Objective To investigate the changes in the prevalence of Babesia microti infections, spleen morphology and proportions of splenic immune cells in BALB/c mice following intravenous and intraperitoneal injections, so as to provide insights into unraveling the immune regulatory mechanisms of Babesia infections. Methods Laboratory - maintained B. microti strains were prepared into whole blood samples with 10% prevalence of B. microti infection. A total of 75 BALB/c mice were randomly divided into three groups, including the normal control group, intravenous injection group, and intraperitoneal injection group, of 25 mice in each group. Mice in the intravenous and intraperitoneal injection groups were administered 100 μL of whole blood samples with 10% prevalence of B. microti infection, with the day of injection recorded as d0, and animals in the normal control group were given no treatments. Blood was sampled from mice in each group via the tail tip on d7, d14, d21, d28 and d35, and prepared into thin-film blood smears, and B. microti infection was observed in red blood cells. Five mice were randomly sampled from each group and sacrificed on d7, d14, d21, d28 and d35, and spleen was collected for measurement of spleen size and weight. In addition, splenic cells were isolated, and the proportions of CD3e⁺ T cells, CD45R⁺ B cells, CD49b⁺ nature killer (NK) cells, and F4/80⁺ macrophages were detected in CD45⁺ lymphocytes using flow cytometry. Results The prevalence of B. microti infection in the intravenous (22.80%) and intraperitoneal injection groups (44.82%) peaked on d7 (χ² = 8.141, P < 0.01) and then rapidly decreased, and no parasites were observed on d35. The longest mouse spleen length [(32.91 ± 2.20) mm] and width [(9.82 ± 0.43) mm], and the greatest weight [(0.78 ± 0.10) g] were found on d14 in the intravenous injection group, and the longest spleen length [(32.42 ± 3.21) mm] and width [(10.25 ± 0.73) mm], and the greatest weight [(0.73 ± 0.09) g] were seen in the intra-peritoneal injection group on d21, d7 and d14, respectively. There were significant differences among the intravenous injection group, intraperitoneal injection group and the normal control group in terms of spleen length (F = 10.310, P < 0.05), width (F = 9.824, P < 0.05), and weight (F = 10.672, P < 0.05) on d21, and the mouse spleen length, width and weight were all significantly greater in the intraperitoneal injection group than in the intravenous injection group (allP values < 0.05). The proportions of splenic CD3e⁺ T cells [(60.60 ± 6.20)% and (39.68 ± 7.62)%], CD45R⁺ B cells [(43.32 ± 2.08)% and (49.53 ± 4.90)%], CD49b⁺ NK cells [(6.88 ± 1.34)% and (7.71 ± 1.59)%], and F4/80⁺ macrophages [(2.21 ± 0.29)% and (3.80 ± 0.35)%] peaked on d14, d21, d21 and d14 in the intravenous and intraperitoneal injection groups, respectively. There were significant differences in the proportions of CD3e⁺ T cells (F = 16.730, P < 0.05) and F4/80⁺ macrophages (F = 15.941, P < 0.05) among the intravenous injection group, intraperitoneal injection group and normal control group on d14, and a higher proportion of CD3e⁺ T cells and a lower proportion of F4/80⁺ macrophages were detected in the intravenous injection group than in the intraperitoneal injection group (both P values < 0.01). There were significant differences among the intravenous injection group, intraperitoneal injection group and normal control group on d21 in terms of proportions of splenic CD3e⁺ T cells (F = 9.252, P < 0.05), CD45R⁺ B cells (F = 14.349, P < 0.05), CD49b⁺ NK cells (F = 13.436,P < 0.05), and F4/80⁺ macrophages (F = 8.180, P < 0.05), and a higher proportion of CD3e⁺ T cells and lower proportions of CD45R⁺ B cells and F4/80⁺ macrophages were detected in the intravenous injection group than in the intraperitoneal injection group (all P values < 0.01). In addition, there was a significant difference in the proportion of CD3e⁺ T cells among the intravenous injection group, intraperitoneal injection group and normal control group on d28 (F = 9.772,P < 0.05), and a lower proportion of CD3e⁺ T cells was found in the intravenous injection group than in the intraperitoneal injection group (P < 0.01). Conclusions Both intraperitoneal and intravenous routes are effective to induce B. microti infections in BALB/c mice, and the prevalence of B. microti infections is higher in BALB/c mice through the intraperitoneal route than through the intravenous route. Intraperitoneal and intravenous injections with B. microti cause diverse spleen morphologies and proportions of splenic immune cells in mice, indicating routes of B. microti infections cause different impacts on immune response mechanisms in mice.

