Objective: To investigate the spatio-temporal clustering characteristics of influenza in Ningxia Hui Autonomous Region from 2014 to 2023, so as to provide the basis for strengthening influenza prevention and control. Methods: Data pertaining to influenza cases reported in Ningxia Hui Autonomous Region from 2014 to 2023 were retrieved from the Infectious Disease Surveillance System of the Chinese Disease Prevention and Control Information System, including age, sex, current residence, onset date, and reporting date. The seasonal incidence of influenza was analyzed using seasonal index. The spatio-temporal clustering characteristics of influenza were identified using spatial autocorrelation analysis and spatio-temporal scan analysis. Results: A total of 20 377 influenza cases were reported in Ningxia Hui Autonomous Region from 2014 to 2023, with a male-to-female ratio of 1.15∶1. The majority were children under 15 years, with 10 950 cases accounting for 53.74%. Influenza was highly prevalent in January, February, March, and December, with seasonal indices of 219.06%, 111.00%, 246.65%, and 366.24%, respectively. The average annual reported incidence was 29.55/100 000, among which Pengyang County, Jinfeng District, Dawukou District, Xiji County, and Litong District had higher average annual reported incidence, at 63.99/100 000, 55.71/100 000, 55.70/100 000, 49.49/100 000, and 49.04/100 000, respectively. Spatial autocorrelation analysis showed that in 2023, there was spatial clustering of influenza cases in Ningxia Hui Autonomous Region (Moran's I=0.333, P<0.05), with a high-high cluster in Jingyuan County, while in other years, the distribution of influenza cases was random (all P>0.05). Spatio-temporal scan analysis showed that from 2014 to 2023, there were four space-time clusters in Ningxia Hui Autonomous Region, including one type Ⅰ cluster in Hongsibao District of Wuzhong City, with the clustering period from January 20 to 26, 2014; and three type Ⅱ clusters, mainly in January, February, March and December, covering one area in Shizuishan City, five areas in Guyuan City, one area in Zhongwei City, three areas in Wuzhong City, and four areas in Yinchuan City. Conclusions From 2014 to 2023, children under 15 years were the primary population affected by influenza in Ningxia Hui Autonomous Region, with distinct spatio-temporal distribution characteristics. The peak incidence occurred during the winter and spring seasons, and the main clustering areas were in the southern regions.

Objective:To discuss whether safranal affects the proliferation,migration,and phenotypic transformation of the vascular smooth muscle cells(VSMCs)in a high-glucose environment and to clarify the function of safranal in the prevention and treatment of diabetic(DM)vascular complications.Methods:The SD rats were selected as experimental subjects;primary VSMCs were cultured from rat thoracic aortas and divided into control group,25 mmol·L-1 high glucose(HG)group,HG+20 μmol·L-1 safranal group,HG+40 μmol·L-1 safranal group,and HG+80 μmol·L-1 safranal group.The cells in control group received no treatment;the cells in 25 mmol·L-1 HG group were pretreated with 25 mmol·L-1 HG;the cells in HG+20,40,and 80 μmol·L-1 safranal groups were further treated with 20,40,and 80 μmol·L-1 safranal respectively for 48 h on the basis of 25 mmol·L-1 HG group.Cell counting kit-8(CCK-8)method was used to determine the appropriate concentration of safranal and detect the viabilities of the VSMCs in various groups;cell scratch healing assay was used to detect the scratch healing rates of the VSMCs in various groups;Transwell chamber assay was used to detect the numbers of the migration VSMCs in various groups;immunofluorescence method was used to detect the fluorescence intensities of alpha-smooth muscle actin(α-SMA)and rabbit anti-osteopontin(OPN)in the VSMCs in various groups;Western blotting method was used to detect the expression levels of OPN,α-SMA,and proliferating cell nuclear antigen(PCNA)in the VSMCs in various groups.Results:Under microscope,on the 4th day of in vitro culture,the spindle-shaped or triangular cells crawled out from the edge of the thoracic aorta tissue blocks,with long spindle being the most common morphology.On the 14th,the cells gradually covered the bottom of the dish;when cell density reached 80%-90%,the characteristic"hills and valleys"growth pattern appeared.Third-generation cells were taken for immunofluorescence