OBJECTIVE: To investigate the effects of removing microglia from spinal cord on nerve repair and functional recovery after spinal cord injury (SCI) in mice. METHODS: Thirty-nine 6-week-old female C57BL/6 mice were randomly divided into control group ( n=12), SCI group ( n=12), and PLX3397+SCI group ( n=15). The PLX3397+SCI group received continuous feeding of PLX3397, a colony-stimulating factor 1 receptor inhibitor, while the other two groups were fed a standard diet. After 14 days, both the SCI group and the PLX3397+SCI group were tested for ionized calcium binding adapter molecule 1 (Iba1) to confirm that the PLX3397+SCI group had completely depleted the spinal cord microglia. The SCI model was then prepared by clamping the spinal cord in both the SCI group and the PLX3397+SCI group, while the control group underwent laminectomy. Preoperatively and at 1, 3, 7, 14, 21, and 28 days postoperatively, the Basso Mouse Scale (BMS) was used to assess the hind limb function of mice in each group. At 28 days, a footprint test was conducted to observe the gait of the mice. After SCI, spinal cord tissue from the injury site was taken, and Iba1 immunofluorescence staining was performed at 7 days to observe the aggregation and proliferation of microglia in the spinal cord. HE staining was used to observe the formation of glial scars at the injury site at 28 days; glial fibrillary acidic protein (GFAP) immunofluorescence staining was applied to astrocytes to assess the extent of the injured area; neuronal nuclei antigen (NeuN) immunofluorescence staining was used to evaluate neuronal survival. And 5-hydroxytryptamine (5-HT) immunofluorescence staining was performed to assess axonal survival at 60 days. RESULTS: All mice survived until the end of the experiment. Immunofluorescence staining revealed that the microglia in the spinal cord of the PLX3397+SCI group decreased by more than 95% compared to the control group after 14 days of continuous feeding with PLX3397 ( P<0.05). Compared to the control group, the BMS scores in the PLX3397+SCI group and the SCI group significantly decreased at different time points after SCI ( P<0.05). Moreover, the PLX3397+SCI group showed a further decrease in BMS scores compared to the SCI group, and exhibited a dragging gait. The differences between the two groups were significant at 14, 21, and 28 days ( P<0.05). HE staining at 28 days revealed that the SCI group had formed a well-defined and dense gliotic scar, while the PLX3397+SCI group also developed a gliotic scar, but with a more blurred and loose boundary. Immunofluorescence staining revealed that the number of microglia near the injury center at 7 days increased in the SCI group than in the control group, but the difference between groups was not significant ( P>0.05). In contrast, the PLX3397+SCI group showed a significant reduction in microglia compared to both the control and SCI groups ( P<0.05). At 28 days after SCI, the area of spinal cord injury in the PLX3397+SCI group was significantly larger than that in SCI group ( P<0.05); the surviving neurons significantly reduced compared with the control group and SCI group ( P<0.05). The axonal necrosis and retraction at 60 days after SCI were more obvious. CONCLUSION The removal of microglia in the spinal cord aggravate the tissue damage after SCI and affecte the recovery of motor function in mice, suggesting that microglia played a neuroprotective role in SCI.