ObjectiveTo understand the infection characteristics of multidrug-resistant organisms (MDROs) in hospitalized patients in a tertiary sentinel hospital in Shanghai, so as to provide an evidence for the development of targeted prevention and control measures. MethodsData of MDROs strains and corresponding medical records of some hospitalized patients in a hospital in Shanghai from 2021 to 2023 were collected, together with an analysis of the basic information, clinical treatment, underlying diseases and sources of sample collection. ResultsA total of 134 strains of MDROs isolated from hospitalized patients in this hospital were collected from 2021 to 2023 , including 63 strains of methicillin-resistant Staphylococcus aureus (MRSA), 57 strains of carbapenem-resistant Acinetobacter baumannii (CRAB), and 14 strains of carbapenem-resistant Klebsiella pneumoniae (CRKP). Of the 134 strains, 30 strains were found in 2021, 47 strains in 2022 and 57 strains in 2023. The male-to-female ratio of patients was 2.05∶1, with the highest percentage (70.90%) in the age group of 60‒<90 years. The primary diagnosis was mainly respiratory disease, with lung and respiratory tract as the cheif infection sites. There was no statistically significant difference in the distribution of strains between different genders and infection sites (P>0.05). However, the differences in the distribution of strains between different ages and primary diagnosis were statistically significant (P<0.05). Patients who were admitted to the intensive care unit (ICU), had urinary tract intubation, were not artery or vein intubated, were not on a ventilator, were not using immunosuppresants or hormones, and were not applying radiotherapy or chemotherapy were in the majority. There was no statistically significant difference in the distribution of strains for whether received radiotherapy or chemotherapy or not (P>0.05), while the differences in the distribution of strains with ICU admission history, urinary tract intubation, artery or vein intubation, ventilator use, and immunosuppresants or hormones use or not were statistically significant (all P<0.05). The type of specimen was mainly sputum, the hospitalized ward was mainly comprehensive ICU, the sampling time was mainly in the first quarter throughout the year, the number of underlying diseases was mainly between 1 to 2 kinds, the application of antibiotics ≥4 kinds, and those who didn’t receive any surgery recently accounted for the most. There were statistically significant differences in the distribution of strains between different specimen types, wards occupied and history of ICU stay (P<0.05), but no statistically significant difference in the distribution of strains between different sampling times, number of underlying diseases and types of antibiotics applied (P>0.05). ConclusionThe situation of prevention and control on MDROs in this hospital is still serious. Focus should be placed on high-risk factors’ and infection monitoring and preventive measures should be strengthened to reduce the incidence rate of MDROs infection.

OBJECTIVE: To determine the role of Tripterygium wilfordii multiglycoside (TGW) in the treatment of psoriatic dermatitis from a cellular immunological perspective. METHODS: Mouse models of psoriatic dermatitis were established by imiquimod (IMQ). Twelve male BALB/c mice were assigned to IMQ or IMQ₊TGW groups according to a random number table. Histopathological changes in vivo were assessed by hematoxylin and eosin staining. Ratios of immune cells and cytokines in mice, as well as PAM212 cell proliferation in vitro were assessed by flow cytometry. Pro-inflammatory cytokine expression was determined using reverse transcription quantitative polymerase chain reaction. RESULTS: TGW significantly ameliorated the severity of IMQ-induced psoriasis-like mouse skin lesions and restrained the activation of CD45⁺ cells, neutrophils and T lymphocytes (all P<0.01). Moreover, TGW significantly attenuated keratinocytes (KCs) proliferation and downregulated the mRNA levels of inflammatory cytokines including interleukin (IL)-17A, IL-23, tumor necrosis factor α, and chemokine (C-X-C motif) ligand 1 (P<0.01 or P<0.05). Furthermore, it reduced the number of γ δ T17 cells in skin lesion of mice and draining lymph nodes (P<0.01). CONCLUSIONS TGW improved psoriasis-like inflammation by inhibiting KCs proliferation, as well as the associated immune cells and cytokine expression. It inhibited IL-17 secretion from γ δ T cells, which improved the immune-inflammatory microenvironment of psoriasis.
Male ; Animals ; Mice ; Tripterygium ; Psoriasis/drug therapy* ; Keratinocytes ; Skin Diseases/metabolism* ; Cytokines/metabolism* ; Imiquimod/metabolism* ; Dermatitis/pathology* ; Disease Models, Animal ; Mice, Inbred BALB C ; Skin/metabolism*