identification;immunofluorescence staining with VSMC-specific marker α-SMA showed positive expression of α-SMA protein in the primarily cultured VSMCs.The CCK-8 assay results showed that compared with control group,the cell viability of the cells in 160 μmol·L-1 safranal group was significantly decreased(P<0.01),indicating toxic damage to the cells.Under the conditions of safranal concentrations at 20,40,and 80 μmol·L-1 respectively,after 48 h intervention on VSMCs,no significant adverse effect on cell viability was observed;considering both the effect and toxicity of safranal,these three concentrations were used in subsequent cell experiments.After 48 h intervention,compared with control group,the activity of the VSMCs in 25 mmol·L-1 HG group was increased(P<0.001);compared with 25 mmol·L-1 HG group,the activities of the VSMCs in HG+20,40,and 80 μmol·L-1 safranal groups were gradually decreased(P<0.05).The cell scratch healing assay and Transwell assay results showed that after 48 h intervention,the scratch healing rate of the VSMCs in 25 mmol·L-1 HG group was significantly higher than that in control group(P<0.01),and the number of transmembrane cells through the Transwell chamber was significantly increased(P<0.05);compared with 25 mmol·L-1 HG group,the scratch healing rates of the VSMCs in HG+20,40,and 80 μmol·L-1 safranal groups were gradually decreased(P<0.05),and the number of transmembrane cells was decreased(P<0.05).The immunofluorescence staining results showed that compared with control group,the fluorescence intensity of α-SMA protein in the VSMCs in 25 mmol·L-1 HG group was significantly weakened(P<0.001),while the fluorescence intensity of OPN protein was significantly enhanced(P<0.001);compared with 25 mmol·L-1 HG group,the fluorescence intensities of α-SMA protein in the VSMCs in HG+20,40,and 80 μmol·L-1 safranal groups were gradually increased(P<0.05),and the fluorescence intensities of OPN were gradually weakened(P<0.05).The Western blotting method results showed that compared with control group,the expression level of α-SMA protein in the VSMCs in 25 mmol·L-1 HG group was decreased(P<0.05),and the expression levels of PCNA and OPN proteins were increased(P<0.01);compared with 25 mmol·L-1 HG group,the expression level of α-SMA protein in the VSMCs in HG+20,40,and 80 μmol·L-1 safranal groups were increased(P<0.05),and the expression levels of PCNA and OPN proteins were decreased(P<0.05).Conclusion:Safranal can inhibit the proliferation,migration,and phenotypic transformation of the VSMCs induced by high glucose.

In order to establish a stable in vitro culture platform for chicken small intestine three-dimensional (3D) organoids, in this study, crypt cells were collected from the small intestine of 18-day-old embryos of AA broilers. On the basis of the L-WRN conditioned medium, we optimized the culture conditions of chicken small intestinal organoids by adjusting the proportions of nicotinamide, N-acetylcysteine, LY2157299, CHIR99021, Jagged-1, FGF, and other cytokines to select the medium suitable for the long-term stable growth of the organoids. The optimization results showed that the addition of 1.5 µmol/L CHIR99021 significantly improved the organoid formation efficiency and organoid diameter. When 0.5 µmol/L Jagged-1 was added, a small amount of bud-like tissue appeared in organoids. After the addition of 50 ng/mL FGF-2, the rate of organoid germination was significantly increased. The 1.5 µmol/L CHIR99021, 0.5 µmol/L Jagged-1, and 50 ng/mL FGF-2 added in the medium can cooperate with each other to improve the formation and speed up the proliferation and differentiation of organoids, while improving the stemness maintenance of cells. The morphology, cell types, and culture characteristics of chicken small intestinal organoids were studied by HE staining, transmission electron microscopy, reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR), indirect immunofluorescence, and immunohistochemistry. The results showed that the 3D organoids of the chicken small intestine cultured in vitro were morphologically consistent with the chicken intestinal tissue and contained differentiated epithelial cells. In summary, we successfully established an in vitro culture system for chicken small intestinal organoids, providing a new method for the subsequent research on chicken intestinal physiology, pathology, and host-pathogen interaction mechanism and the development of relevant drugs.