Animals ; Spinal Cord Injuries/surgery* ; Microglia/pathology* ; Female ; Mice ; Mice, Inbred C57BL ; Nerve Regeneration/drug effects* ; Spinal Cord/pathology* ; Pyrroles/administration & dosage* ; Aminopyridines/administration & dosage* ; Recovery of Function ; Disease Models, Animal ; Calcium-Binding Proteins/metabolism* ; Receptors, Granulocyte-Macrophage Colony-Stimulating Factor/antagonists & inhibitors* ; Microfilament Proteins/metabolism* ; Glial Fibrillary Acidic Protein/metabolism*

OBJECTIVE: To summarize the research progress of rodent models of secondary lymphedema (SL) and provide a reference for selecting appropriate animal models in SL research. METHODS: Recent literature on rodent SL models at home and abroad was comprehensively analyzed, summarizing model categories, development techniques, strengths, and weaknesses. RESULTS: Current research primarily utilizes rats and mice to establish SL models. The main model types include hind limb, forelimb, tail, and head/neck models. The hind limb model is the most frequently employed, typically requiring surgery combined with irradiation to induce stable chronic edema. Forelimb models primarily simulate upper limb lymphedema, but exhibit relatively rapid edema resolution. Tail models offer operational simplicity and are predominantly used for studying acute edema mechanisms and interventions; however, they demonstrate poor clinical relevance. Emerging head/neck models provide a valuable tool for investigating head and neck cancer-associated lymphedema. These models exhibit variations in lymphedema duration, degree of fibrosis, and edema incidences. CONCLUSION Existing models still fall short in faithfully replicating the chronicity, fibrosis, fat deposition, and complex microenvironment characteristic of human chronic lymphedema. Future research must integrate multidisciplinary approaches, optimize model construction strategies, and explore novel modeling approaches to more accurately mimic the human disease and advance SL prevention and treatment research.
Lymphedema/pathology* ; Animals ; Disease Models, Animal ; Rats ; Mice ; Humans ; Head and Neck Neoplasms/complications* ; Rodentia

OBJECTIVE: To investigate the effects of 4-methylcatechol (4MC) on the migration and inflammatory response in rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS), as well as its underlying mechanisms of action. METHODS: RA-FLS was isolated from synovial tissue donated by RA patients, and the optimal concentration of 4MC was determined by cell counting kit 8 method for subsequent experiments, and the effect of 4MC on the migratory ability of RA-FLS was evaluated via a cell scratch assay. An inflammation model of RA-FLS was induced by tumor necrosis factor α (TNF-α). Real-time fluorescence quantitative PCR and ELISA were employed to detect the gene and protein expression levels of interleukin-1β (IL-1β) and IL-6 in RA-FLS and their culture supernatants, respectively, thereby investigating the anti-inflammatory effects of 4MC. Western blot was used to examine the expressions of nuclear factor κB (NF-κB) signaling pathway-related proteins, including inhibitor of NF-κB-α (IKBα), phosphorylated (P)-IκBα, NF-κB-inducing kinase α (IKKα), P-IKKαβ, P-p65, and p65. Cellular immunofluorescence was utilized to detect the expression and localization of p65 in RA-FLS, exploring whether 4MC exerts its anti-inflammatory effects by regulating the NF-κB signaling pathway. Finally, a collagen-induced arthritis (CIA) mouse model was established. The anti-RA effect of 4MC in vivo was evaluated by gross observation and histological examination. RESULTS: 4MC inhibited RA-FLS migration in a concentration-dependent manner. In the TNF-α-induced RA-FLS inflammation model, 4MC significantly decreased the gene and protein expression levels of IL-1β and IL-6. Furthermore, 4MC markedly reduced the ratios of P-IΚBα/IΚBα, P-IKKαβ/IKKα, and P-p65/p65, thereby blocking the transcriptional activity of p65 by inhibiting its nuclear translocation. This mechanism effectively suppressed the activation of the TNF-α-mediated NF-κB signaling pathway. Animal studies demonstrated that 4MC [10 mg/(kg·day)] significantly lowered serum levels of IL-1β, IL-6, and TNF-α, and alleviated arthritis severity and bone destruction in CIA mice. CONCLUSION 4MC not only inhibits the migration of RA-FLS but also mitigates their inflammatory response by suppressing the NF-κB signaling pathway, thereby effectively exerting its anti-RA effects.