目的：探讨苍术素（ATR）通过调节受体相互作用蛋白激酶（RIPK）1/RIPK3/混合谱系激酶结构域样（MLKL）信号通路对非小细胞肺癌（NSCLC）A549细胞程序性死亡及裸鼠移植瘤生长的影响。方法：使用0~160 μmol/L的ATR处理A549细胞，MTT法检测细胞存活率以确定后续实验给药浓度。使用ATR和/或RIPK1抑制剂Nec-1（necrostatin-1）、caspase抑制剂Z-VAD-FMK处理A549细胞，验证ATR是否诱导A549细胞发生程序性坏死。将A549细胞分为对照组、ATR-L组、ATR-M组、ATR-H组（分别用0、10、20、40 μmol/L ATR处理）、ATR+Nec-1组（40 μmol/L ATR+50 μmol/L Nec-1处理），处理24 h后，采用PI单染及Hoechst33342/PI双染法检测细胞死亡情况、透射电镜观察细胞死亡形态、DCFH-DA荧光探针法检测细胞内ROS水平、JC-1染色法检测线粒体膜电位、WB法检测细胞中RIPK1/RIPK3/MLKL信号通路相关蛋白质的表达水平。构建A549细胞裸鼠移植瘤模型，用10 mg/kg ATR（溶于玉米油中）对裸鼠灌胃给药5周，观察ATR对移植瘤生长的影响，WB法检测移植瘤组织中RIPK1/RIPK3/MLKL信号通路相关蛋白质的表达水平。结果：10~160 μmol/L的ATR可显著抑制A549细胞增殖，选择10、20、40 μmol/L的ATR进行后续实验。ATR组A549细胞存活率显著低于对照组（P<0.01）和ATR+Nec-1组（P<0.01），而ATR+z-VAD组细胞存活率显著低于z-VAD组（P<0.01），说明ATR可诱导A549细胞发生程序性坏死而非凋亡。与对照组比较，ATR处理组A549细胞发生肿胀，线粒体内脊消失呈空泡化，细胞内容物向外泄漏，细胞核聚集，表现为坏死特征，ATR-L组、ATR-M组、ATR-H组A549细胞死亡率、ROS水平及p-RIPK1、p-RIPK3、p-MLKL表达水平均显著升高，线粒体膜电位显著降低（均P<0.01），且呈药物浓度依赖性；与ATR-H组比较，ATR+Nec-1组细胞死亡率、ROS及p-RIPK1、p-RIPK3、p-MLKL表达水平降低，线粒体膜电位显著升高（均P<0.01）。裸鼠移植瘤实验结果显示，与对照组比较，ATR组裸鼠移植瘤体积、移植瘤质量均降低（P<0.05，或P<0.01），而与瘤组织中p-RIPK1、p-RIPK3、p-MLKL蛋白表达水平均显著升高（均P<0.01）。结论：ATR可能通过激活RIPK1/RIPK3/MLKL信号通路诱导A549细胞发生程序性坏死，抑制A549细胞及其裸鼠移植瘤的生长。

To establish and optimize a method for the detection of recombinant human midkine (rhMK) activity and verify its methodology, cell counting kit-8 (cck-8) method was used to measure the proliferation activity of rat knee chondrocytes. The specificity, accuracy, precision, linearity and robustness of the method were also verified in this study. The established method was proven to have good specificity because the buffer of rhMK and recombinant human interleukin-1 receptor antagonist have no obvious active effect; the recoveries of the samples with relative activities of 50%, 75%, 100%, 125%, 150% were in the range of 80.0% to 124.0% by statistical analysis, the relative standard deviations (RSD) of relative potency were all within 20%, the linear correlation coefficient, R² ≥ 0.98, suggesting that the accuracy, precision and linearity of the method were good; the robustness correlation coefficient, R² ≥ 0.92 and the ratio of maximum to minimum of sigmoidal dose-response were no less than 1.5, indicating that robustness of the methods was good. In conclusion, a bioactivity measurement method for rhMK was established and fully validated in this study and it provides a reliable method for the bioactivity analysis of rhMK routine samples during the development. This study was approved by the Animal Care and Use Committee of Shanghai Model Organisms Center, Inc. (approval number: 2019-0008-06).

