Objective:To explore the application and clinical significance of pathological diagnostic criteria for medically refractory epilepsy in children.Methods:Cross-sectional study.A retrospective analysis was conducted on 490 children(pathology involved) with medically refractory epilepsy treated continuously in the Pediatric Epilepsy Center of Peking University First Hospital from January 2019 to May 2022.The distribution of different pathological types was observed, and the differences in clinical characteristics among different pathological types were analyzed through Kruskal-Wallis or χ2 tests.The impact of clinical and pathological features on patient prognosis was evaluated through regression analysis. Results:Focal cortical dysplasia (FCD) was the predominant lesion (49.59%, 243/490).The electroencephalograms ( χ2=6.720, P=0.035) and clinical seizure characteristics ( χ2=26.370, P<0.001) in FCDⅡ were more focal than those in FCD Ⅰ and Ⅲ.Moreover, the proportions of focal resection in surgery ( χ2=24.286, P<0.001) and central involvement ( χ2=22.849, P<0.001) in FCDⅡ were higher than those in FCD Ⅰ and Ⅲ.Univariate and multivariate regression analyses revealed that FCD Ⅱ had a better prognosis than other dysplastic patients among the 375 cases of dysplasia ( P=0.049).Next-generation sequencing was performed on 35 cases of cortical malformations with such morphological characteristics as increased numbers of neurons in the white matter and Olig2-positive glial cell hyperplasia, and SLC35A2 mutations were detected in 2 cases (5.71%). Conclusions:Pathology of refractory epilepsy is specialized and continuously evolving.Standardized specimen processing and the accumulation of morphological, immunohistochemical, and molecular genetic data provide the foundation for clarifying the neuropathological nature of epilepsy, improving integrated classification, and advancing prognosis prediction and targeted therapy.

Objective:To evaluate the effect of cathepsin B(CTSB)/NOD-like receptor pyrin domain containing 3(NLRP3)pathway on the polarization of macrophages induced by LPS.Methods:The well-growing RAW264.7 mouse mononuclear macrophage lines were cultured in vitro and divided into 3 groups(n=6)according to the random number table method:control group(C group),LPS group(L group)and LPS+CA074-me(CTSB inhibitors)group(B group).C group was cultured normally for 24 h,L group was cultured with LPS concentration of 1 μg/ml medium for 24 h.B group was pretreated with CTSB inhibitor CA074-me 30 μmol/L for 1 h before LPS induction,and co-cultured with LPS concentration of 1 μg/ml medium for 24 h.After 24 hours,the morphological changes of the cells were observed by microscope,the concentrations of IL-1β and IL-18 in the supernatant were determined by ELISA.The ex-pressions of cathepsin B precursor(pro-CTSB),mature cathepsin B(mature-CTSB),NLRP3,apoptosis-related speck protein(ASC)and apoptosis-related speck protein-1(caspase-1)were detected by Western blot.The mRNA expression levels of CD32,inducible ni-tric oxide synthase(iNOS),arginase 1(Arg-1)and CD206 were detected by qRT-PCR.The positive expression rates of M1 macro-phage surface marker CD86 and M2 macrophage surface marker CD206 were detected by flow cytometry.Results:Compared with group C,the morphology of cells in groups L and B became larger and pseudopodia appeared.The concentrations of IL-1β and IL-18 in cell supernatant were increased,the expressions of pro-CTSB,mature-CTSB,NLRP3,ASC and caspase-1 were increased,and the expressions of CD32,iNOS mRNA were up-regulated and the positive rates of CD86 and CD206 were increased(P<0.01).Arg-1 and CD206 mRNA in group B were up-regulated(P<0.01).Compared with group L,the pseudopodia of group B were reduced,and the morphology was closer to group C.The concentration of IL-1β and IL-18 in the supernatant,the expression of mature-CTSB,NLRP3,ASC and caspase-1,CD32 and iNOS mRNA and the positive rate of CD86 were down-regulated in group B.The expression of pro-CTSB,Arg-1 and CD206 mRNA and the positive rate of CD206 were increased(P<0.01).Conclusion:Inhibition of CTSB/NLRP3 pathway can reduce the inflammatory response,reduce the LPS-induced polarization of RAW264.7 cells to M1 macrophages,and pro-mote their polarization to M2 macrophages.