Synoviocytes/metabolism* ; Arthritis, Rheumatoid/metabolism* ; Animals ; Cell Movement/drug effects* ; Humans ; Catechols/therapeutic use* ; Fibroblasts/drug effects* ; Mice ; Tumor Necrosis Factor-alpha/pharmacology* ; Interleukin-1beta/metabolism* ; Interleukin-6/metabolism* ; Signal Transduction/drug effects* ; NF-kappa B/metabolism* ; Transcription Factor RelA/metabolism* ; Synovial Membrane/cytology* ; Cells, Cultured ; Male ; Arthritis, Experimental ; Anti-Inflammatory Agents/pharmacology* ; NF-KappaB Inhibitor alpha ; Inflammation

OBJECTIVE: To explore the construction of a canine model of vascularized allogeneic spinal cord transplantation (vASCT) and preliminarily evaluate its therapeutic efficacy for spinal cord injury (SCI). METHODS: Sixteen female Beagle dogs aged 8-12 months were randomly selected, with 8 dogs serving as donors for the harvesting of spinal cord tissue with a vascular pedicle [dorsal intercostal artery (DIA) at the T₁₀ level and accompanying vein]. The remaining 8 dogs underwent a 1.5-cm-length spinal cord defect at the T₁₀ level, followed by transplantation of the donor spinal cord tissue for repair. Polyethylene glycol (PEG) was applied to both ends to spinal cord graft; then, using a random number table method, the dogs were divided into an experimental group (n=4) and a control group (n=4). The experimental group received immunosuppressive intervention with oral tacrolimus [0.1 mg/(kg∙d)] postoperatively, while the control group received no treatment. The operation time and ischemia-reperfusion time of two groups were recorded. The recovery of hind limb function was estimated by Olby score within 2 months after operation; the motor evoked potentials (MEP) was measured through neuroelectrophysiological examination, and the spinal cord integrity was observed through MRI. RESULTS: There was no significant difference in the operation time and ischemia-reperfusion time between the two groups (P>0.05). All dogs survived until the completion of the experiment. Within 2 months after operation, all dogs in the control group failed to regain the movement function of hind limbs, and Olby scores were all 0. In the experimental group, the movement and weight-bearing, as well as walking abilities of the hind limbs gradually recovered, and the Olby scores also showed a gradually increasing trend. There was a significant difference between the two groups from 3 to 8 weeks after operation (P<0.05). Neuroelectrophysiological examination indicated that the electrical signals of the experimental group passed through the transplanted area, and the latency was shortened compared to that at 1 month after operation (P<0.05), showing continuous improvement, but the amplitude did not show significant improvement (P>0.05). The control group was unable to detect any MEP changes after operation. MRI examination showed that the transplanted spinal cord in the experimental group survived and had good continuity with normal spinal cord tissue, while no relevant change was observed in the control group. CONCLUSION The vASCT model of dogs was successfully constructed. This surgical procedure can restore the continuity of the spinal cord. The combination of tacrolimus anti-immunity is a key factor for the success of transplantation.
Animals ; Dogs ; Female ; Spinal Cord/blood supply* ; Spinal Cord Injuries/surgery* ; Transplantation, Homologous ; Disease Models, Animal ; Recovery of Function ; Plastic Surgery Procedures/methods* ; Tacrolimus ; Immunosuppressive Agents

OBJECTIVE: To investigate the effect of mild hypothermia on neurological function in rats with spinal cord injury (SCI) based on the adenosine monophosphate activated protein kinase (AMPK)/Nod-like receptor protein 3 (NLRP3) pathway. METHODS: Fifty 7-8 weeks old SPF male Sprague Dawley rats were used to establish rat model of SCI by Allen's method. Among them, 48 successfully modeled rats were randomly divided into SCI group, mild hypothermia group (SCI+mild hypothermia treatment), and Compound C group (SCI+mild hypothermia+intraperitoneal injection of 20 mg/kg AMPK/NLRP3 pathway inhibitor Compound C), with 16 rats in each group. Another 16 normal rats with laminectomy were selected as sham-operation group. Basso-Beattie-Bresnahan (BBB) score was used to evaluate the motor ability of rats at 1, 3, 7, 14 days after treatment. After 14 days, the rats were sacrificed, and the spinal cord histopathological morphology was observed by HE staining, the neuronal apoptosis in spinal cord tissue was detected by TUNEL assay, and the serum levels of interleukin 2 (IL-2), IL-6, transforming growth factor β ₁ (TGF-β ₁), malondialdehyde (MDA), and superoxide dismutase (SOD) were detected by ELISA. The expressions of AMPK/NLRP3 pathway proteins in spinal cord tissue, including B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax), and cleaved-Caspase-9 were detected by Western blot. RESULTS: At 1 day after treatment, the rats in SCI group, mild