Objective To investigate the role and mechanism of ubiquitin-specific protease 46 (USP46) in hypoxia/reoxygenation (H/R)-induced pyroptosis of renal tubular epithelial cells. Methods Renal tubular epithelial cells were divided into negative control siRNA group (si-CTL group), USP46 knockdown group (si-USP46 group), negative control siRNA + H/R treatment group (si-CTL+H/R group), and USP46 knockdown + H/R treatment group (si-USP46+H/R group). Flow cytometry was used to detect cell apoptosis in each group. Real-time quantitative polymerase chain reaction (RT-qPCR) was employed to measure the messenger RNA (mRNA) expression of USP46, NOD-like receptor protein 3 (NLRP3), gasdermin D (GSDMD), interleukin (IL)-18, and IL-1β. Western blotting was used to detect the protein expression of USP46, NLRP3, GSDMD, and cleaved cysteinyl aspartate specific proteinase (C-Caspase)-1. The levels of inflammatory factors and lactate dehydrogenase (LDH) in the cell supernatants were detected, and the levels of reactive oxygen species (ROS) and malondialdehyde (MDA) in the cells were detected. Co-immunoprecipitation was used to verify the interaction between USP46 and NLRP3. Results Compared with the si-CTL group, the si-CTL+H/R group exhibited increased cell apoptosis, elevated protein expression of USP46, NLRP3, GSDMD-N and C-Caspase-1, increased mRNA expression of USP46, NLRP3, GSDMD, IL-18 and IL-1β, higher levels of IL-18, IL-1β, TNF-α and LDH, and increased ROS and MDA levels (all P < 0.05). Compared with the si-CTL+H/R group, the si-USP46+H/R group showed decreased cell apoptosis, reduced protein expression of USP46, NLRP3, GSDMD-N and C-Caspase-1, decreased mRNA expression of USP46, GSDMD and IL-18, lower levels of IL-18, IL-1β, TNF-α and LDH, and decreased ROS and MDA levels (all P < 0.05). Co-immunoprecipitation results indicated that USP46 could bind to NLRP3. Conclusions Downregulation of USP46 alleviates H/R-induced pyroptosis in renal tubular epithelial cells, possibly by inhibiting USP46-dependent NLRP3 deubiquitination and promoting NLRP3 ubiquitination and degradation.

Objective To evaluate the effect of spliced X-box binding protein 1 (XBP1s) on hypoxia/reoxygenation (H/R) injury of mouse renal tubular epithelial cells and unravel underlying mechanism. Methods Mouse renal tubular epithelial cells were divided into adenovirus negative control group (Ad-shNC group), targeted silencing XBP1s adenovirus group (Ad-shXBP1s group), Ad-shNC+H/R group and Ad-shXBP1s+H/R group. The apoptosis level, mitochondrial reactive oxygen activity, mitochondrial membrane potential and mitochondrial calcium ion level were detected in each group. Chromatin immunocoprecipitation followed by sequencing (ChIP-seq) was employed to analyze the binding sites of XBP1s in regulating the inositol 1,4,5-trisphosphate receptor (ITPR) family. The expression levels of XBP1s and ITPR family messenger RNA (mRNA) and protein were determined in each group. Results Compared with the Ad-shNC group, the apoptosis level was higher, mitochondrial reactive oxygen species level was increased, mitochondrial membrane potential was decreased and mitochondrial calcium ion level was elevated in the Ad-shNC+H/R group. Compared with the Ad-shNC+H/R group, the apoptosis level was lower, mitochondrial reactive oxygen species level was decreased, mitochondrial membrane potential was elevated, and mitochondrial calcium ion level was decreased in the Ad-shXBP1s+H/R group (all P<0.05). Compared with the Ad-shNC group, relative expression levels of XBP1s, ITPR1, ITPR2 and ITPR3 mRNAs and proteins were down-regulated in the Ad-shXBP1s group (all P<0.05). Compared with the Ad-shNC group, relative expression levels of XBP1s, ITPR1, ITPR2 and ITPR3 proteins were up-regulated in the Ad-shNC+H/R group. Compared with the Ad-shNC+H/R group, relative expression levels of XBP1s, ITPR1, ITPR2 and ITPR3 were down-regulated in the Ad-shXBP1s+H/R group (all P<0.05). ChIP-seq results showed that XBP1s could bind to the promoter and exon of ITPR1, the exon of ITPR2, and the exon of ITPR3. Conclusions XBP1s may affect mitochondria-associated endoplasmic reticulum membrane structure and function by directly regulating ITPR transcription and translation. Down-regulating XBP1s may inhibit ITPR expression and mitigate mitochondrial damage.

