Hypoxic-ischemic (HI) brain damage poses a high risk of death or lifelong disability, yet effective treatments remain elusive. Here, we demonstrated that miR-100-5p levels in the lesioned cortex increased after HI insult in neonatal mice. Knockdown of miR-100-5p expression in the brain attenuated brain injury and promoted functional recovery, through inhibiting the cleaved-caspase-3 level, microglia activation, and the release of proinflammation cytokines following HI injury. Engineered extracellular vesicles (EVs) containing neuron-targeting rabies virus glycoprotein (RVG) and miR-100-5p antagonists (RVG-EVs-Antagomir) selectively targeted brain lesions and reduced miR-100-5p levels after intranasal delivery. Both pre- and post-HI administration showed therapeutic benefits. Mechanistically, we identified protein phosphatase 3 catalytic subunit alpha (Ppp3ca) as a novel candidate target gene of miR-100-5p, inhibiting c-Fos expression and neuronal apoptosis following HI insult. In conclusion, our non-invasive method using engineered EVs to deliver miR-100-5p antagomirs to the brain significantly improves functional recovery after HI injury by targeting Ppp3ca to suppress neuronal apoptosis.
Animals ; MicroRNAs/metabolism* ; Extracellular Vesicles/metabolism* ; Mice ; Recovery of Function/physiology* ; Hypoxia-Ischemia, Brain/therapy* ; Mice, Inbred C57BL ; Antagomirs/administration & dosage* ; Male ; Animals, Newborn ; Apoptosis/drug effects* ; Brain Injuries/metabolism* ; Glycoproteins ; Peptide Fragments ; Viral Proteins

Objective To explore the effects of transcutaneous electrical acupoint stimulation(TEAS)combined with multi-model analgesia on infrared thermal imaging characteristics and pain after total knee arthroplasty(TKA).Methods A total of 74 patients with TKA were divided into the treatment group and the control group according to random number table method,with 37 cases in each group.The control group was treated with multi-model analgesia,and the treatment group was treated with TEAS on the basis of multi-model analgesia for 30 min,once in the morning and afternoon before the patient's rehabilitation exercise 1-7 days after surgery.The infrared thermal imaging data,visual analogue scale(VAS)score,Western Ontario and McMaster Universities Osteoarthritis Index(WOMAC)score and pain threshold of the two groups were recorded and analyzed before and at different time points after operation.Results There were 2 cases dropped out in each group.Compared with before surgery,both groups had an increase in knee joint temperature on the surgical side 14 days after surgery(P<0.05),and the temperature in the treatment group was lower than that in the control group(P<0.05).The self knee temperature difference in the treatment group was lower than that in the control group 14 days after surgery(P<0.05).Compared with before surgery,the VAS score of the treatment group decreased 3 and 7 days after surgery(P<0.05),while the VAS score of the control group decreased 7 days after surgery(P<0.05);compared with the control group at 3 and 7 days after surgey,the VAS score of the treatment group was lower than that of the control group.Compared with before surgery,the WOMAC scores of both groups increased 7 days after surgery(P<0.05);after 7 days of surgery,the WOMAC score of the treatment group was lower than that of the control group(P<0.05).Compared with before surgery,the pain threshold values of both groups increased 7 days after surgery(P<0.05);compared with the control group at 3 and 7 days after surgery,the pain threshold values of the treatment group was higher than that of the control group(P<0.05).Conclusion TEAS combined with multi-model analgesia can reduce the temperature of the knee joint,relieve the pain of the operated limb,and promote the early functional recovery of the knee joint after TKA.Infrared thermal imaging technology has a certain application value in indirectly assessing the degree of postoperative pain and detecting early infection of the affected limb.

