney disease, which is why makes it necessary to do a more strict clinical follow-up.
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La enfermedad glomerular comprende un grupo heterogéneo de entidades que se caracterizan por la pérdida de la arquitectura o función del glomérulo secundario a proceso inflamatorio del mismo de etiología autoinmune, infecciosa, paraneoplásica, que puede ser identificada con estudios de histopatología. Su reconocimiento durante la gestación representa un reto diagnóstico por la sobreposición de cambios fisiológicos, el debut de enfermedades autoinmunitarias o de enfermedades genéticas, entre otros. La presentación clínica suele encajar en grupos sindromáticos específicos, sin embargo, es frecuente que sean clínicamente indistinguibles o sobrepuestos. El debut de la enfermedad renal con curso clínico de rápida instauración y de evolución desfavorable con respecto a la función renal, hace mandatorio un estudio completo desde el abordaje clínico hasta la interpretación de los hallazgos histopatológicos, encaminado en la distinción de causas primarias y secundarias. Si bien las glomerulonefritis primarias no son las más frecuentes en la gestación, la identificación certera del diagnóstico y su adecuada clasificación permite el manejo dirigido y óptimo de las mismas. Se presentan los casos clínicos de dos gestantes con enfermedad glomerular primaria, con discrepancia en su diagnóstico, enfatizando en sus manifestaciones durante el curso de la gestación, el algoritmo diagnóstico utilizado, el tratamiento inicial y de mantenimiento utilizado. Se resalta la utilidad de la biopsia renal, específicamente la inmunofluorencia para aclarar el mismo.
Glomerular disease involves a heterogeneous group of entities that are characterized by loss of the architecture and function of the glomerulus and this can be caused by immunity, infectious and paraneoplastic etiologies. The aforementioned can be identified in histopathological studies. The recognition of this entity during pregnancy represents a diagnostic challenge due to the superposition of physiological changes, the development of autoimmune diseases and / or genetic disease, among others. Clinical manifestations can be into specific syndromic groups; however we can find indistinguishable manifestations and overlapping of this. When the disease is present its common to find rapidly establishment and unfavorable evolution about renal function. With this it's necessary to complete studies involving the initial clinical approach until histopathological findings with the goal to find primary and secondary causes. As it's known primary glomerulonephritis is not the most frequent in pregnancy, the accuracy in the diagnosis and the proper classification allows the direct and soon management. In this case report we describe 2 pregnant women with primary glomerular disease with discrepancy in their diagnosis. We talk about manifestations during pregnancy, the algorithm used in the diagnosis and finally the initial treatment and the maintenance used in these patients.
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Abstract Thymoquinone (TQ) has shown hepatoprotective effects in various experimental studies. We aimed to investigate the possible beneficial effects of TQ regarding its prevention of alpha-amanitin induced hepatotoxicity in human C3A hepatocytes. After administering alpha-amanitin in a concentrations of 1 and 10µg/mL on the cells in a hepatocyte cell line, TQ was administered in various concentrations (10, 5, 1, 0.5, 0.1, 0.05, 0.01, 0.005 µg/mL). The MTT test was used to determine cell viability. For the groups given only TQ at various concentrations, the cell viability rates at 48 hours post-administration were found at 82.6, 98.3, 102.1, 102.5, 99.4, 99.4, 101.9 and 106.3%, respectively. For the group with 1μg/mL alpha-amanitin and various TQ concentrations, the cell viability rates were found at 74.6, 88.5, 87.4, 88.7, 85.7, 86.8, 88.4, and 92.9%, respectively. For the group with 10μg/mL alpha-amanitin and various TQ concentrations, the cell viability rates for each TQ subgroup were found at 65.2, 79.2, 81.4, 81.1, 81.8, 81.8, 82.2 and 91.9%, respectively. Our study is the first in vitro study that investigates TQ's effects on alpha-amanitin induced hepatotoxicity. Although TQ had beneficial effect in low doses did not significantly increase cell viability in liver damage due to alpha-amanitin toxicity.
