Objective:To investigate the role of serotonergic (5-HT) neurons activation in the dorsal raphe nucleus (DRN) in heat-induced seizures and its influence on breathing patterns in Scn1a + /- mice. Methods:24-day-old male Scn1a + /- mice were used for experiments, and sudden unexpected death in epilepsy(SUDEP) model was established through heat induction for 5 consecutive days. (1) Fluoxetine intervention experiment: 20 male mice were randomly divided into model group( n=10) and fluoxetine group ( n=10) according the weight-matched method. The fluoxetine group received intraperitoneal injection of fluoxetine (10 mg/kg) 45 min before heat induction each day for 5 consecutive days, while the model group received equal volume of 0.9% NaCl solution. (2) Chemogenetic activation experiment: 20 male mice were randomly divided into vector control group( n=10) and chemogenetic activation group ( n=10) according the weight-matched method. Empty vector or rAAV-TPH2-hM3d(Gq)-EGFP-WPREs was stereotaxically injected into DRN 14 d prior to seizure induction, and deschloroclozapine (5 mg/kg) was intraperitoneally injected 30 min before heat induction. Seizure characteristics and survival were assessed through video monitoring, and respiratory parameters were monitored. Immunofluorescence was used to detect colocalization of tryptophan hydroxylase 2(TPH2) and c-Fos in DRN. Statistical analysis was performed using SPSS 24.0 and GraphPad Prism 9.0. The independent sample t-test or Mann-Whitney test was used for inter-group comparison. Results:(1) In the fluoxetine intervention experiment: the survival rates between the model group and fluoxetine group showed no statistically significant difference ( χ2=2.23, P>0.05). As for the frequency of grade Ⅳ seizures, the model group (1.50(1.25, 2.35)min) demonstrated higher frequency than the fluoxetine group (0.43(0.20, 0.67)min) ( t=-3.40, P<0.05).With respect to respiratory parameters, the model group demonstrated shorter expiratory time ((0.10±0.02) s) and inspiratory time ((0.15±0.02) s) compared to the fluoxetine group ((0.16±0.05) s, (0.19±0.04) s) ( t=-3.47, -3.73, both P<0.01). The respiratory rate in the model group ((269.96±44.84) times/min) was significantly higher compared to the fluoxetine group ((195.04±52.37) times/min) ( t=3.44, P<0.01). The tidal volume in the model group ((0.10±0.02) mL) was significantly lower than the fluoxetine group ((0.13±0.04) mL) ( t=-2.19, P<0.05).The number of TPH2+ /c-Fos+ co-expressing cells in the model group (11.00±4.00) was lower than that in the fluoxetine group (33.00±8.39)( t=-4.16, P<0.05). (2) In the chemogenetic activation experiment: compared to the vehicle group, the chemogenetic activation group demonstrated significantly enhanced survival rates ( χ2=5.83, P<0.05). As for the frequency of grade Ⅴ seizures, the vehicle group (2.11(1.62, 3.44) times/min) showed higher frequency compared to the chemogenetic activation group (0.81(0.00, 1.62) times/min) ( t=18.00, P<0.05). In terms of respiratory parameters, the vehicle group showed shorter expiratory time ((0.10±0.01) s) and inspiratory time ((0.14±0.01) s) compared to the chemogenetic activation group ((0.12±0.01) s, (0.15±0.01) s) ( t=-2.78, -2.50, both P<0.05). The respiratory rate in the vehicle group ((208.37±9.73) times/min) was significantly higher than the chemogenetic activation group ((191.85±8.83) times/min) ( t=3.98, P<0.01). The tidal volume in the vehicle group ((0.09±0.01) mL) was significantly lower than the chemogenetic activation group ((0.12±0.02) mL) ( t=-4.77, P<0.001).The number of TPH2+ /c-Fos+ co-expressing cells in the vehicle group (9.00±3.46) was lower than that in the chemogenetic activation group (43.00±11.02)( t=-5.20, P<0.01). Conclusion:Specific activation of serotonergic neurons in the DRN can ameliorate heat-induced epileptic symptoms, improve respiratory function, and prolong survival time in Scn1a + /- mice.