Objective To explore the treatment effects and mechanism of Fangji Huangqi Xiaozhong Prescription in metabolic syndrome phenotype osteoarthritis(MS-OA)based on PPARγ/NF-κB signaling pathway.Methods SD rats were randomly divided into sham-operation group,OA group,MS-OA group,Western medicine group,and TCM high-and low-dasage groups.The modified Hulth method was used to make the OA model,and OA model was added with high-carbohydrate high-fat diet to make the MS-OA model.TCM high-and low-dosage groups were given 15.12,7.56 g/kg Fangji Huangqi Xiaozhong Prescription for gavage.The Western medicine group was given 16.2 mg/kg of losoprofen sodium by gavage,while the other groups were given physiological saline by gavage once a day for 6 consecutive weeks.Rat body mass was measured,biochemical detection of blood lipids and blood glucose was conducted,ELISA was used to detect the contents of serum TNF-α,IL-1β,IL-10 and leptin,morphological changes in cartilage tissue were observed using safranin O-fixed green and HE staining,immunohistochemical staining was used to detect expressions of Acan,ColⅩ,MMP13,TNF-α,IL-1β and PPARγ in cartilage tissue,Western blot was used to detected the expression of PPARγ,NF-κBp65 and p-NF-κBp65 protein in cartilage tissue.Results Compared with the sham-operation group,body mass and serum TC,TG,LDL-C,TNF-α,IL-1β and leptin of MS-OA group increased significantly(P<0.01),the contents of HDL-C and IL-10 decreased(P<0.01),cartilage tissue degeneration was significant,and the Mankin score increased(P<0.01),the expression of ColⅩ,MMP13,TNF-α,IL-1β,p-NF-κBp65 protein increased(P<0.05,P<0.01),and the expression of Acan and PPARγ protein decreased(P<0.01).Compared with the MS-OA group,the contents of serum TC,TG,LDL-C,TNF-α and leptin decreased in TCM high-dosage group(P<0.05,P<0.01),the content of IL-10 increased(P<0.05),the pathological damage of cartilage tissue improved,the Mankin score decreased(P<0.01),the expressions of ColⅩ,MMP13,TNF-α,IL-1β and p-NF-κBp65 protein in cartilage tissue decreased(P<0.05,P<0.01),and the protein expressions of Acan and PPARγ protein increased(P<0.01,P<0.05).Conclusion Fangji Huangqi Xiaozhong Prescription can improve lipid metabolism disorder,improve intra-articular inflammatory environment,balance cartilage metabolism,and delay cartilage degeneration in MS-OA rats.Its mechanism may be related to the regulation of PPARγ/NF-κB signaling pathway.

Objective:To establish a rapid quantitative PCR (qPCR) technique for Mycobacterium marinum skin infections, and to analyze its clinical diagnostic efficiency. Methods:DNA was extracted from Mycobacterium marinum colonies and serially diluted (10 -1 to 10 -8). Twelve pairs of previously reported primers and probes, as well as 6 pairs of newly designed primers and probes in this study, were used for qPCR amplification to identify the most sensitive primers and probes for the detection of Mycobacterium marinum. Skin lesion tissues were collected from 72 patients with confirmed Mycobacterium marinum infections (experimental group) and 68 with other mycobacterial infections (control group) at Shandong Provincial Hospital for Skin Diseases & Shandong Provincial Institute of Dermatology and Venereology, Shandong First Medical University & Shandong Academy of Medical Sciences in 2021. These skin tissues were subjected to qPCR amplification, interferon-gamma release assay (IGRA), acid-fast staining, and tissue culture to evaluate the diagnostic efficacy. Results:The newly designed primers and probes targeting the mycobacterial enhanced infection locus 2 (Mel2) demonstrated the highest sensitivity, with a detection limit of 0.86 copies/μl (cycle threshold value = 37) ; the qPCR amplification with the Mel2 primers/probes did not yield positive results when used for the detection of other mycobacteria (including Mycobacterium leprae and Staphylococcus spp) . Among the 72 patients in the experimental group, 44 were positive for qPCR with a sensitivity of 61.1% (95% CI: 49.6% - 71.5%), and 47 were positive for culture with a sensitivity of 65.2% (95% CI: 53.8% - 75.3%) ; all the 68 controls were negative for both qPCR and culture, with their specificities both being 100%. Among 65 patients subjected to IGRA, 31 were positive with a sensitivity of 47.7% (95% CI: 36.0% - 59.6%), while 16 out of 25 controls were negative for IGRA with a specificity of 64.0% (95% CI: 44.5% - 79.8%). Among 58 patients subjected to acid-fast staining, 37 were positive with a sensitivity of 63.8% (95% CI: 50.9% - 74.9%), and 52 out of 66 controls were negative for acid-fast staining with a specificity of 78.8% (95% CI: 67.5% - 86.9%). The combination of qPCR and culture resulted in a sensitivity of 93% and a specificity of 100% for the detection of Mycobacterium marinum. Conclusion:In this study, a highly sensitive qPCR assay was developed for the detection of Mycobacterium marinum, and its combination with culture could further improve the detection sensitivity.