hypothermia group, and Compound C group did not recover their motor ability. With the prolongation of time, the motor function of rats in each group gradually recovered. Among them, the BBB score of SCI group was significantly lower than that of sham-operation group and mild hypothermia group ( P<0.05), and the BBB score of Compound C group was significantly lower than that of mild hypothermia group ( P<0.05). Compared with the sham-operation group, the SCI group displayed obvious pathological changes in the spinal cord tissue, with disordered tissue architecture, inflammatory infiltration, and blurred interstitial boundaries. The neuronal apoptosis rate, Bax/Bcl-2 ratio, cleaved Caspase-9 expression, NLRP3 protein expression, serum IL-2, IL-6, and MDA levels were elevated, whereas serum TGF-β ₁, SOD levels, and spinal cord phosphorylation AMPK/AMPK protein expression significantly decreased ( P<0.05). Compared with the SCI group, the above phenomena significantly improved in the mild hypothermia group ( P<0.05), while the Compound C group showed the opposite trend of change compared to the mild hypothermia group ( P<0.05). CONCLUSION Mild hypothermia can attenuate neurological dysfunction after SCI in rats, potentially by activating the AMPK/NLRP3 pathway.
Animals ; Spinal Cord Injuries/physiopathology* ; Rats, Sprague-Dawley ; Male ; Rats ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; AMP-Activated Protein Kinases/metabolism* ; Hypothermia, Induced ; Signal Transduction ; Spinal Cord/pathology* ; Apoptosis ; Interleukin-6/metabolism* ; Disease Models, Animal ; bcl-2-Associated X Protein/metabolism* ; Superoxide Dismutase/metabolism* ; Proto-Oncogene Proteins c-bcl-2/metabolism* ; Caspase 9/metabolism*

Systemic lupus erythematosus (SLE) is a complex autoimmune disease with diverse clinical symptoms, which involves multiple systems and organs such as the kidney, cardiovascular system, and nervous system. The main characteristic of SLE is the mistaken attacks on self-tissues by the immune system, but the specific pathogenesis is still unclear. This paper reviews and compares the modeling mechanisms and main pathological manifestations of spontaneous and experimental mouse models of SLE, which can provide evidences for selecting appropriate mouse models in studies on the pathogenesis, clinical diagnosis and treatment of SLE.
Animals ; Lupus Erythematosus, Systemic/immunology* ; Disease Models, Animal ; Mice ; Humans

Objective To explore the effect of Evodiamine (Evo) on the immune function of allergic rhinitis (AR) rats and the regulatory mechanism on C-C motif chemokine ligand 2 (CCL2)/ C-C motif chemokine receptor 2 (CCR2) pathway. Methods The related targets of Evo-AR-immune function were screened by network pharmacology, and the protein interaction network diagram of intersecting targets was constructed. The AR rat model was established by ovalbumin (OVA) combined with aluminium hydroxide, and the rats were divided into six groups: a normal control (NC) group, a model group, a Loratadine (LOR) group, an Evodiamine low dose (Evo-L) group, a Evodiamine high dose (Evo-H) groups, and an Evo-H combined with CCL2 group. After the last administration, the symptoms of rats in each group were scored; ELISA was applied to detect the levels of histamine, immunoglobulin E (IgE), interleukin 4 (IL-4), IL-13 and interferon γ (IFN-γ); Diff-Quick staining solution was applied to detecte the number of cells in the nasal lavage fluid (NALF); hematoxylin eosin (HE) staining was applied to observe the pathological changes of nasal mucosa tissue; real-time quantitative PCR was applied to detect the levels of CCL2 and CCR2 mRNA in tissue; Western blot was applied to detect the expression levels of CCL2, CCR2 and CXC motif chemokine ligand 8 (CXCL8) proteins in nasal mucosa. Results There were eight intersection targets of EVo-AR-immune function, and protein interaction network diagram showed that CXCL8 was the core target. Compared with the NC group, the score of nasal symptoms, the levels of histamine, IgE, IL-4 and IL-13, the numbers of eosinophil, macrophages, neutrophils, lymphocytes and total cells, the mRNA and protein expression levels of CCL2 and CCR2, and the expression of CXCL8 protein in the model group were increased, while the level of IFN-γ was decreased. Compared with the model group, the score of nasal symptoms, the levels of histamine, IgE, IL-4 and IL-13, the numbers of eosinophil, macrophages, neutrophils, lymphocytes and total cells, the mRNA and protein expression levels of CCL2 and CCR2, and the expression of CXCL8 protein in LOR and Evo groups were decreased, while the level of IFN-γ was increased. Further use of CCL2 recombinant protein for compensatory experiments revealed that the improvement effect of Evo on immune function in AR rats was reversed by CCL2. Conclusion Evo can improve the immune function of AR rats, and its mechanism may be related to the inhibition of the CCL2/CCR2 pathway.
