Objective This study reports on the obligatory parasitism of water mites Arrenurus sp. on Anopheles messeae at the Manzhouli Port, Inner Mongolia, China. Methods Duing July 2024, a survey on the mosquito diversity was conducted at the Manzhouli Port. Captured mosquitoes and their ectoparasites were identified to species level. Results A total of 1840 adult mosquitoes were collected, representing species from three genera: Culex(Cx. modestus, Cx. pipiens pallens), Aedes(Ae. dorsalis, Ae. flavidorsalis, Ae. flavescens), and Anopheles (An. messeae). Among all the mosqutioes specimens,3 out of 150 captured An. messeae were found to carry ectoparasitic mites, with number of 2,4,27 mites separately. Morphological and molecular identification reached the same result as water mites(Hydrachnidiae, Hydracrina). COI gene sequence showed 94% similarity with the closest species Arrenurus truncatellus. Conlusions Literature review suggests water mites are host-specific parasitism of mosquito species and herein with the first record of Arrenurus sp. parasiting on An. Messeae in the most high-latitude region globally.

ObjectiveTo investigate the protective effect and mechanism of Shaoyaotang （SYD） on the lipopolysaccharide （LPS）-induced damage of tight junction proteins in the human colorectal adenocarcinoma （Caco-2） cell model of inflammation via the Ras homolog gene family member A （RhoA）/Rho-associated coiled-coil forming protein kinase （ROCK） pathway. MethodsCaco-2 cells were grouped as follows： Blank， model （LPS， 10 mg·L^-1）， SYD-containing serum （10%， 15%， and 20%）， and inhibitor （Fasudil， 25 μmol·L^-1）. After 24 hours of intervention， the cell viability in each group was examined by the cell-counting kit 8 （CCK-8） method. Enzyme-linked immunosorbent assay was employed to determine the levels of endothelin-1 （ET-1）， tumor necrosis factor-α （TNF-α）， interleukin-1β （IL-1β）， and interleukin-6 （IL-6）. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） and Western blot were employed to determine the mRNA and protein levels， respectively， of RhoA， ROCK2， claudin-5， and zonula occludens-1 （ZO-1） in cells of each group. ResultsCompared with the blank group， the model group showcased a marked reduction in the cell viability （P<0.01）， elevations in the levels of ET-1， TNF-α， IL-1β， and IL-6 （P<0.01）， declines in both mRNA and protein levels of ZO-1 and claudin-5 （P<0.01）， and rises in mRNA and protein levels of RhoA and ROCK2 （P<0.01）. Compared with the model group， the Shaoyaotang-containing serum （10%， 15%， and 20%） groups had enhanced cell viability （P<0.01）， lowered levels of ET-1， TNF-α， IL-1β， and IL-6 （P<0.01）， up-regulated mRNA and protein levels of ZO-1 and claudin-5 （P<0.05， P<0.01）， and down-regulated mRNA and protein levels of RhoA and ROCK2 （P<0.01）. Moreover， the inhibitor group and the 15% and 20% Shaoyaotang-containing serum groups had lower levels of ET-1， TNF-α， IL-1β， and IL-6 （P<0.05， P<0.01）， higher mRNA and protein levels of ZO-1 and claudin-5 （P<0.05， P<0.01）， and lower mRNA and protein levels of RhoA and ROCK2 （P<0.05， P<0.01） than the 10% Shaoyaotang-containing serum group. ConclusionThe Shaoyaotang-containing serum can lower the levels of LPS-induced increases in levels of inflammatory cytokines and endothelin to ameliorate the damage of tight junction proteins of the Caco-2 cell model of inflammation by regulating the expression of proteins in the RhoA/ROCK pathway.