Animals ; Organoids/metabolism* ; Intestine, Small/drug effects* ; Chickens ; Cell Culture Techniques/methods* ; Culture Media ; Chick Embryo ; Tissue Culture Techniques/methods*

Objective:To analyze the tendency of elderly patients with breast cancer in the choice of treatment methods and related influencing factors.Methods:The data of 312 elderly patients with unilateral breast cancer treated in Yuncheng Central Hospital from January 2013 to December 2017 were collected for retrospective analysis. All patients' treatment options were analyzed, and the age of patients who chose different treatment options was compared. Univariate and logistic regression were used to analyze the chemotherapy choice tendency of elderly breast cancer patients, and Cox proportional risk model was used to analyze the influencing factors of 5-year survival of elderly breast cancer patients.Results:In the whole patient population, the selection rates of surgery, chemotherapy, radiotherapy, endocrine therapy and targeted therapy were respectively 97.44% (304/312), 81.41% (254/312), 7.05% (22/312), 68.27% (213/312), 3.85% (32/312). The mean age of all patients was (67.94±6.55) years. There were no statistically significant differences in the age of patients with different treatment methods (all P>0.05). The results of univariate analysis showed that, there were statistically significant differences in the depth of invasive cancer ( t=3.11, P=0.002), number of axillary lymph node metastasis ( t=6.54, P<0.001), comorbidities ( t=-4.85, P<0.001) and Eastern Cooperative Oncology Group (ECOG) score ( t=-4.56, P<0.001) between chemotherapy and non-chemotherapy patients, and there were no statistically significant differences in age ( t=-0.52, P=0.604), pathological type ( χ2=4.96, P=0.084), surgical type ( χ2=0.21, P=0.899), tumor differentiation degree ( χ2=3.28, P=0.194), estrogen receptor ( χ2=0.99, P=0.321), progesterone receptor ( χ2=0.89, P=0.346), and human epidermal growth factor receptor-2 ( χ2=0.58, P=0.445). The results of multifactor analysis showed that types of comorbidities ( OR=0.91, 95% CI: 0.85-0.99, P=0.024) and ECOG score ( OR=0.95, 95% CI: 0.92-0.99, P=0.007) were independent influencing factors for the use of chemotherapy after surgery in elderly breast cancer patients. A total of 74 patients died within 5 years after surgery, and the 5-year overall survival rate was 76.28%. More axillary lymph node metastasis ( RR=1.26, 95% CI: 1.09-1.46, P=0.001) and more complications ( RR=1.07, 95% CI: 1.02-1.13, P=0.007) were risk factors for prognosis. Conclusion:Surgery and chemotherapy are the main treatment methods for elderly patients with breast cancer. ECOG score and number of complications can directly affect the results of chemotherapy selection for such patients, the number of axillary lymph node metastasis and complications had significant influence on the long-term survival of the patients.

Objective:To investigate the clinical efficacy of ultrasound-guided vacuum-assisted excision (VAE) in the treatment of breast benign phyllode tumor and the risk factors of local recurrence after operation.Methods:A total of 256 cases of patients with breast benign phyllode tumor admitted to Yuncheng Central Hospital from January 2012 to January 2020 were included in the retrospective study, they were all female, of which 160 cases received ultrasound-guided VAE surgical treatment and 96 cases received open surgical treatment. Measurement data were expressed as mean ± standard deviation ( ± s), and t-test was used for comparison between groups. The Chi-square test or Fisher exact probability method was used to compare the data groups. Univariate analysis was performed on the factors related to the local recurrence of benign phyllode tumor after surgery, and then the factors with statistically significant differences in the univariate analysis were further incorporated into multivariate Logistic regression analysis. Results:Univariate analysis results showed that the maximum diameter of tumor ≥25 mm and the history of ipsilateral breast fibroadenoma were associated with postoperative local recurrence of breast benign phyllodes tumor ( P<0.05). Multivariate Logistic regression analysis results showed that tumor diameter ≥25 mm was an independent risk factor for postoperative local recurrence of breast benign phyllodes tumor ( P<0.05). Conclusion:Ultrasound-guided VAE in the treatment of breast benign phyllodes tumor patients with maximum diameter less than 25 mm can reduce the postoperative local recurrence rate, and patients with tumor maximum diameter greater than 25 mm should have the higher local recurrence risk.