Objective To evaluate the effect of glycogen synthase kinase 3β (GSK3β) on hypoxia/reoxygenation (H/R)-induced injury of senescent renal tubular epithelial cell (RTEC) in aged mice and its regulatory mechanism. Methods RTEC were divided into the Young group (young RTEC with normal growth), Old group (aged RTEC induced by Etoposide), Old+Ad-shNC+H/R group [aged RTEC induced by Etoposide and then transfected with adenovirus negative control (Ad-shNC) for H/R treatment], and Old+Ad-shGSK3β+H/R group (aged RTEC induced by Etoposide and then transfected with short-hairpin RNA-expressing adenovirus with targeted silencing GSK3β for H/R treatment), respectively. Apoptosis level and mitochondrial reactive oxygen species level were detected by flow cytometry. Calcium ion level was determined by immunofluorescence staining. The expression and phosphorylation levels of GSK3β, mitochondria-associated endoplasmic reticulum membrane (MAM)-related proteins of inositol 1,4,5-trisphosphate receptor1 (ITPR1), voltage dependent anion-selective channel 1(VDAC1) and glucose-regulated protein 75 (GRP75) were detected by Western blot. The interaction between GSK3β and MAM-related proteins was analyzed by immunoprecipitation. Results Compared with the Young group, the apoptosis, mitochondrial reactive oxygen species and mitochondrial calcium ion levels were higher in the Old group. Compared with the Old group, the apoptosis, mitochondrial reactive oxygen species and mitochondrial calcium ion levels were higher in the Old+Ad-shNC+H/R group. Compared with the Old+Ad-shNC+H/R group, the apoptosis, mitochondrial reactive oxygen species and mitochondrial calcium ion levels were lower in the Old+Ad-shGSK3β+H/R group, and the differences were statistically significant (all P<0.05). Compared with the Young group, the expression levels of ITPR1, GRP75 and GSK3β proteins were up-regulated, the phosphorylation levels of ITPR1 and GRP75 were increased, whereas the total protein and phosphorylation levels of VDAC1 were decreased in the Old group. Compared with the Old group, the expression level of GSK3β protein was unchanged, the total protein and phosphorylation levels of ITPR1 and GRP75 were increased, the expression level of total VDAC1 protein remained unchanged and the phosphorylation level was increased in the Old+Ad-shNC+H/R group. Compared with the Old+Ad-shNC+H/R group, the expression level of GSK3β protein was decreased, the expression levels of total ITPR1, GRP75 and VDAC1 proteins were unchanged, whereas the phosphorylation levels were decreased in the Old+Ad-shGSK3β+H/R group. Immunoprecipitation showed that GSK3β could interact with ITPR1, GRP75 and VDAC1 proteins. Conclusions The expression level of GSK3β is up-regulated in senescent RTEC. Down-regulating GSK3β expression may reduce the phosphorylation level of ITPR1-GRP75-VDAC1 complex, constrain the overload of mitochondrial calcium ion, protect mitochondrial function and mitigate cell damage during reperfusion.