Objective:To evaluate the effect of cathepsin B(CTSB)/NOD-like receptor pyrin domain containing 3(NLRP3)pathway on the polarization of macrophages induced by LPS.Methods:The well-growing RAW264.7 mouse mononuclear macrophage lines were cultured in vitro and divided into 3 groups(n=6)according to the random number table method:control group(C group),LPS group(L group)and LPS+CA074-me(CTSB inhibitors)group(B group).C group was cultured normally for 24 h,L group was cultured with LPS concentration of 1 μg/ml medium for 24 h.B group was pretreated with CTSB inhibitor CA074-me 30 μmol/L for 1 h before LPS induction,and co-cultured with LPS concentration of 1 μg/ml medium for 24 h.After 24 hours,the morphological changes of the cells were observed by microscope,the concentrations of IL-1β and IL-18 in the supernatant were determined by ELISA.The ex-pressions of cathepsin B precursor(pro-CTSB),mature cathepsin B(mature-CTSB),NLRP3,apoptosis-related speck protein(ASC)and apoptosis-related speck protein-1(caspase-1)were detected by Western blot.The mRNA expression levels of CD32,inducible ni-tric oxide synthase(iNOS),arginase 1(Arg-1)and CD206 were detected by qRT-PCR.The positive expression rates of M1 macro-phage surface marker CD86 and M2 macrophage surface marker CD206 were detected by flow cytometry.Results:Compared with group C,the morphology of cells in groups L and B became larger and pseudopodia appeared.The concentrations of IL-1β and IL-18 in cell supernatant were increased,the expressions of pro-CTSB,mature-CTSB,NLRP3,ASC and caspase-1 were increased,and the expressions of CD32,iNOS mRNA were up-regulated and the positive rates of CD86 and CD206 were increased(P<0.01).Arg-1 and CD206 mRNA in group B were up-regulated(P<0.01).Compared with group L,the pseudopodia of group B were reduced,and the morphology was closer to group C.The concentration of IL-1β and IL-18 in the supernatant,the expression of mature-CTSB,NLRP3,ASC and caspase-1,CD32 and iNOS mRNA and the positive rate of CD86 were down-regulated in group B.The expression of pro-CTSB,Arg-1 and CD206 mRNA and the positive rate of CD206 were increased(P<0.01).Conclusion:Inhibition of CTSB/NLRP3 pathway can reduce the inflammatory response,reduce the LPS-induced polarization of RAW264.7 cells to M1 macrophages,and pro-mote their polarization to M2 macrophages.

Objective:To explore the application and clinical significance of pathological diagnostic criteria for medically refractory epilepsy in children.Methods:Cross-sectional study.A retrospective analysis was conducted on 490 children(pathology involved) with medically refractory epilepsy treated continuously in the Pediatric Epilepsy Center of Peking University First Hospital from January 2019 to May 2022.The distribution of different pathological types was observed, and the differences in clinical characteristics among different pathological types were analyzed through Kruskal-Wallis or χ2 tests.The impact of clinical and pathological features on patient prognosis was evaluated through regression analysis. Results:Focal cortical dysplasia (FCD) was the predominant lesion (49.59%, 243/490).The electroencephalograms ( χ2=6.720, P=0.035) and clinical seizure characteristics ( χ2=26.370, P<0.001) in FCDⅡ were more focal than those in FCD Ⅰ and Ⅲ.Moreover, the proportions of focal resection in surgery ( χ2=24.286, P<0.001) and central involvement ( χ2=22.849, P<0.001) in FCDⅡ were higher than those in FCD Ⅰ and Ⅲ.Univariate and multivariate regression analyses revealed that FCD Ⅱ had a better prognosis than other dysplastic patients among the 375 cases of dysplasia ( P=0.049).Next-generation sequencing was performed on 35 cases of cortical malformations with such morphological characteristics as increased numbers of neurons in the white matter and Olig2-positive glial cell hyperplasia, and SLC35A2 mutations were detected in 2 cases (5.71%). Conclusions:Pathology of refractory epilepsy is specialized and continuously evolving.Standardized specimen processing and the accumulation of morphological, immunohistochemical, and molecular genetic data provide the foundation for clarifying the neuropathological nature of epilepsy, improving integrated classification, and advancing prognosis prediction and targeted therapy.