Animals ; Receptors, CCR2/immunology* ; Signal Transduction/drug effects* ; Chemokine CCL2/immunology* ; Rats ; Rhinitis, Allergic/metabolism* ; Immunoglobulin E/blood* ; Quinazolines/pharmacology* ; Male ; Interferon-gamma ; Rats, Sprague-Dawley ; Interleukin-13 ; Histamine ; Interleukin-4/immunology* ; Disease Models, Animal

Objective To explore the effects of S100 calcium-binding protein A9 (S100A9) gene knockout on the phenotype of systemic lupus erythematosus (SLE) in mice and to clarify the role of S100A9 in the pathogenesis of SLE. Methods Ten female C57BL/6 wild-type and S100A9 knockout (S100A9-KO ) mice were selected, with five wild-type and five S100A9-KO B6 mice receiving imiquimod (IMQ) cream to establish SLE mouse model. The other five wild-type and five S100A9-KO B6 mice were treated as control groups by wiping the skin of the right ear with a cotton swab. After 8 weeks, the mice were sacrificed. The serum was collected from each mouse to detect the levels of anti-double-stranded DNA (dsDNA) antibodies, immunoglobulin G (IgG), B cell activating factor (BAFF), and interleukin 6 (IL-6) using ELISA. The levels of serum creatinine were determined using a sarcosine oxidase method. Urine was collected to measure urinary protein concentration. Kidneys were collected and stained with hematoxylin and eosin (H&E) for evaluating histological changes. Results After IMQ treatment, the length and weight of spleen, levels of serum creatinine, anti-dsDNA antibodies, IgG, BAFF, IL-6, and urinary protein in the IMQ B6 group and IMQ S100A9-KO B6 group were significantly higher than those of the control groups. Lupus-like changes including increased glomerular volume and tubular epithelial swelling were observed in kidneys from the IMQ and IMQ S100A9-KO groups. However, compared with the IMQ B6 group, the IMQ S100A9-KO B6 group exhibited milder levels of serum and urine indicators as well as the lupus-like symptoms. Conclusion IMQ could induce lupus-like symptoms in both wild-type B6 mice and S100A9-KO B6 mice, but the lesions in S100A9 knockout mice are milder. Theses results suggested that S100A9 is involved in and promotes the pathogenesis of SLE.