ObjectiveTo investigate the protective effect and mechanism of Shaoyaotang （SYD） on the lipopolysaccharide （LPS）-induced damage of tight junction proteins in the human colorectal adenocarcinoma （Caco-2） cell model of inflammation via the Ras homolog gene family member A （RhoA）/Rho-associated coiled-coil forming protein kinase （ROCK） pathway. MethodsCaco-2 cells were grouped as follows： Blank， model （LPS， 10 mg·L^-1）， SYD-containing serum （10%， 15%， and 20%）， and inhibitor （Fasudil， 25 μmol·L^-1）. After 24 hours of intervention， the cell viability in each group was examined by the cell-counting kit 8 （CCK-8） method. Enzyme-linked immunosorbent assay was employed to determine the levels of endothelin-1 （ET-1）， tumor necrosis factor-α （TNF-α）， interleukin-1β （IL-1β）， and interleukin-6 （IL-6）. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） and Western blot were employed to determine the mRNA and protein levels， respectively， of RhoA， ROCK2， claudin-5， and zonula occludens-1 （ZO-1） in cells of each group. ResultsCompared with the blank group， the model group showcased a marked reduction in the cell viability （P<0.01）， elevations in the levels of ET-1， TNF-α， IL-1β， and IL-6 （P<0.01）， declines in both mRNA and protein levels of ZO-1 and claudin-5 （P<0.01）， and rises in mRNA and protein levels of RhoA and ROCK2 （P<0.01）. Compared with the model group， the Shaoyaotang-containing serum （10%， 15%， and 20%） groups had enhanced cell viability （P<0.01）， lowered levels of ET-1， TNF-α， IL-1β， and IL-6 （P<0.01）， up-regulated mRNA and protein levels of ZO-1 and claudin-5 （P<0.05， P<0.01）， and down-regulated mRNA and protein levels of RhoA and ROCK2 （P<0.01）. Moreover， the inhibitor group and the 15% and 20% Shaoyaotang-containing serum groups had lower levels of ET-1， TNF-α， IL-1β， and IL-6 （P<0.05， P<0.01）， higher mRNA and protein levels of ZO-1 and claudin-5 （P<0.05， P<0.01）， and lower mRNA and protein levels of RhoA and ROCK2 （P<0.05， P<0.01） than the 10% Shaoyaotang-containing serum group. ConclusionThe Shaoyaotang-containing serum can lower the levels of LPS-induced increases in levels of inflammatory cytokines and endothelin to ameliorate the damage of tight junction proteins of the Caco-2 cell model of inflammation by regulating the expression of proteins in the RhoA/ROCK pathway.

This study aims to investigate the molecular mechanism by which naringin alleviates cerebral ischemia/reperfusion(CI/R) injury through DRP1/LRRK2/MCU signaling axis. A total of 60 SD rats were randomly divided into the sham group, the model group, the sodium Danshensu group, and low-, medium-, and high-dose(50, 100, and 200 mg·kg~(-1)) naringin groups, with 10 rats in each group. Except for the sham group, a transient middle cerebral artery occlusion/reperfusion(tMCAO/R) model was established in SD rats using the suture method. Longa 5-point scale was used to assess neurological deficits. 2,3,5-Triphenyl tetrazolium chloride(TTC) staining was used to detect the volume percentage of cerebral infarction in rats. Hematoxylin-eosin(HE) staining and Nissl staining were employed to assess neuronal structural alterations and the number of Nissl bodies in cortex, respectively. Western blot was used to determine the protein expression levels of B-cell lymphoma-2 gene(Bcl-2), Bcl-2-associated X protein(Bax), cleaved cysteine-aspartate protease-3(cleaved caspase-3), mitochondrial calcium uniporter(MCU), microtubule-associated protein 1 light chain 3(LC3), and P62. Mitochondrial structure and autophagy in cortical neurons were observed by transmission electron microscopy. Immunofluorescence assay was used to quantify the fluorescence intensities of MCU and mitochondrial calcium ion, as well as the co-localization of dynamin-related protein 1(DRP1) with leucine-rich repeat kinase 2(LRRK2) and translocase of outer mitochondrial membrane 20(TOMM20) with LC3 in cortical mitochondria. The results showed that compared with the model group, naringin significantly decreased the volume percentage of cerebral infarction and neurological deficit score in tMCAO/R rats, alleviated the structural damage and Nissl body loss of cortical neurons in tMCAO/R rats, inhibited autophagosomes in cortical neurons, and increased the average diameter of cortical mitochondria. The Western blot results showed that compared to the sham group, the model group exhibited increased levels of cleaved caspase-3, Bax, MCU, and the LC3Ⅱ/LC3Ⅰ ratio in the cortex and reduced protein levels of Bcl-2 and P62. However, naringin down-regulated the protein expression of cleaved caspase-3, Bax, MCU and the ratio of LC3Ⅱ/LC3Ⅰ ratio and up-regulated the expression of Bcl-2 and P62 proteins in cortical area. In addition, immunofluorescence analysis showed that compared with the model group, naringin and positive drug treatments significantly decreased the fluorescence intensities of MCU and mitochondrial calcium ion. Meanwhile, the co-localization of DRP1 with LRRK2 and TOMM20 with LC3 in cortical mitochondria was also decreased significantly after the intervention. These findings suggest that naringin can alleviate cortical neuronal damage in tMCAO/R rats by inhibiting DRP1/LRRK2/MCU-mediated mitochondrial fragmentation and the resultant excessive mitophagy.