Objective:To construct a clinical prediction scale for focal cortical dysplasia (FCD)type Ⅱ in the malformation of cortical development (MCD) disease spectrum in children.Methods:A case-sectional study.From January 2014 to June 2019, patients who underwent surgery at the Pediatric Epilepsy Center of Peking University First Hospital and were pathologically diagnosed with MCD after surgery were enrolled and randomly divided into the training set and the validation set using random numbering.Clinical, electrophysiological, and imaging data of patients in the training set were analyzed.Variables that could predict FCD type Ⅱ were screened out using a Logistic regression model, and a rating scale was constructed.The diagnostic efficiency of the scale was validated in the validation set to determine the optimum cut-off value, and a consistency test was performed.Results:A total of 381 patients were enrolled in the study, with 260 in the training set and 121 in the validation set.Five clinical factors that exhibited a significant correlation with FCD type Ⅱ were identified in the training set through the logistic regression model: (1) age of seizure onset (<24 months); (2) lesion involving the frontal lobe; (3) epileptic spasms; (4) family history of epilepsy; (5) hippocampal atrophy ± signal change.Based on these 5 variables, the FCD type Ⅱ prediction scale was developed and validated in the validation set with an area under the curve of 0.732.The optimum cut-off value for the prediction scale was 1, at which point the Youden index was 0.384.The scale′s positive predictive value was 0.836, and the negative predictive value was 0.500.The diagnostic consistency between the pathological diagnosis and the FCD type Ⅱ prediction scale was acceptable (Kappa value=0.351), and there was no statistically significant difference between the two diagnostic methods ( P value of the McNemar test=0.065). Conclusions:The FCD type Ⅱ prediction scale has clinical practicability.The application of this scale to predict the pathological type of MCD before operation can help doctors choose the appropriate surgical strategy.