Objective:To study whether cisplatin may induce apoptosis in the pericytes of cochlear stria vascularis and underlying mechanisms.Methods:Twenty male C57BL/6J mice aged 6-8 weeks were divided into control group and a cisplatin group. Primary cultured mouse cochlear vascular peristriatal cells were identified and divided into control group, cisplatin group, and N-acetylcysteine+cisplatin group. Auditory brainstem response (ABR) was used to detect hearing in mice. Hematoxylin eosin (HE) staining was used to observe morphological changes in the stria vascularis of the cochlea. The total superoxide dismutase (SOD) activity test kit (WST-1 method) and thiobarbituric acid (TBA) method were used to detect SOD activity and Malondialdehyde (MDA) content, respectively. DCFH-DA fluorescence probe was used to detect the content of reactive oxygen species in peripheral cells. Hoechst 33 342 and flow cytometry were used to detect the apoptosis rate of pericytes. Immunofluorescence technology was used to detect the distribution and expression of apoptosis related proteins in pericytes of cochlear stria vascularis. Immunohistochemistry and Western blotting (WB) were used to detect the expression of apoptosis-and mitochondrial-related proteins. Mito SOX TM-red and JC-1 were used to detect the mitochondrial function of pericytes. Evans blue staining was used to observe the permeability of the blood labyrinth barrier (BLB). Statistical analysis was conducted using SPSS 18.0 software. Results:Compared with the control group, the cisplatin group significantly increased in the hearing threshold ( t=4.72, P<0.01), Ⅰ-wave latency ( t=12.25, P<0.05), and the levels of oxidative stress in the cochlea and pericytes ( t=38.34, P<0.01), and also cisplatin caused disorder and contraction of the cochlear stria vascularis structure, increased BLB permeability [Evans blue leakage (1.08±0.42) AU vs (0.55±0.23) AU, t=4.64, P<0.05], with a statistically significant difference, enhanced the expressions of apoptotic proteins c-Caspase-3 ( t=5.01, P<0.01) and Bax ( t=6.33, P<0.01) in the peristriatal cells of cochlear blood vessels in mice treated with cisplatin increased. And cisplatin can induce apoptosis of primary cultured pericytes and up-regulate the expression of c-Caspase-3 and Bax ( P<0.05). The NAC+cisplatin group partially reversed cisplatin-induced pericyte apoptosis ( P<0.05). Cisplatin caused damage to the mitochondrial function of peripheral cells, and induced the release of apoptosis-inducing factor (AIF) and cytochrome C (cyt-c) into the cytoplasm ( P<0.05). The NAC+cisplatin group partially reversed cisplatin induced pericyte apoptosis ( P<0.05) and mitochondrial damage ( P<0.05). Conclusion:Cisplatin can increase the level of oxidative stress in the cochlea and cause mitochondrial pathway apoptosis in C57BL/6J mouse cochlear vascular peristriatal cells.