Animals ; Rats, Sprague-Dawley ; Reperfusion Injury/genetics* ; Flavanones/administration & dosage* ; Rats ; Dynamins/genetics* ; Male ; Brain Ischemia/genetics* ; Protein Serine-Threonine Kinases/genetics* ; Signal Transduction/drug effects* ; Humans ; Drugs, Chinese Herbal/administration & dosage*

OBJECTIVE: To explore the preventive and therapeutic effects of Dahuang Zhechong Pill (DZP) on pulmonary fibrosis and the underlying mechanisms. METHODS: The first key rate-limiting enzyme hexokinase 2 (HK2) of glycolysis was silenced and over-expressed through small interfering RNA and lentivirus using lung fibroblast MRC-5 cell line, respectively. The cell viability, migration, invasion and proliferation were detected by cell counting kit-8, wound healing assay, transwell assay, and flow cytometry. The mRNA and protein expression levels of HK2 were detected by RT-PCR and Western blotting, respectively. The contents of glucose, adenosine triphosphate (ATP) and lactate in MRC-5 cells were determined by enzyme-linked immunosorbnent assay (ELISA). Then, the relationship between miR-29b-2-5p and HK2 was explored by luciferase reporter gene assay. Pulmonary fibrosis cell model was induced by transforming growth factor-β 1 (TGF-β 1) in MRC-5 cells, and the medicated serum of DZP (DMS) was prepared in rats. MRC-5 cells were divided into control, TGF-β 1, TGF-β 1+10% DMS, TGF-β 1+10% DMS+miR-29b-2-5p inhibitor, TGF-β 1+10% DMS+inhibitor negative control, TGF-β 1+10% DMS+miR-29b-2-5p mimic and TGF-β 1+10% DMS+mimic negative control groups. After miR-29b-2-5p mimics and inhibitors were transfected into MRC-5 cells, all groups except control and model group were treated with DMS. The effect of DMS on MRC-5 cells were detected using aforementioned methods and immunofluorescence. Similarly, the contents of glucose, ATP and lactate in each group were measured by ELISA. RESULTS: The mRNA and protein expressions of HK2 in MRC-5 cells were successfully silenced and overexpressed through si-HK2-3 and lentiviral transfection, respectively. After silencing HK2, the mRNA and protein expressions of HK2 were significantly decreased (P<0.01), and the concentrations of glucose, ATP and lactate were also significantly decreased (P<0.05). The proliferation, migration and invasion of MRC-5 cells were significantly declined (P<0.05 or P<0.01), while the apoptosis of MRC-5 cells was significantly increased (P<0.01). After overexpressing HK2, the mRNA and protein expressions of HK2 were significantly increased (P<0.05), and the concentrations of glucose, ATP and lactate were also significantly increased (P<0.05 or P<0.01). The proliferation, migration and invasion of MRC-5 cells were significantly increased (P<0.05 or P<0.01), while the apoptosis of MRC-5 cells was significantly decreased (P<0.05). The relative luciferase activity of 3'UTR-WT+hsa-miR-29b-2-5p transfected with HK2 was significantly decreased (P<0.01). After miR-29b-2-5p mimic and inhibitor were transfected into the MRC-5 cells, DMS intervention could significantly reduce the concentration of glucose, ATP and lactate, and the mRNA and proteins expressions of HK2, phosphofructokinase and pyruvate kinase isoform M2 (P<0.05 or P<0.01). The proliferation, migration and invasion of MRC-5 cells were alleviated (P<0.05 or P<0.01), and the deposition of fibronectin, α-smooth muscle actin, and collagen I were significantly decreased (P<0.05 or P<0.01). CONCLUSIONS Glycolysis is closely related to pulmonary fibrosis. DZP reduced glycolysis and inhibited fibroblasts' excessive differentiation and abnormal collagen deposition through the miR-29b-2-5p/HK2 pathway, which played a role in delaying the process of pulmonary fibrosis.