Objective:To explore the value of arterial spin labeling (ASL) in detecting epileptogenic zone (EZ) in children with drug-refractory epilepsy (DRE).Methods:From March 2018 to December 2019, 28 children with DRE were collected prospectively in Peking University First Hospital. Structural MRI, ASL sequence, and PET-CT were performed on 28 DRE children. All children underwent surgical treatment. Intraoperative electrocorticogram findings combined with postoperative MRI results were considered the gold standard for locating EZ. A total of 29 EZ were resected in 28 children. Based on the pathological results, the EZ was divided into focal cortical dysplasia (FCD) Ⅰb and Ⅱa group ( n=12), FCD Ⅱ b group ( n=11) and malformation of cortical dysplasia (MCD) group ( n=6). Structural MRI was observed for finding any abnormal changes that could induce epilepsy and was divided into the normal MRI group ( n=13) and the abnormal MRI group ( n=16). The spatial relationship between abnormal areas in the cerebral blood flow (CBF) map and PET images and the gold standard was observed, and the accurate detection rate of EZ was calculated. The region of interest (ROI) on CBF and PET images was drawn. ROIs were defined as EZ, EZ contralateral zone (EZCZ), EZ adjacent zone (EZAZ), EZAZ contralateral zone (EZAZCZ). The CBF and maximum standardized uptake value (SUV max) were measured, and the asymmetry index (AI) value of EZ and EZAZ of CBF and SUV max was calculated respectively. One-way ANOVA was used to compare the difference among 4 regions and 3 pathological types of CBF, SUV max, and AI. The independent sample t-test was used to compare the difference in AI between normal and abnormal MRI groups. Results:In CBF map, the EZ was accurately localized in 89.7% (26/29) of the lesions, in which 24 EZ had decreased perfusion, and 2 EZ had increased perfusion. Among the 24 EZ with decreased perfusion, the CBF of EZ, EZCZ, EZAZ, and EZAZCZ were significantly different( F=8.79, P<0.001). In PET-CT, the EZ was accurately localized in 93.1% (27/29) of the lesions, in which 25 EZ had decreased metabolism, and 2 EZ had increased metabolism. Among the 25 EZ with decreased metabolism, the SUV max of EZ, EZCZ, EZAZ, and EZAZCZ were significantly different ( F=6.40, P=0.001). The AI value of CBF and SUV max of EZ in the abnormal MRI group were larger than those of the normal MRI group, and the difference was statistically significant ( t=3.34, 3.09, P=0.002 , 0.004). There was no statistical difference in the AI values of CBF and SUV max among FCD Ⅰb and Ⅱa group, FCD Ⅱb group and MCD group ( F=2.05, 1.54, P=0.149, 0.234). Conclusions:ASL technology is accurate in detecting EZ. The changes in perfusion and metabolism of normal structural MRI EZ are greater than abnormal structural MRI EZ. There is no obvious difference in CBF and SUVmax changes in different pathological EZ.

Humans ; Asthma/drug therapy* ; Biological Therapy

Background: There is no standard cut-off value of serum IgG4 concentration and serum IgG4/total IgG ratio for the diagnosis of IgG4-related disease (IgG4-RD) or as a marker of treatment responses. We aimed to explore this issue through a retrospective cohort analysis of adults in southwest China. Methods: The diagnostic performance of serum IgG4 concentration and IgG4/IgG ratio for IgG4-RD was evaluated in a retrospective analysis of 177 adults newly diagnosed as having IgG4-RD and 877 adults without IgG4-RD. Dynamic analysis was performed to evaluate the significance of serum IgG4 concentration on IgG4-RD treatment responses. Results: The serum IgG4 concentration differed according to sex. The optimal cut-off values of serum IgG4 concentration and IgG4/IgG ratio for IgG4-RD diagnosis were 1.92 g/L and 0.12 in males and 1.83 g/L and 0.11 in females, respectively. For patients with serum IgG4 concentration >2.01 g/L, the cut-off values in the total population were >3.00 g/L and 0.19, respectively. The median serum IgG4 concentration decreased over time, and the decrease rate increased over time. The serum IgG4 concentration significantly decreased at >1 week post-treatment (P=0.004), and the median decrease rate was close to 50% at >4 weeks post-treatment. Conclusions Serum IgG4 can be a good indicator for IgG4-RD diagnosis; however, different diagnostic cut-off values should be determined according to sex. The decreasing rate is more conducive than the serum IgG4 concentration to monitor treatment efficacy. The IgG4/IgG ratio did not improve the diagnostic efficacy for IgG4-RD.