Objective: To investigate the role of connexin 43 ( Cx43) in acute sympathetic stress induced by phe- nylephrine (PE) overactivation of α1-adrenergic receptor ( α1-AR) in cardiomyocytes． Methods: Cardiomyocytes of H9C2 rats were randomly divided into control group，PE alone treatment group，Gap26 ( Cx43 specific inhibitor) intervention group and Gap26 alone treatment group．PE alone treatment group was treated with 50 μmol / L PE for 15 min．The Gap26 intervention group was pretreated with 0. 5 μmol / L Gap26 for 30 min，and then treated with 50 μmol / L PE for 15 min．The protein and mRNA expression levels of Cx43，NLRP3 inflammasome，interleukin-1 β (IL-1 β) ，Caspase-1，interleukin-18 (IL-18 ) were detected by Western blot and qRT-PCR. The expression and co-location of Cx43 in cardiomyocytes were observed by immunofluorescence assay，and the expression of inflamma- tory cytokines IL-1 β and IL-18 in cardiomyocytes was detected by ELISA. Results: Compared with control group， the protein and mRNA levels of Cx43，NLRP3，Caspase-1 and IL-18 in PE group increased．Compared with PE a- lone treatment group，the protein and mRNA levels of Cx43，NLRP3，Caspase-1 and IL-18 decreased after Gap26 intervention，but they were still higher than those of control group． Similarly ，immunofluorescence showed that Cx43 protein expression increased in PE alone group，while Cx43 expression was down-regulated in Gap26 inter- vention group compared with PE alone group．ELISA results showed that the expression of IL-1 β and IL-18 was sig- nificantly up-regulated in PE alone group，but down-regulated in Gap26 intervention group． Conclusion Cx43 is involved in α1-AR activation induced cardiac acute sympathetic stress by regulating NLRP3 inflammasome.

Objective:To investigate whether idebenone can improve behavioral disorders in mice with Parkinson disease (PD) by increasing PHB2 mediated mitophagy.Methods:In the first small experiment, thirty mice were randomly divided into normal control group, model group and treatment group according to the random number table method, with 10 animals in each group.The aim of this study was to observe the effect of idebenone on the behavior of Parkinson disease model mice. In the second experiment, 20 mice were randomly divided into blank control group, MPTP group, shRNA-PHB2 group and shRNA-PHB2+ MPTP group, with 5 mice in each group. The changes of tyrosine dehydrogenase (TH) in brain tissue were detected by immunofluorescence assay. In the third experiment, 30 mice were randomly divided into blank control group, shRNA-PHB2 group, MPTP+ idebenone group, shRNA-PHB2+ MPTP group, shRNA-PHB2+ idebenone group and shRNA-PHB2+ MPTP+ idebenone group, with 5 animals in each group. The aim of this study was to investigate the effect of idebenone on mitochondrial autophagy in mouse brain.C57BL-6 mice were intraperitoneally injected with MPTP to establish the animal model of chronic PD. Then 200 mg / kg idebenone was given by gavage for 21 days. And the expression of PHB2 in brain was inhibited by microinjection of adeno-associated virus 9 (AAV9) shRNA inhibin 2(PHB2) into lateral ventricle. The behavioral changes of the PD mice were detected by Morris water maze, and the changes of tyrosine dehydrogenase (TH) induced by inhibiting PHB2 were detected by immunohistochemistry. The protein expression of LC3 and PHB2 in substantia nigra of midbrain was detected by Western blot.The data were analyzed by GraphPad 7.0 and SPSS 22.0.Results:(1) In the water maze test data of the first small experiment, the repeated measurement ANOVA showed that the group-time interaction effects of latency of mice from 1 to 7 days were significant ( Ftime×group=20.51, P<0.05). Simple effect analysis showed that on the 5th, 6th and 7th day, the incubation period of the treatment group was significantly shortened (all P<0.05). Univariate analysis of variance showed that on the 7th day of the test, the differences between the control group and the model group, the model group and the treatment group, the control group and the treatment group were all statistically significant( t=-49.95, -21.81, 28.14; all P<0.01). In the third small experiment, repeated measurement analysis of variance showed that the interaction