Objective To investigate the role and mechanism of spliced X-box binding protein 1 (XBP1s) in the senescence of primary renal tubular epithelial cells induced by hypoxia/reoxygenation (H/R). Methods Primary renal tubular epithelial cells were divided into the normal control group (NC group), H/R group, empty adenovirus negative control group (Ad-shNC group), targeted silencing XBP1s adenovirus group (Ad-shXBP1s group), empty adenovirus+H/R treatment group (Ad-shNC+H/R group) and targeted silencing XBP1s adenovirus+H/R treatment group (Ad-shXBP1s +H/R group), respectively. The expression levels of XBP1s in the NC, H/R, Ad-shNC and Ad-shXBP1s groups were measured. The number of cells stained with β-galactosidase, the expression levels of cell aging markers including p53, p21 and γH2AX, and the levels of reactive oxygen species (ROS), malondialdehyde (MDA) and superoxide dismutase (SOD) were determined in the Ad-shNC, Ad-shNC+H/R and Ad-shXBP1s+H/R groups. Chromatin immunoprecipitation was employed to verify Sirtuin 3 (Sirt3) of XBP1s transcription regulation, and the expression levels of Sirt3 and downstream SOD2 after down-regulation of XBP1s were detected. Mitochondrial reactive oxygen species (mtROS) were detected by flow cytometry. Results Compared with the NC group, the expression level of XBP1s was up-regulated in the H/R group. Compared with the Ad-shNC group, the expression level of XBP1s was down-regulated in the Ad-shXBP1s group (both P<0.001). Compared with the Ad-shNC group, the number of cells stained with β-galactosidase was increased, the expression levels of p53, p21 and γH2AX were up-regulated, the levels of ROS, MDA and mtROS were increased, the SOD activity was decreased, the expression level of Sirt3 was down-regulated, and the ratio of Ac-SOD2/SOD2 was increased in the Ad-shNC+H/R group. Compared with the Ad-shNC+H/R group, the number of cells stained with β-galactosidase was decreased, the expression levels of p53, p21 and γH2AX were down-regulated, the levels of ROS, MDA and mtROS were decreased, the SOD activity was increased, the expression level of Sirt3 was up-regulated and the ratio of Ac-SOD2/SOD2 was decreased in the Ad-shXBP1s+H/R group (all P<0.05). Conclusions Down-regulation of XBP1s may ameliorate the senescence of primary renal tubular epithelial cells induced by H/R, which probably plays a role through the Sirt3/SOD2/mtROS signaling pathway.

Objective To analyze the effect of a high salt diet on ovarian mitochondrial function.Methods Twenty female ICR mice were randomly divided into a normal salt diet(NSD)group and a high salt diet(HSD)group(n = 10 each).The NSD group was given a normal salt diet and the HSD group was given an 8%NaCl diet for 4 weeks.A high salt-treated cell model was established by inducing COV-434 cells cultured in vitro with NaCl.Western blotting was used to detect the protein expression of superoxide dismutase(SOD)and ComplexesⅠ-Ⅴ.The activity of SOD and succinate dehydrogenase(SDH)was detected kinetically.A chemiluminescence assay was used to detect adenosine triphosphate(ATP)levels.Results Compared with the NSD,the HSD significantly reduced the expression level of ComplexⅠin ovarian mitochondria(P＜0.01),significantly increased the expression level of ComplexⅤ(P＜0.05),and significantly reduced the activity of SDH and content of ATP(P＜0.01).The expression level of ComplexesⅠandⅡdecreased significantly(P＜0.05),expression level of ComplexⅤ increased significantly(P＜0.05),activity of SDH decreased significantly(P＜0.01),and content of ATP was insufficient(P＜0.01)in COV-434 cells cultured under high salt conditions.Conclusion High salt can lead to mitochondrial dys-function in the mouse ovary,such as imbalanced oxidative homeostasis,changed expression level of electron transport chain complexes,blocked tricarboxylic acid cycle,and insufficient ATP level.

OBJECTIVE To establish a GC-MS/MS analytical method for the determination of 1-chloroethyl cyclohexyl carbonate in candesartan cilexetil tablets. METHODS The analytical column was DB-5MS(30 m×0.25 mm, 0.25 μm). The column temperature was maintained at 80 ℃, then was raised to 300 ℃ at the rate of 20 ℃·min–1 and was maintained for 5 min. Helium was used as carrier gas, and its flow rate was 1.0 mL·min–1. The detection was achieved in multiple reaction monitoring mode. RESULTS The calibration curve of 1-chloroethyl cyclohexyl carbonate had good linearity in the concentration range of 4.4−437.8 ng·mL–1. The limits of quantification and detection were 4.4 and 2.2 ng·mL–1, respectively. The average recovery was 95.6%(RSD=6.3%, n=9). CONCLUSION This method has satisfactory convenience, good sensitivity and high accuracy, and it is suitable for the determination of 1-chloroethyl cyclohexyl carbonate in candesartan cilexetil tablets.