MicroRNAs/genetics* ; Glycolysis/genetics* ; Animals ; Pulmonary Fibrosis/metabolism* ; Humans ; Drugs, Chinese Herbal/therapeutic use* ; Hexokinase/genetics* ; Cell Line ; Cell Proliferation/drug effects* ; Rats, Sprague-Dawley ; Rats ; Cell Movement/drug effects* ; Male ; Cell Survival/drug effects* ; Signal Transduction/drug effects*

OBJECTIVE: To explore the functional roles and underlying mechanisms of neferine in the context of angiotensin II (Ang II)-induced hypertension and vascular dysfunction. METHODS: Male mice were infused with Ang II to induce hypertension and randomly divided into treatment groups receiving neferine or a control vehicle based on baseline blood pressure using a random number table method. The hypertensive mouse model was constructed by infusing Ang II via a micro-osmotic pump (500 ng/kg per minute), and neferine (0.1, 1, or 10 mg/kg), valsartan (10 mg/kg), or double distilled water was administered intragastrically once daily for 6 weeks. A non-invasive blood pressure system, ultrasound, and hematoxylin and eosin staining were performed to assess blood pressure and vascular changes. RNA sequencing and network pharmacology were employed to identify differentially expressed transcripts (DETs) and pathways. Vascular ring tension assay was used to test vascular function. A7R5 cells were incubated with neferine for 24 h and then treated with Ang II to record the real-time Ca²⁺ concentration by confocal microscope. Immunohistochemistry (IHC) and Western blot were used to evaluate vasorelaxation, calcium, and the extracellular signal-regulated kinase (ERK)1/2 pathway. RESULTS: Neferine treatment effectively mitigated the elevation in blood pressure, pulse wave velocity, aortic thickening in the abdominal aorta of Ang II-infused mice (P<0.05). RNA sequencing and network pharmacology analysis identified 355 DETs that were significantly reversed by neferine treatment, along with 25 potential target genes, which were further enriched in multiple pathways and biological processes, such as ERK1 and ERK2 cascade regulation, calcium pathway, and vascular smooth muscle contraction. Further investigation revealed that neferine treatment enhanced vasorelaxation and reduced Ca²⁺-dependent contraction of abdominal aortic rings, independent of endothelium function (P<0.05). The underlying mechanisms were mediated, at least in part, via suppression of receptor-operated channels, store-operated channels, or voltage-operated calcium channels. Neferine pre-treatment demonstrated a reduction in intracellular Ca²⁺ release in Ang II stimulated A7R5 cells. IHC staining and Western blot confirmed that neferine treatment effectively attenuated the upregulation of p-ERK1/2 both in vivo and in vitro, which was similar with treatment of ERK1/2 inhibitor PD98059 (P<0.05). CONCLUSIONS Neferine remarkably alleviates Ang II-induced elevation of blood pressure, vascular dysfunction, and pathological changes in the abdominal aorta. This beneficial effect is mediated by the modulation of multiple pathways, including calcium and ERK1/2 pathways.