Objective:To investigate the role of serotonergic (5-HT) neurons activation in the dorsal raphe nucleus (DRN) in heat-induced seizures and its influence on breathing patterns in Scn1a + /- mice. Methods:24-day-old male Scn1a + /- mice were used for experiments, and sudden unexpected death in epilepsy(SUDEP) model was established through heat induction for 5 consecutive days. (1) Fluoxetine intervention experiment: 20 male mice were randomly divided into model group( n=10) and fluoxetine group ( n=10) according the weight-matched method. The fluoxetine group received intraperitoneal injection of fluoxetine (10 mg/kg) 45 min before heat induction each day for 5 consecutive days, while the model group received equal volume of 0.9% NaCl solution. (2) Chemogenetic activation experiment: 20 male mice were randomly divided into vector control group( n=10) and chemogenetic activation group ( n=10) according the weight-matched method. Empty vector or rAAV-TPH2-hM3d(Gq)-EGFP-WPREs was stereotaxically injected into DRN 14 d prior to seizure induction, and deschloroclozapine (5 mg/kg) was intraperitoneally injected 30 min before heat induction. Seizure characteristics and survival were assessed through video monitoring, and respiratory parameters were monitored. Immunofluorescence was used to detect colocalization of tryptophan hydroxylase 2(TPH2) and c-Fos in DRN. Statistical analysis was performed using SPSS 24.0 and GraphPad Prism 9.0. The independent sample t-test or Mann-Whitney test was used for inter-group comparison. Results:(1) In the fluoxetine intervention experiment: the survival rates between the model group and fluoxetine group showed no statistically significant difference ( χ2=2.23, P>0.05). As for the frequency of grade Ⅳ seizures, the model group (1.50(1.25, 2.35)min) demonstrated higher frequency than the fluoxetine group (0.43(0.20, 0.67)min) ( t=-3.40, P<0.05).With respect to respiratory parameters, the model group demonstrated shorter expiratory time ((0.10±0.02) s) and inspiratory time ((0.15±0.02) s) compared to the fluoxetine group ((0.16±0.05) s, (0.19±0.04) s) ( t=-3.47, -3.73, both P<0.01). The respiratory rate in the model group ((269.96±44.84) times/min) was significantly higher compared to the fluoxetine group ((195.04±52.37) times/min) ( t=3.44, P<0.01). The tidal volume in the model group ((0.10±0.02) mL) was significantly lower than the fluoxetine group ((0.13±0.04) mL) ( t=-2.19, P<0.05).The number of TPH2+ /c-Fos+ co-expressing cells in the model group (11.00±4.00) was lower than that in the fluoxetine group (33.00±8.39)( t=-4.16, P<0.05). (2) In the chemogenetic activation experiment: compared to the vehicle group, the chemogenetic activation group demonstrated significantly enhanced survival rates ( χ2=5.83, P<0.05). As for the frequency of grade Ⅴ seizures, the vehicle group (2.11(1.62, 3.44) times/min) showed higher frequency compared to the chemogenetic activation group (0.81(0.00, 1.62) times/min) ( t=18.00, P<0.05). In terms of respiratory parameters, the vehicle group showed shorter expiratory time ((0.10±0.01) s) and inspiratory time ((0.14±0.01) s) compared to the chemogenetic activation group ((0.12±0.01) s, (0.15±0.01) s) ( t=-2.78, -2.50, both P<0.05). The respiratory rate in the vehicle group ((208.37±9.73) times/min) was significantly higher than the chemogenetic activation group ((191.85±8.83) times/min) ( t=3.98, P<0.01). The tidal volume in the vehicle group ((0.09±0.01) mL) was significantly lower than the chemogenetic activation group ((0.12±0.02) mL) ( t=-4.77, P<0.001).The number of TPH2+ /c-Fos+ co-expressing cells in the vehicle group (9.00±3.46) was lower than that in the chemogenetic activation group (43.00±11.02)( t=-5.20, P<0.01). Conclusion:Specific activation of serotonergic neurons in the DRN can ameliorate heat-induced epileptic symptoms, improve respiratory function, and prolong survival time in Scn1a + /- mice.