【Objective】 To observe the effects of mirror neuron system theory(MNST) on hand dexterity and fine motor function in preschool children with developmental coordination disorder(DCD), so as to provide reference for the rehabilitation of children with DCD. 【Methods】 A total of 51 children with DCD treated at Nantong Maternal and Child Health Care Hospital from June 2021 to April 2023 were enrolled in this study, and were randomly assigned to treatment group (n=26) and control group (n=25) except for 5 cases who missed in the follow-up. Both groups received conventional rehabilitation training, while the treatment group received MNST additionally. The hand dexterity and fine motor function of both groups were assessed using the hand dexterity subscale of the Movement Assessment Battery for Children-Second Edition (MABC-2), Peabody Developmental Motor Scale-Fine Motor (PDMS-FM), and Function Independence Measure for Children (WeeFIM) before and after 12 weeks of treatment. 【Results】 Before treatment, there were no significant differences in hand dexterity subscale of MABC-2, PDMS-FM, and WeeFIM scores between the two groups (P>0.05). After treatment, both groups showed improvements in hand dexterity subscale of MABC-2, PDMS-FM, and WeeFIM scores (treatment group: t=35.620, 42.084, 40.072; control group: t=14.000, 12.017, 14.054, P<0.001), with the treatment group showing significantly greater improvements compared to the control group (t=2.611, 3.120, 2.331, P<0.05). 【Conclusion】 MNST combined with conventional rehabilitation training can enhance hand dexterity and fine motor function in children with DCD, thereby improving children′s activities of daily living.

Objective To explore the clinicopathological and immunohistochemical features of nephrogenic adenoma(NA).Methods Clinical data of NA patients diagnosed in the Department of Pathology,Zhongshan Hospital,Fudan University from July 2016 to October 2022 were collected and analyzed to explore their clinicopathological features.Results A total of 13 NA cases were enrolled.There were 11 males and 2 females.Organs involved:ureter(n=7),bladder(n=5),bladder and ureter(n=1),renal pelvis(n=2).NA patients performed as ureteral stenosis(6/7),rough bladder wall(3/5),and renal pelvis polyp(2/2).The typical microscopical features of NA were tubular(13/13)and papillary(4/13)structures,covered with cuboidal or columnar epithelium(13/13),or a mixed hobnail-spike eosinophilic epithelium(12/13);the interstitium was loose,containing varied amounts of vasculature and inflammatory cells(13/13).Immunohistochemistry revealed specific expressions of CK7,PAX-8,CK19 and CK8.Conclusions NA is a rare neoplasm of the urinary system with unique histological features.NA has the risk of misdiagnosis and over-treatment,and the potential of recurrence and malignant conversion.The diagnosis of NA depends on pathology,and the immunohistochemistry can be helpful for its pathological diagnosis.

OBJECTIVE To provide reference for improving the level of hospital pharmaceutical management for operating room drugs. METHODS The operating room pharmacy of our hospital utilized the concept and means of the Internet of Things （IoT） to build an intelligent IoT system for operating room drugs （hereinafter referred to as the “IoT system”）， and optimized and improved it. The quality of drug management in the operating room of our hospital during the initial phase of the IoT system （Q1 2022） and after optimization and improvement （Q1 2023） were compared by setting indicators from four aspects： quality， efficiency， cost， and satisfaction. RESULTS After more than a year of optimization and improvement， our hospital has built a traceable IoT system for the entire drug process that integrated surgical anesthesia systems and hospital information systems， with the direction of drug circulation in the operating room as the axis， using intelligent drug vehicles as the hardware foundation， and anesthesia doctor’s order information system as the software medium. After the optimization and improvement of the IoT system， the standardized score of anesthesia orders in the operating room increased from （68.5±3.5） points in the initial period to （97.0± 2.7） points； the consistency rate between accounts and materials increased from （82.40±8.85）% to （96.50±4.80）%； the time of taking medicine was shortened from （40±8） min to （12±3） min； the frequency of drug withdrawal was reduced from （36.0± 6.5） times/day to （15.5±3.0） times/day； the cost of loss drugs was decreased from （1 292.61±305.90） yuan to （594.24±195.05） yuan； the satisfaction was increased from （80.5±6.5） points to （96.0±3.0） points. All indicators were significantly improved with statistically significant differences （P＜0.05）. CONCLUSIONS The intelligent IoT system constructed by our hospital effectively ensures the accessibility， timeliness， and safety of intraoperative medication， which is conducive to improving the quality of drug management in the operating room.