Objective To establish a highly sensitive,stable,and universally applicable ultra-high-performance liquid mass spectrometry tandem method(UPLC-MS/MS)for simultaneous determination of nirmatrelvir and ritonavir blood concentrations in human plasma.Methods The separation was performed on an ACQUITY UPLC BEH C18 column(2.1 mm× 50 mm,1.7 μm)with gradient elution,and the mobile phase consisted of 0.1％formic acid-water and 100％acetonitrile at the flow rate of 0.3 mL·min-1.The column temperature was 45℃,and the injection volume was 2 μL.Electrospray ionization as ion source(ESI+)was used as the ion source and multiple reactions monitoring mode(nirmatrelvir m/z 500.20→319.10,nirmatrelvir-D9 m/z 508.59→328.10,ritonavir m/z 721.30→426.10,13C,2H3-ritonavir m/z 725.30→426.10)was adopted.Thirty patients with coronavirus disease 2019(COVID-19)treated with nirmatrelvir and ritonavir at the People's Hospital of Changxing County in Jan.2023 were selected to measure their steady-state trough concentrations of nirmatrelvir and ritonavir after 3 days of treatment.Results The linear range of nirmatrelvir was 0.100-10.0 μg·mL-1(R2=0.997 2),and the linear range of nirmatrelvir was 0.050-5.00 μg·mL-1(R2=0.995 2).The recovery rates of nirmatrelvir and lopinavir were both＞90％and the intra-batch and inter-batch precision relative standard deviations(RSDs)were both＜10％.Additionally,the recovery ranges for nirmatrelvir and lopinavir were 91.5％-97.0％,and the matrix effects ranged from 92.4％to 97.7％.The results of clinical samples showed that the plasma concentrations of nirmatrelvir and ritonavir in patients with COVID-19 varied greatly among individuals.Conclusion The method for simultaneous determination of nirmatrelvir and ritonavir concentrations in human plasma established in this study is convenient,highly specific,highly accurate,with high precision,which is suitable for monitoring the concentrations of nirmatrelvir and ritonavir in patients.

OBJECTIVE To provide a reference for pharmaceutical care in patients with hemorrhagic transformation after cerebral infarction complicated with atrial fibrillation. METHODS Clinical pharmacists participated in the treatment practice of a patient with hemorrhagic transformation after cerebral infarction complicated with atrial fibrillation. Because the patient had a hemorrhagic transformation after cerebral infarction after stent implantation and arterial thrombolysis， the clinical pharmacists recommended stopping antiplatelet drugs and giving plasma and cold precipitation； because D-dimer was significantly elevated， the clinical pharmacists recommended anticoagulant therapy with low-molecular heparin. Due to the patient’s recurrence of hemorrhagic transformation after cerebral infarction， the clinical pharmacists recommended discontinuing rivaroxaban and administering human prothrombin complex concentrate. RESULTS The physician adopted the clinical pharmacists’ recommendation. After treatment， the patient’s condition tended to improve steadily and was allowed to be discharged with medication. CONCLUSIONS The clinical pharmacists assessed the individualized risk and optimized the patient’s medication regimen by suggesting discontinuation of antiplatelet and anticoagulant drugs， carrying out pharmaceutical care such as coagulation index monitoring， determining the time and indications for restarting anticoagulation， and pharmaceutical monitoring， to ensure the safety and efficacy of the patient’s medication.

CRISPR/Cas system, an adaptive immune system with clustered regularly interspaced short palindromic repeats, may interfere with exogenous nucleic acids and protect prokaryotes from external damages, is an effective gene editing and nucleic acid detection tools. The CRISPR/Cas system has been widely applied in virology and bacteriology; however, there is relatively less knowledge about the application of the CRISPR/Cas system in parasitic diseases. The review summarizes the mechanisms of action of the CRISPR/Cas system and provides a comprehensive overview of their application in gene editing and nucleic acid detection of parasitic diseases, so as to provide insights into future studies on parasitic diseases.