Objective:To investigate the association between programmed cell death protein 1 (PD-1) and its ligand PD-L1 and cytokines in patients with polycystic ovary syndrome (PCOS).Methods:Using the GSE54248 dataset from the GEO database, differentially expressed PD1/PD-L1 pathway-related genes in PCOS were identified and subjected to GO and KEGG pathway enrichment analysis. In this case-control study, totally 105 patients with PCOS (named PCOS group) and 109 non-PCOS patients (named control group) who were treated at the Reproductive Assisted Reproduction Center of the First Affiliated Hospital of Xinjiang Medical University from January 2022 to June 2023 were recruited. The QBPlex flow cytometry high-throughput multiplex assay was utilized to assess the peripheral blood levels of PD-L1, PD-L2, PD-1, and cytokines in PCOS group and control group. Pearson's method was used for correlation analysis.Results:In PCOS group, the PD-1 level in peripheral blood [2.890 (0.020, 4.540) ng/L] was significantly lower than that of control group [3.370 (2.460, 4.360) ng/L, P=0.008], the PD-L1 level [9.820 (8.860, 10.880) ng/L] was lower than that in control group [10.410 (9.700, 11.160) ng/L, P=0.001]. There was no significant difference in the expression level of PD-L2 between the two groups ( P>0.05). From the GSE54248 dataset, 26 differentially expressed genes were identified, primarily enriched in the PD-1/PD-L1 pathway, Th1 and Th2 cell differentiation, and pathways associated with the production of cytokines involved in inflammatory responses. Compared with control group, PCOS group exhibited a significant decrease in the peripheral blood concentrations of interleukin (IL)-5, IL-9, IL-25, IL-10, growth stimulation expressed gene 2 (ST-2), and Granzyme B, and a significant increase in IL-8, IL-1RA, and tumor necrosis factor-α (TNF-α) levels, with all differences being statistically significant (all P<0.05). PD-1 exhibited positive correlations with the levels of IL-1RA, ST-2, and TNF-α ( r=0.270, P=0.005; r=0.213, P=0.029; r=0.291, P=0.003), while it exhibited negative correlations with the levels of IL-9, IL-25, and Granzyme B ( r=-0.322, P<0.001; r=-0.211, P=0.031; r=-0.369, P<0.001). PD-L1 demonstrated positive correlations with the levels of IL-9, IL-25, and Granzyme B ( r=0.254, P=0.009; r=0.330, P<0.001; r=0.340, P<0.001), and a negative correlation with IL-10 level ( r=-0.373, P=0.009). Conclusion:The expression of PD-1 and PD-L1 in the peripheral blood of PCOS patients is down-regulated, which may be associated with an imbalance in Th1/Th2 cytokines and serve as potential molecular biomarkers for the treatment of PCOS.

Objective:To investigate the association between programmed cell death protein 1 (PD-1) and its ligand PD-L1 and cytokines in patients with polycystic ovary syndrome (PCOS).Methods:Using the GSE54248 dataset from the GEO database, differentially expressed PD1/PD-L1 pathway-related genes in PCOS were identified and subjected to GO and KEGG pathway enrichment analysis. In this case-control study, totally 105 patients with PCOS (named PCOS group) and 109 non-PCOS patients (named control group) who were treated at the Reproductive Assisted Reproduction Center of the First Affiliated Hospital of Xinjiang Medical University from January 2022 to June 2023 were recruited. The QBPlex flow cytometry high-throughput multiplex assay was utilized to assess the peripheral blood levels of PD-L1, PD-L2, PD-1, and cytokines in PCOS group and control group. Pearson's method was used for correlation analysis.Results:In PCOS group, the PD-1 level in peripheral blood [2.890 (0.020, 4.540) ng/L] was significantly lower than that of control group [3.370 (2.460, 4.360) ng/L, P=0.008], the PD-L1 level [9.820 (8.860, 10.880) ng/L] was lower than that in control group [10.410 (9.700, 11.160) ng/L, P=0.001]. There was no significant difference in the expression level of PD-L2 between the two groups ( P>0.05). From the GSE54248 dataset, 26 differentially expressed genes were identified, primarily enriched in the PD-1/PD-L1 pathway, Th1 and Th2 cell differentiation, and pathways associated with the production of cytokines involved in inflammatory responses. Compared with control group, PCOS group exhibited a significant decrease in the peripheral blood concentrations of interleukin (IL)-5, IL-9, IL-25, IL-10, growth stimulation expressed gene 2 (ST-2), and Granzyme B, and a significant increase in IL-8, IL-1RA, and tumor necrosis factor-α (TNF-α) levels, with all differences being statistically significant (all P<0.05). PD-1 exhibited positive correlations with the levels of IL-1RA, ST-2, and TNF-α ( r=0.270, P=0.005; r=0.213, P=0.029; r=0.291, P=0.003), while it exhibited negative correlations with the levels of IL-9, IL-25, and Granzyme B ( r=-0.322, P<0.001; r=-0.211, P=0.031; r=-0.369, P<0.001). PD-L1 demonstrated positive correlations with the levels of IL-9, IL-25, and Granzyme B ( r=0.254, P=0.009; r=0.330, P<0.001; r=0.340, P<0.001), and a negative correlation with IL-10 level ( r=-0.373, P=0.009). Conclusion:The expression of PD-1 and PD-L1 in the peripheral blood of PCOS patients is down-regulated, which may be associated with an imbalance in Th1/Th2 cytokines and serve as potential molecular biomarkers for the treatment of PCOS.