Objective:To compare the precision and stability of handheld full-course visual navigation and robot-assisted total hip arthroplasty (THA), as well as to evaluate perioperative laboratory indicators and clinical efficacy.Methods:A retrospective analysis was performed on 68 patients with unilateral hip joint disorders who underwent primary THA at the General Hospital of the People's Liberation Army of China from January to June 2024. Patients were divided into two groups based on the assistance method: the Visual Treatment Solution (VTS) group, which included 34 patients (13 males, 21 females; mean age 56.92±8.31 years; body mass index[BMI] 24.20±3.55 kg/m 2), and the MAKO group, also with 34 patients (18 males, 16 females; mean age 57.97±6.54 years; BMI 25.20±3.91 kg/m 2). Among the VTS group, 1 there were 18 cases on the left side and 16 cases on the right side, including 14 cases of femoral head necrosis, 8 cases of congenital hip dysplasia, and 12 cases of hip osteoarthritis. In the MAKO group, there were 19 cases on the left side and 15 cases on the right side, including 13 cases of femoral head necrosis, 10 cases of congenital hip dysplasia, and 11 cases of hip osteoarthritis. Preoperative planning, intraoperative verification deviations, postoperative acetabular prosthesis anteversion and abduction angles, and limb length discrepancies were analyzed. Acetabular angle distribution scatter plots were generated for intraoperative verification and postoperative imaging. The operation time, hospital stay, and laboratory values, including hemoglobin (Hb) loss, fibrinogen degradation products (FDP), D-dimer, high-sensitivity C-reactive protein (hCRP), as well as postoperative pain (measured by the visual analogue scale, VAS), hip Harris score (HHS), and Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) scores were recorded. Results:No statistically significant differences were observed between postoperative imaging measurements of anteversion and abduction angles and intraoperative verification values ( P>0.05). Postoperative imaging indicated that 91% (31/34) of the VTS group and 100% (34/34) of the MAKO group were within the Lewinnek safety zone. The scatter distribution of angles in the MAKO group was more concentrated than that in the VTS group. Furthermore, the angle deviation between intraoperative verification and preoperative planning was significantly greater in the VTS group for both anteversion (2.00° [-1.00°, 4.00°] vs. 0.00° [-1.00°, 1.00°]) and abduction angles (-1.00° [-5.00°, 0.00°] vs. 0.00° [-1.00°, 1.00°]; P<0.05). The bilateral limb length discrepancies were 1.35(0.67, 2.45) mm in the VTS group and 1.65(0.50, 2.52) mm in the MAKO group, with no significant difference noted ( P>0.05). Both methods achieved effective hip biomechanical reconstruction. Additionally, no significant differences in operation time, Hb loss, FDP, D-dimer, hCRP, or postoperative hospital stay were found ( P>0.05). Follow-up at one and three months postoperatively showed no significant differences in VAS (2.94±0.92 vs. 2.97±0.83), HHS (69.88±4.20 vs. 70.06±3.88), or WOMAC scores (22.44±3.15 vs. 22.76±3.39) between the two groups ( P>0.05). Conclusion:The stability of preoperative planning and design in robot-assisted total hip arthroplasty is superior to that of handheld navigation methods. Nevertheless, both approaches demonstrate comparable postoperative prosthesis stability, laboratory indicators, and clinical outcomes.