Animals ; Lupus Erythematosus, Systemic/chemically induced* ; Female ; Calgranulin B/genetics* ; Mice, Knockout ; Mice, Inbred C57BL ; Phenotype ; Mice ; Interleukin-6/blood* ; Disease Models, Animal ; Antibodies, Antinuclear/blood* ; B-Cell Activating Factor/blood* ; Immunoglobulin G/blood* ; Kidney/pathology*

Objective To investigate the effects of cucurbitacin B (CucB) on alleviating skin lesions and inflammation in psoriasis mice via the cGAS-STING signaling pathway. Methods The expression of genes associated with the cGAS-STING signaling pathway in psoriatic lesions and non-lesional skin was analyzed, and hallmark gene set enrichment analysis was performed. The cytotoxicity of CucB on BMDMs was evaluated using the CCK-8 assay. The expression levels of genes and proteins related to the cGAS-STING signaling pathway, along with the secretion of inflammatory cytokines, were measured at different concentrations of CucB using quantitative PCR, Western blotting, and ELISA. Imiquimod-induced psoriasis BALB/c mice were divided into four groups: normal group, model group, low-dose CucB group [0.1 mg/ (kg.d)], and high-dose CucB group [0.4 mg/ (kg.d)], with five mice per group. PASI scoring was performed to assess the severity of psoriasis after 6 days of treatment, and HE staining was conducted to observe pathological damage. Meanwhile, the mRNA levels of inflammatory cytokines and their secretion were detected by qPCR and ELISA. Results Most cGAS-STING signaling-related genes were upregulated in lesional skin of psoriasis patients, and the hallmark gene set enrichment analysis revealed that the most significantly upregulated genes were primarily associated with immune response signaling pathways. CucB inhibited dsDNA-induced phosphorylation of interferon regulatory factor 3 (IRF3) and STING proteins in both bone-marrow derived macrophages(BMDMs) and THP-1 cells. CucB also suppressed dsDNA-induced mRNA expression of IFNB1, TNF, IFIT1, CXCL10, ISG15, and reduced the secretion of cytokines such as IFN-β, IL-1β, and TNF-α in THP-1 cells. In the imiquimod-induced psoriasis mouse model, CucB treatment reduced psoriatic symptoms, alleviated skin lesions, and attenuated inflammation. ELISA and qPCR results showed that CucB significantly reduced serum secretion levels of IL-6, TNF-α, and IL-1β, as well as the mRNA levels of IL23A, IL1B, IL6, TNF, and IFNB1. Conclusion CucB inhibits cytoplasmic DNA-induced activationc of the GAS-STING pathway. CucB significantly attenuates skin lesions and inflammation in IMQ-induced psoriatic mice, and the potential molecular mechanism may be related to the down-regulation of the cGAS-STING pathway.
Animals ; Psoriasis/pathology* ; Signal Transduction/drug effects* ; Membrane Proteins/genetics* ; Mice ; Nucleotidyltransferases/genetics* ; Disease Models, Animal ; Mice, Inbred BALB C ; Skin/metabolism* ; Triterpenes/therapeutic use* ; Humans ; Cytokines/metabolism* ; Inflammation/drug therapy* ; Male

Objective To generate and characterize natural killer cell (NK cell)-conditional Cd226 gene knockout mice using Cre-loxP technology, and to explore the role of CD226 on NK cells in alleviating intestinal inflammation in a murine model of ulcerative colitis (UC). Methods NK cell-conditional Cd226 gene knockout mice were generated by crossing loxP-flanked Cd226 mice with Ncr1-Cre mice via the Cre-loxP system. Polymerase chain reaction (PCR) and agarose gel electrophoresis were used for genotyping. A UC model was established by dextran sulfate sodium (DSS) induction. Flow cytometry was performed to analyze CD226 expression levels on NK cells and the infiltration of related immune cells in colon tissues. Hematoxylin-eosin (HE) staining was performed to assess the degree of colonic inflammation. Results DNA gel electrophoresis and flow cytometry confirmed the successful generation of NK cell-specific Cd226 knockout mice. After conditional knockout of Cd226 in NK cells, inflammation in the UC mouse model was alleviated. Flow cytometry results showed a reduced proportion of NK cells in peripheral blood and the colon lamina propria, while HE staining demonstrated attenuated inflammatory responses. Conclusion Specific knockout of Cd226 in NK cells mitigates intestinal inflammation in UC mice by reducing NK cell numbers and inhibiting their pro-inflammatory functions.
Animals ; Colitis, Ulcerative/pathology* ; Killer Cells, Natural/metabolism* ; Mice, Knockout ; T Lineage-Specific Activation Antigen 1 ; Antigens, Differentiation, T-Lymphocyte/genetics* ; Mice ; Disease Models, Animal ; Mice, Inbred C57BL ; Male