Purpose: Laparoscopy is being increasingly accepted for pancreaticoduodenectomy. Stapled anastomosis (SA) is used extensively to facilitate laparoscopic pancreaticoduodenectomy (LPD); however, the incidence of anastomotic bleeding after stapled gastrointestinal anastomosis is still high. Methods: One hundred and thirty-nine patients who underwent LPD using Whipple method were enrolled in our study. We performed the SA with our reinforced method (n = 68, R method) and without the method (n = 71, NR method). We compared the clinical characteristics and anastomosis methods of patients with or without gastrointestinal-anastomotic hemorrhage (GAH), and operative parameters were also compared between the anastomotic methods. Results: Of the 139 patients undergoing LPD, 15 of them developed GAH. The clinical characteristics of patients with or without GAH were not significantly different except in the anastomotic method (P < 0.001). In the univariate logistic regression analyses, only the anastomotic method was associated with GAH. Furthermore, patients with the NR method had significantly higher incidences of GAH (P < 0.001) and Clavien-Dindo grade ≥ III complications (P < 0.001). Conclusion Our retrospective analysis showed that the SA performed with reinforced method might be a reform of SA without the reinforcement, as indicated by the lower incidence of GAH. However, further research is necessary to evaluate the utility of this reinforced method.

Dendritic Cells ; cytology ; metabolism ; Enzyme-Linked Immunosorbent Assay ; HEK293 Cells ; Humans ; Interferon Regulatory Factor-7 ; metabolism ; Interferon Type I ; metabolism ; Interferon-gamma ; analysis ; Interleukin-6 ; analysis ; Oligonucleotides ; metabolism ; RNA Interference ; RNA, Small Interfering ; metabolism ; Real-Time Polymerase Chain Reaction ; Receptors, Tumor Necrosis Factor ; antagonists & inhibitors ; genetics ; metabolism

Objective To explore the establishment of a rat model of acute radiation-induced liver injury and sig-nificance of the dynamic changes of TGF-β1 expression.Methods Forty healthy 6-week old male SD rats were randomly divided into model group (n=30) and control group (n=10).The right liver of rats in the model group was given a single dose of 25 Gy 6 MV X-ray irradiation.Histopathological examination using HE staining and transmission electron microsco-py were conducted to observe the liver pathological changes in rats at 3, 5, and 10 days after irradiation, serum TGF-β1 was detected, and relevant indicators of liver function ( ALT, AST, ALP) were determined.Statistical analysis was per-formed using SPSS 17.0 software.Results At 3, 5 and 10 days after irradiation, early pathological changes in the liver cells were observed by electron microscopy, the expression of TGF-β1 was gradually increased with the time prolongation, and significant differences were found between the model group and the control group at different time points (P<0.05). The light microscopic observation of liver tissues did not show significant differences between the control group and model group.The liver ALT, AST, ALP at different time points did not show significant differences between the two groups ( P>0.05).Conclusion Electron microscopy can be used to evaluate the early changes of radiation-induced liver injury, pri-or to the alterations visible by routine light microscopy.TGF-β1 can be used to predict the degree of radiation-induced liver injury, and may be used as a sensitive serum cytokine in predicting the degree of radiation-induced acute liver injury.