Subject(s)
Cell Line/classification , In Vitro Techniques/methods , Alpha-Amanitin/administration & dosage , Liver/physiopathologyABSTRACT
Objective: To investigate the expression of programmed death ligand-1 (PD-L1, SP142) and PD-L1 (22C3) in triple-negative breast cancer (TNBC), and analyze their correlation with the clinicopathological factors and prognosis. Methods: The clinicopathologic data of 259 patients with TNBC treated in Cancer Hospital from August 2010 to December 2013 were collected. Whole section of surgical tissue samples were collected to conduct PD-L1 (SP142) and PD-L1 (22C3) immunohistochemical (IHC) staining. The PD-L1 expression in tumor cells and tumor infiltrating immune cells were visually assessed respectively, the relationship between PD-L1 expression and clinicopathologic characterizes were analyzed. Univariable and multivariable Cox proportional hazards regression models were used to test the correlations between PD-L1 expression and disease-free survival (DFS) and overall survival (OS). Results: The positive rates of SP142 (immune cell score, ICs≥1%) and 22C3 (combined positive score, CPS≥1) were 42.1%(109/259) and 41.3%(107/259) in TNBC tissues, respectively, with a total coincidence rate of 82.3%. The Kappa value of positive expression cases was 0.571 and the distribution difference of SP142 and 22C3 positive expression cases was statistically significant (P<0.001). The PD-L1 positive patients were less likely to have vascular invasion (P<0.05), but with higher histological grade and Ki-67 proliferation index (P<0.05). The recurrence/metastasis cases(8) of the patients with positive PD-L1 (SP142) was significantly lower than that of patients with negative PD-L1(SP142, 27, P=0.016). The positive expression of PD-L1 (SP142) patients were longer DFS (P=0.019). The OS of patients with positive PD-L1 (SP142) were longer than those with negative PD-L1 (SP142), but without significance (P=0.116). The positive expression of PD-L1 (22C3) was marginally associated with DFS and OS of patients (P>0.05). Conclusions: The expression of PD-L1 (22C3) is different from that of PD-L1 (SP142) in TNBC, and the two antibodies can't be interchangeable for each other in clinical tests. PD-L1 (SP142) status is an independent prognostic factor of DFS in TNBC. The DFS is significantly prolonged in patients with positive expression of PD-L1 (SP142).
Subject(s)
Humans , B7-H1 Antigen/genetics , Immunohistochemistry , Prognosis , Triple Negative Breast Neoplasms/pathologyABSTRACT
Objective To validate whether the expression of human cluster of differentiation 55 (hCD55) protein in porcine islet cells could inhibit the activation of complement components in human serum. Methods Four adult pigs with WT (wild type), GTKO [α-1, 3-galactosyltransferase (GGTA1) knockout], GTKO/hCD55 and hCD55 genotypes were selected. Islet cells were isolated from WT, GTKO and GTKO/hCD55 pigs, and the purity and insulin secretion function were detected. The expression of hCD55 at the DNA, RNA and protein levels was analyzed by agarose gel electrophoresis, reverse transcription polymerase chain reaction (RT-PCR) and flow cytometry, respectively. Complement-dependent cytotoxicity assay and complement deposition assay were performed under the incubation conditions with fresh human serum. Results The purity of isolated porcine islet cells from three genotype pigs was > 75%, and the glycemic index was > 1. The expression of hCD55 messenger RNA(mRNA) and protein in GTKO/hCD55 porcine islet cells decreased the deposition of human complement component C3c and membrane-attacking complex C5b-9, and reduced the cytotoxicity. Conclusions The expression of hCD55 protein in porcine islet cells could inhibit the activation of human complement and reduce complement-mediated killing effect, indicating that hCD55 protein could exert complement protection effect on porcine islet cells. These findings provide theoretical basis for the application of hCD55 in islet xenotransplantation.