Animals ; Angiotensin II ; Male ; Benzylisoquinolines/therapeutic use* ; Network Pharmacology ; Blood Pressure/drug effects* ; Sequence Analysis, RNA ; Mice ; Hypertension/chemically induced* ; Mice, Inbred C57BL ; Calcium/metabolism*

Objective:To analyze the status of registration of clinical trials of radiopharmaceuticals in China, and to provide reference for the development and clinical application of radiopharmaceuticals.Methods:By searching the clinical trial registration and information disclosure platform of the Center for Drug Evaluation of the National Medical Products Administration (NMPA), the data on clinical trials of radiopharmaceuticals registered from 2014 to 2024 were collected and analyzed for trial design, administered dose, common indications, and geographical distribution.Results:A total of 77 clinical trials were included. The Compound Annual Growth Rate for the number of projects from 2014 to 2024 was 40%. Diagnostic radiopharmaceuticals were predominantly based on 18F and 99Tc m, while therapeutic radiopharmaceuticals primarily utilized 177Lu and 90Y. All indications were concentrated in the field of oncology. Regarding trial design, non-randomized (71.4%), open-label (89.6%), and single-arm (66.2%) trials accounted for the highest proportions. Geographical distribution showed Beijing (29 trials), Shanghai (18 trials), and Jiangsu province (14 trials) as the regions with the highest concentration of clinical trials. Conclusions:Radiopharmaceutical clinical trials in China have shown rapid growth. However, research remains predominantly focused on oncology, with a relatively high proportion of early-stage trials. In order to fully utilize the potentials of radiopharmaceuticals and improve the quality of clinical trials, nuclear medicine researches should broaden therapeutic applications, implement prudently administerd dose in clinical trials, and implement optimized radiation protection procedures across all clinical trial centers.

Aim To investigate the intervention effect of Rehmannia radix water extract on bleomycin(BLM)-induced pulmonary fibrosis in mice combined with metabolomics and to reveal the potential mechanism,in order to provide new ideas for clinical treatment of pul-monary fibrosis.Methods Male C57BL/6N mice were randomly divided into the control group,model group,pirfenidone group(positive control,PFD,270 mg·kg-1),and low dose(DH-L,4.55 g·kg-1)group,medium dose(DH-M,9.1 g·kg-1)group and high dose(DH-H,18.2 g·kg-1)group of Rehman-nia.Except for the control group,BLM(5 mg·kg-1)was instilled into the trachea to establish the model of pulmonary fibrosis in the other groups.The survival rate,lung index and blood oxygen saturation of mice in each group were evaluated.HE and Masson staining were used to observe the pathological changes of lung tissue.WBP was used to detect lung function.Flow cytometry was used to detect the apoptosis of primary lung cells,ROS and immune cells.ELISA was used to detect the levels of fibrosis markers and inflammatory factors(α-SMA,collagen Ⅰ,collagen Ⅲ,TGF-β1,TNF-α,IL-1 β,and IL-6).Biochemical method was employed to detect the contents of GSH-Px,T-SOD and MDA.Liquid chromatograph mass spectrometer(LC-MS)metabolomics was used to analyze the changes of serum metabolic profile.Results Water extract of Re-hmannia significantly increased the survival rate,oxy-gen saturation and lung function of mice with pulmona-ry fibrosis,reduced the lung coefficient,ameliorated pathological damage and collagen deposition in lung tissue,reduced the levels of apoptosis and oxidative stress,and down-regulated the levels of inflammatory factors in lung tissue.It regulated the levels of metabo-lites such as bile acid metabolism,sphingolipid metabo-lism,and unsaturated fatty acid metabolism.Conclu-sions Water extract of Rehmannia inhibits lung injury and collagen deposition in mice with pulmonary fibrosis by inhibiting inflammatory response,which may be a-chieved by regulating the levels of inflammatory factors through the metabolic pathways of bile acid and sphin-golipid.