Objective:To investigate the efficacy and safety of geritinib in the treatment of acute myeloid leukemia (AML) with FLT3 mutation.Methods:The clinical data of 5 AML patients with FLT3 mutation who were diagnosed in the University of Hong Kong-Shenzhen Hospital, Shenzhen People's Hospital, Shenzhen Second People's Hospital, Shenzhen University General Hospital from March 2020 to April 2021 were retrospectively analyzed. Relapsed patients concurrently received two- or three-drug chemotherapy combined with geritinib. Blood routine was checked once a week; liver function and renal function were checked once every 2 weeks during treatment. Bone marrow puncture was performed once every 1 to 3 months to monitor the bone marrow morphology, minimal residual disease (MRD) and FLT3 mutation expression levels. The efficacy, side effects, overall survival of these patients were analyzed after treatment with geritinib.Results:The white blood cell was increased in all the 5 patients at the initial diagnosis. FLT3 mutations analysis showed FLT3-internal tandem duplication (ITD) (3 cases) and FLT-3 tyrosine-kinase domain (TKD) (2 cases). Among 5 patients, 1 patient was relapse-free with maintenance therapy of oral geritinib after hematological stem cell transplantation (HSCT) for 60 days; among other 4 relapsed and refractory patients, 1 female patient after pregnancy relapsed after transplantation and then achieved complete remission followed by the maintenance therapy with geritinib after oral geritinib, 1 16-year-old patient achieved treatment outcome close to the complete remission after treatment with geritinib, 1 patient achieved complete remission after treatment with geritinib, and then underwent haplo-HSCT followed by the maintenance therapy with geritinib and the other 1 relapsed patient achieved complete remission after treatment with geritinib. After transplantation, 3 patients receiving maintenance treatment of geritinib did not relapse. The main side effects included anemia, decreased neutrophil count, rash, and increased aminotransferase. The median follow-up time of 5 patients was 15 months (6-20 months). All 5 cases survived until the last follow-up in November 2021 and 4 patients were disease-free.Conclusions:Relapsed and refractory AML patients with FLT3 mutation can achieve complete remission after treatment with geritinib and get a chance for transplantation. Geritinib may reduce the risk of recurrence after transplantation and improve survival rate. No serious side effects occur in geritinib treatment.

Objective:To evaluate the role of cathepsin B (CTSB) in mechanical ventilator-induced lung injury (VILI) in rats and the relationship with NOD-like receptor pyrin domain containing 3 (NLRP3) inflammasome.Methods:Thirty-six SPF-grade healthy male Sprague-Dawley rats, aged 6-8 weeks, weighing 220-300 g, were divided into 3 groups ( n=12 each) by the random number table method: control group (group C), VILI group (group V) and VILI + CA074-me group (group Me). CA074-me 5 mg/kg was intraperitoneally injected in group Me, while the equal volume of normal saline was given instead in group C and group V. Group C kept spontaneous breathing for 4 h, and the animals were mechanically ventilated (tidal volume 20 ml/kg, respiratory rate 80 breaths/min, fraction of inspired oxygen 21%, PEEP 0 cmH 2O). Blood samples from femoral artery were collected for arterial blood gas analysis before tracheal intubation and after spontaneous breathing or ventilation, and PaO 2 was recorded.Rats were sacrificed, and bronchoalveolar lavage fluid (BALF) was collected and lung tissues were collected for determination of the wet/dry lung weight ratio (W/D ratio), serum interleukin-1beta (IL-1β) and IL-18 concentrations in BALF (by enzyme-linked immunosorbent assay), expression of CTSB, NLRP3, apoptosis-associated speck-like protein containing a caspase-1 recruitment domain (ASC) and caspase-1 mRNA in lung tissues (quantitative real-time polymerase chain reaction), and expression of CTSB, NLRP3, ASC and caspase-1 in lung tissues (by Western blot) and for microscopic examination of the pathological changes (using HE staining). Lung injury was assessed and scored. Results:Compared with group C, PaO 2 was significantly decreased after the end of ventilation, the lung injury score, W/D ratio and concentrations of IL-1β and IL-18 in serum and BALF were increased, and the expression of CTSB, NLRP3, ASC and caspase-1 protein and mRNA in lung tissues was up-regulated in group V and group Me ( P<0.01). Compared with group V, PaO 2 was significantly increased after the end of ventilation, the lung injury score, W/D ratio and concentrations of IL-1β and IL-18 in serum and BALF were decreased, and the expression of CTSB, NLRP3, ASC and caspase-1 protein and mRNA in lung tissues was down-regulated in group Me ( P<0.01). Conclusions:CTSB is involved in VILI in the rats, and the mechanism may be related to activation of NLRP3 inflammasomes.