Objective Network pharmacology,molecular docking,metabolomics and experimental verification were used to investigate the effect of Gegen Decoction on epileptic seizure under cerebral hypoxia.Methods The core targets of Gegen Decoction,epilepsy and cerebral hypoxia were screened by network pharmacology,and the correlation between ligand and receptor was verified by molecular docking.Further animal experiments were carried out to verify the effect of Gegen Decoction on the rat model with epilepsy accompanied by cerebral hypoxia,which was established by lithium chlorine-pilocarpine method combined with unilateral common carotid artery ligation and placing in a closed chamber with oxygen content of 10%-15%.Sixty rats were randomly divided into blank group,model group,phenytoin sodium(26 g·kg-1)group,high-,medium-and low-dose(10.62,5.31,2.655 g·kg-1)Gegen Decoction groups.After treatment,the number of epileptic seizures rats in each group was counted,and the recovery of nerve function and epileptic seizures were evaluated.HE staining was used to observe the morphological changes of the CA1 region of hippocampus in rats.The expression of interleukin-6(IL-6)and interleukin-17(IL-17)were detected by immunohistochemistry.The changes of brain neurotransmitters were analyzed by targeted metabolomics.Results Network pharmacological results suggested that 128 active components and 228 targets were obtained.A PPI network map of drug-disease was constructed by STRING.The results showed that STAT3,IL-6 and AKT1 were the core targets.GO enrichment for biological processes(BP)involves responses to external irritant,oxygen levels,hypoxia and metabolic processes of reactive oxygen species.Cell components(CC)mainly involve in the impact on cell membrane and protein kinase complex,while molecular function(MF)involves in influence on the binding of ubiquitin-like protein and ubiquitin-like protein ligase.The key pathways of Gegen Decoction in the treatment of hypoxic-induced epilepsy in the brain,which were analyzed by KEGG,involved lipid and atherosclerosis signaling pathway,IL-17 signaling pathway,Th-17 signaling pathway and so on.The experimental results showed that the number of seizures in the Gegen Decoction group was significantly decreased compared with that in the model group(P＜0.01),the morphology and structure of hippocampal cells were improved to varying degrees,and cell vacuolation was reduced.The expression of IL-17A and IL-6 in all treatment groups was lower than those in model group,and there was statistical significance in medium-dose Gegen Decoction group(P＜0.01,P＜0.05).Brain metabolomics test showed that compared with model group,monoamine neurotransmitters including 5-hydroxytryptophan(5-HTP)and norepinephrine(NE)were increased in high-dose Gegen Decoction group,while tryptophan metabolite kynurenine(Kyn)was decreased.Conclusion Gegen Decoction can reduce the levels of serum inflammatory factors IL-6 and IL-17A in epileptic rats under hypoxia.Moreover,Gegen Decoction can reduce epileptic seizures by improving hypoxia-induced inflammatory response,increasing the content of monoamine neurotransmitter(5-HTP and NE),as well as reducing the content of tryptophan metabolite Kyn and the toxin of tryptophan metabolism pathway.

bstract: Objective To elucidate the distribution characteristics of mosquito species and the population genetic diversity of important mosquito species in the Wuyi Mountain Nature Reserve, with the aim of providing a scientific basis for the prevention and control of mosquito-borne diseases in the region. Methods Within Wuyi Mountain Nature Reserve 2022- 2023, 16 sampling sites were selected, where larvae were collected using the pipette method and adult mosquitoes were collected using the mosquito trap lamp method. Mosquito species identification was accomplished by integrating morphological characteristics and molecular identification techniques based on CO Ⅰ and CO Ⅱ gene sequences. Software tools such as ClustalX, DnaSP v5.0, and Network v4.6.1.0 were utilized to analyze the genetic differentiation (Fst), gene flow (Nm) among populations of Aedes japonicus, Aedes albopictus and Armigeres subalbatus. Results The mosquito specimens collected from the Wuyi Mountain Nature Reserve were identified as 30 species belonging to 8 genera within the Culicidae family. Among them, there were 10 species of Aedes, 13 species of Culex, 1 species each of Armigeres, Orthopodomyia, Uranotaenia, Tripyeroides, and Anopheles, and 2 species of Toxorhynchites. Analysis of the genetic structure of important mosquito populations showed that the haplotype diversity index (Hd) of Ae. japonicus was 0.994 7, with a generally moderate degree of differentiation between populations and a higher degree of genetic differentiation between populations 6 and 12; Aedeslbopictus had a haplotype diversity index of 0.634 0, with significant genetic differentiation between populations 1 and 2 compared to other groups; Armigeres subalbatus had a haplotype diversity index of 0.703 3, with substantial genetic differentiation and limited gene flow between population 2 and populations 3, 4, 6, and 7. Conclusions The mosquito species are rich in Wuyi Mountain Nature Reserve. A comprehensive survey of the composition and distribution of mosquito populations was carried out, and the characteristics of the population genetic structure of the important species of Ae. japonicus, Ae. Albopictus, and Ar. subalbatus populations were analyzed, providing valuable scientific data reference for local mosquito-borne ecological research and disease prevention and control.