between time and group was significant ( Ftime×group=42.11, P<0.01). Simple effect analysis showed that compared with MPTP+ idebenone group, the latency of shRNA-PHB2+ MPTP+ idebenone group was significantly prolonged (all P<0.05). There were no significant difference between shRNA-PHB2+ MPTP+ idebenone group and shRNA-PHB2+ MPTP group except the 4th day ( P<0.05). On the 7th day, compared with MPTP+ idebenone group, the residence time of shRNA-PHB2+ MPTP+ idebenone group was significantly increased ( t=-34.36, P<0.001), but there was no significant difference between shRNA-PHB2+ MPTP group and shRNA-PHB2+ MPTP+ idebenone group ( t=2.94, P>0.05). (2)The results of immunofluorescence experiment showed that the relative expression of TH in the control group, model group, shRNA-PHB2 group and shRNA-PHB2+ MPTP group were (41.03±3.01), (24.20±4.18), (38.39±3.31) and (13.12±2.65), respectively. Compared with the control group, the expression of TH in the midbrain of the MPTP group was significantly down-regulated, the difference was statistically significant( t=7.98, P<0.01). Compared with the MPTP group, the expression of TH in shRNA-PHB2 group was down regulated ( t=-6.73, P<0.05). (3) Western blot results showed that the relative expression of LC3 in midbrain tissue of control group, shRNA-PHB2 group, MPTP+ idebenone group, shRNA-PHB2+ MPTP group, shRNA-PHB2+ idebenone group and shRNA-PHB2+ MPTP+ idebenone group were (0.86±0.07), (0.77±0.08), (0.42±0.05), (0.21±0.05), (0.66±0.09) and (0.27±0.07). The relative expression of PHB2 were (1.13±0.14), (0.56±0.11), (1.08±0.14), (0.27±0.07), (0.68±0.14) and (0.24±0.10). Compared with MPTP+ idebenone group, the relative expression of LC3 and PHB2 in shRNA-PHB2+ MPTP+ idebenone group was significantly decreased ( F=1.96, P<0.01). Conclusion:Idebenone can increase the level of mitophagy in PD mice through PHB2, thus improving the behavioral disorder.

Objective:To evaluate the role of silent information regulator 1 (SIRT1)/nuclear factor E2 related factor 2 (Nrf2) signaling pathway in sleep deprivation-induced cognitive dysfunction in rats.Methods:Thirty-six adult male Sprague-Dawley rats, aged 54-56 weeks, weighing 600-750 g, were divided into 3 groups ( n=12 each) using a random number table method: control group (group C), sleep deprivation group (group SD) and sleep deprivation+ SIRT1 agonist Srt1720 group (group SD+ Srt). Sleep deprivation model was established by modified multi-platform water environment method.Srt1720 10 mg/kg was injected intraperitoneally every 12 h starting from 24 h before establishing the model in group SD+ Srt, while the equal volume of 0.9% normal saline was injected intraperitoneally in C and SD groups.After the end of sleep deprivation, Morris water maze test was performed to evaluate the cognitive function.Animals were then sacrificed, and their hippocampi were removed for determination of neuronal degeneration rate in hippocampal CA1 region (using HE staining), the apoptosis rate in hippocampal CA1 (using TUNEL assay ), the expression of SIRT1 and Nrf2 in hippocampal CA1 (by immunohistochemistry) and the contents of reactive oxygen species (ROS) and superoxide dismutase (SOD) (by microplate method). Results:Compared with group C, the escape latency was significantly prolonged, the time of staying at the platform quadrant was shortened, and the frequency of crossing the platform was decreased on 2-4 days, the apoptotic rate and neuronal degeneration rate were increased, the expression of SIRT1 and Nrf2 was down-regulated, ROC content was increased, and SOD content was decreased in SD and SD+ Srt groups ( P<0.05). Compared with group SD, the escape latency was significantly shortened, the time of staying at the platform quadrant was prolonged, and the frequency of crossing the platform was increased on 3 and 4 days, the apoptotic rate and neuronal degeneration rate were decreased, the expression of SIRT1 and Nrf2 was up-regulated, ROC content was decreased, and SOD content was increased in group SD+ Srt ( P<0.05). Conclusions:Sleep deprivation can induce oxidative stress response in hippocampus by inhibiting the activation of SIRT1/Nrf2 signaling pathway, leading to cognitive dysfunction in rats.