Multisystem inflammatory syndrome in children(MIS-C)is a complex syndrome characterized by multi-organ involvement that has emerged in the context of the severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)outbreak.The clinical presentation of MIS-C is similar to Kawasaki disease but predominantly presents with fever and gastrointestinal symptoms,and severe cases can involve toxic shock and cardiac dysfunction.Epidemiological findings indicate that the majority of MIS-C patients test positive for SARS-CoV-2 antibodies.The pathogenesis and pathophysiology of MIS-C remain unclear,though immune dysregulation following SARS-CoV-2 infection is considered a major contributing factor.Current treatment approaches for MIS-C primarily involve intravenous immunoglobulin therapy and symptomatic supportive care.This review article provides a comprehensive overview of the definition,epidemiology,pathogenesis,clinical presentation,diagnosis,treatment,and prognosis of MIS-C.

ObjectiveTo investigate the status of liver function abnormalities in patients with severe mental disorder (SMD) in Shanghai community, to explore the related factors to abnormal liver function in patients with SMD, and to analyze the effects of the types of mental disorders and medication status on liver function abnormalities. MethodsThe patients with SMD in Jinshan District, Minhang District, Hongkou District and Xuhui District of Shanghai were selected as the research subjects. Questionnaire survey, physical examination and laboratory tests were conducted to obtain their demographic characteristics and liver function indicators such as alanine aminotransferase (ALT) and total bilirubin (TBil). The types of mental disorders and medication status of patients in Jinshan District were also investigated. Abnormalities in liver function were determined by abnormalities in either ALT or TBil. Binary logistic regression analysis was used for multivariate analysis of the status of abnormal liver function, and the effects of mental disorder types and medication status on liver function were analyzed, simultaneously. ResultsA total of 7 251 patients with SMD were finally included into this study, and the rate of liver function abnormality was 22.7%, of which 694 cases (9.6%) had ALT abnormality and 1 084 cases (14.9%) had TBil abnormality. Univariate analysis showed that the rate of liver function abnormalities was higher in males than that in females (χ²=45.026, P<0.001), higher in suburbs than that in urban areas (χ²=25.317, P<0.001), higher in those with higher BMI than in those with lower BMI (χ²=63.748, P<0.001), higher in those with elevated blood pressure (BP) than in those without elevated BP (χ²=24.774, P<0.001), higher in those with elevated blood glucose than in those without elevated blood glucose (χ²=43.345, P<0.001), higher in those with abnormal triglyceride (TG) than in those with normal TG (χ²=15.551,P<0.001), and higher in those with abnormal total cholesterol (TC) than in those with normal TC (χ²=10.962, P=0.001). Multivariate analysis showed that the rate of abnormal liver function was higher in males than that in females (OR=1.53, 95%CI: 1.36‒1.73), higher in suburbs than that in urban areas (OR=1.43, 95%CI: 1.21‒1.70), higher in those with overweight than in those with normal BMI (OR=1.16, 95%CI: 1.01‒1.33), higher in those with obesity than in those with those with normal BMI (OR=1.61, 95%CI: 1.36‒1.91), higher in those with elevated blood glucose than in those without elevated blood glucose (OR=1.39, 95%CI: 1.23‒1.58), and higher in those with abnormal TC than in those with normal TC (OR=1.36, 95%CI: 1.13‒1.65).The difference in the rate of ALT abnormalities among the SMD patients in Jinshan District with different medication status was statistically significant (χ²=21.928, P<0.001). Whereas, the differences in the effects of the types of mental disorders and medication status on the status of liver function abnormalities were not statistically significant(P>0.05). ConclusionThe detection rate of liver function abnormalities in community-based SMD patients in Shanghai is high, and male, suburban, and accompanied by elevated BMI, BP, blood glucose, and TC are risk factors for liver function abnormalities in patients with SMD. Primary healthcare providers should pay more attention to the liver function of patients with SMD and initiate targeted and tailored prevention, detection and treatment measures.