BACKGROUNDThe signal transducer and activator of transcription 6 (STAT6) expression in lung epithelial cells plays a pivotal role in asthma pathogenesis. Activation of STAT6 expression results in T helper cell type 2 (Th2) cell differentiation leading to Th2-mediated IgE production, development of allergic airway inflammation and hyperreactivity. Therefore, antagonizing the expression and/or the function of STAT6 could be used as a mode of therapy for allergic airway inflammation.

METHODSIn this study, we synthesized a 20-mer phosphorothioate antisense oligonucleotide (ASODN) overlapping the translation starting site of STAT6 and constructed STAT6 antisense RNA (pANTI-STAT6), then transfected them into murine spleen lymphocytes and analyzed the effects of antagonizing STAT6 function in vitro and in a murine model of asthma.

RESULTSIn vitro, we showed suppression of STAT6 expression and interleukin (IL)-4 production of lymphocytes by STAT6 ASODN. This effect was more prominent when cells were cultured with pANTI-STAT6. In a murine model of asthma associated with allergic pulmonary inflammation in ovalbumin (OVA)-sensitized mice, local intranasal administration of fluorescein isothiocyanate (FITC)-labeled STAT6 ASODN to DNA uptake in lung cells was accompanied by a reduction of intracellular STAT6 expression. Such intrapulmonary blockade of STAT6 expression abrogated signs of lung inflammation, infiltration of eosinophils and Th2 cytokine production.

CONCLUSIONThese data suggest a critical role of STAT6 in the pathogenesis of asthma and the use of local delivery of STAT6 ASODN as a novel approach for the treatment of allergic airway inflammation such as in asthma.

Animals ; Asthma ; drug therapy ; metabolism ; Blotting, Western ; Cell Differentiation ; drug effects ; Cells, Cultured ; Female ; Interleukin-4 ; metabolism ; Lymphocytes ; drug effects ; metabolism ; Mice ; Mice, Inbred C57BL ; Oligonucleotides, Antisense ; chemistry ; pharmacology ; Phosphates ; pharmacology ; RNA, Antisense ; chemistry ; pharmacology ; Reverse Transcriptase Polymerase Chain Reaction ; STAT6 Transcription Factor ; genetics ; metabolism ; Th2 Cells ; drug effects ; metabolism

AIMTo prepare verapamil hydrochloride controlled-onset extended-release pellets (VH-COERP) and study its release behavior in vitro. To compare the pharmacokinetic characteristics and bioavailability in six Beagle dogs after oral administration of VH-COERP and verapamil hydrochloride delayed-release pellets (VH-DRP) as reference.

METHODSThe core of VH-COERP were prepared in the fluidized bed (mini-glatt) by spraying water solution containing drugs onto sucrose-starch pellets with hydroroxy propyl methyl cellulose (HPMC) as the inner coating swelling layer and ethylcellulouse aqueous dispersion as the outer coating controlled layer. Through modifying the coating level of inner and outer layer, the VH-COERP with the optimized cumulative release profile was obtained. The concentration of VH in plasma of six dogs and its pharmacokinetic behaviors after oral administration of VH-COERP and VH-DRP at different times were studied by RP-HPLC. The pharmacokinetic parameters were computed by software program 3P97.

RESULTSThe lag time, the release behavior and the amount of VH from VH-COERP within 24 hours were not influenced by the pH of dissolution medium and post-process, but obviously influenced by the different kinds of added material in swelling layer and the coating level of the inner swelling layer and the outer controlled layer. In vitro the lag time of release profile of VH from VH-COERP was 5 h and then VH was extended release from VH-COERP in the following time. Compared with the VH-DRP, VH-COERP in vivo has an obviously lag time (4 h) , Tmax was also delayed (8 h) and the relative bioavailability was (94.56 +/- 7.64)%.

CONCLUSIONThe release profile of VH from VH-COERP was shown to be extended-release after an conspicuous lag time in vitro and in vivo. So the drug can be taken by the patient before bed time and begin to work at the morning.

Administration, Oral ; Animals ; Biological Availability ; Calcium Channel Blockers ; administration & dosage ; pharmacokinetics ; Cellulose ; analogs & derivatives ; chemistry ; Delayed-Action Preparations ; Dogs ; Drug Stability ; Hypromellose Derivatives ; Methylcellulose ; analogs & derivatives ; chemistry ; Microscopy, Electron, Scanning ; Verapamil ; administration & dosage ; chemistry ; pharmacokinetics