Abstract: Objective　This project aimed to study the Miao medicine helleborus thibetanus franchon,including investigating its anti-inflammatory activity in collagen-induced arthritis CIA rats and its mechanism of VEGF/VEGFR2/P38MAPK pathway regulation. Methods　Sixty female Wistar rats were randomly divided into six groups: normal; model; positive drug; and low, medium, high dose groups, with 10 rats in each group. Bovine type Ⅱ collagen solution was injected into the tail of rats to construct the rheumatoid arthritis model, and the positive drug group was given MTX2.0 mg/(kg·d) by gavage once every other day. The three groups of helleborus thibetanus franchon low, medium, and high dose were gavaged with helleborus thibetanus franchon ethanol extract at 0.25, 0.5 and 1 g/(kg·d) once a day. The normal and model groups were given an equivalent volume of NaCl solution, with continuous administration lasting for 28 days. During treatment, the general condition of the rats was observed, body weight changes recorded, and foot thickness measured. After treatment and euthanasia, the rats’ hind limbs were removed for Micro-CT to detect bone destruction;hematoxylin and eosin staining for pathological investigattion of the synovial membrane; immunohistochemistry to observe neovascularization in the synovium;quantitative reverse-transcription PCR to detect mRNA levels of VEGF-A, VEGFR2, TNF-α in the synovial tissue; and Western Blot to detect the expression of VEGF, VEGFR2, p-P38, p-AKT. The analyses were used to explore the potential mechanisms of action of the Miao medicine helleborus thibetanus franchon in treating rheumatoid arthritis. Results　Compared with the normal group, the model group showed significant weight loss (P<0.01), increased foot swelling (P<0.01), visible proliferative synovial tissue with inflammatory cell infiltration, erosive lesions on bone surfaces, increased neovascularization in the synovium, and significant bone destruction in Micro-CT, with reduced bone percentage, trabecular thickness, and bone density. The levels of VEGF-A, VEGFR2, TNF-α mRNA and VEGF-A, VEGFR2, p-P38, p-AKT proteins were significantly elevated (P<0.01). Compared with the model group,the helleborus thibetanus franchon ethanol extract-treated groups showed improvements in these conditions in a dose-dependent manner, with the high-dose group receiving the best effect. There was a significant increase in the rats’ body weight (P<0.05); reduction in foot swelling (P<0.05); amelioration of synovial and erosive bone lesions; reduction in neovascularization in the synovium; and significantly lower levels of VEGF-A, VEGFR2, and TNF-α mRNA, and VEGF-A, VEGFR2, p-P38, and p-AKT protein (P<0.01). Conclusions　The Miao medicine plant helleborus thibetanus franchon may alleviate joint inflammatory damage in CIA rats by modulating the VEGF/VEGFR2 signaling pathway, thereby exerting therapeutic effects on rheumatoid arthritis.

Abstract: Objective　To explore method of establishing and evaluating an asthma rat model with phlegm and blood stasis syndrome. Methods　60 SD male rats were randomly divided into 5 groups, a normal group, asthma group, combination of disease and syndrome (combination) group, DM group, and KCLW group, with 12 rats in each group. Asthma models were established using ovalbumin (OVA). A syndrome model of phlegm and blood stasis was established using a high-fat diet combined with the ice water bath method . We evaluated the asthma model through animal behavior observation, pathological section observation, inflammation index detection, and airway reactivity measurements. The phlegm and blood stasis syndrome model was evaluated via measurements of rat body mass, blood glucose, blood lipids, coagulation function, and hemorheological indexes and by observing symptoms and syndrome determination by Kechuan Liuwei mixture. Results　(1) After OVA induction, the rats in the asthma model group and combination group showed symptoms such as shortness of breath, open mouth breathing, abdominal movement, restlessness, and irritability. HE staining showed the disordered arrangement of the bronchial mucosa in lung tissue, local detachment, thickening of the basement membrane and the bronchial tube wall, narrowing of the lumen, extensive infiltration of inflammatory cells, and congestion of capillaries. Compared with the normal group, the asthma model group and combination group (P<0.05) had increased serum IL-4, IL-6, and TGF-β1. Penh values were increased after stimulation with various concentrations of Mch (P<0.05). (2) Rats in the combination group showed symptoms such as chills, curling up with minimal movement, purple and dark claws, purple and black bruises on the tail, loose stools, and unclean perianal area. Compared with the rats in the asthma model group, rats in the combination group had increased body mass (P<0.05) and blood glucose, triglyceride, and total cholesterol levels (P<0.05), a shortened thrombin time (P<0.05), increased fibrinogen content (P<0.05), and significantly increased whole-blood viscosity at low, medium, and high shear rates (P<0.05). The indexes were significantly improved after Kechuan Liuwei mixture administration. Conclusions　The asthma rat model with phlegm and blood stasis syndrome can be established through OVA induction and high-fat diet combined with ice water bath. The model can be evaluated through behavioral observation, index measurements, and syndrome determination via formulas.

Abstract: Objective　Establishment of a mouse model of orthotopic lung cancer and lymph node metastasis to simulate lymph node metastasis during the occurrence and development of lung cancer and to study the immune cells in the microenvironment before lung cancer metastasis occurs. Methods　Mouse Lewis lung cancer cells were cultured and then injected through the chest walls of mice to establish an orthotopic mouse model of lung cancer. Samples were taken at 7, 14, 21, and 28 days after tumor transplantation, and the changes in the orthotopically transplanted tumors, lymph node tissue structure, and B cells were observed by HE staining and immunohistochemistry. Results　The orthotopic C57BL/6J 　　　　　　mouse model of lung cancer was successfully constructed, and the mediastinal lymph nodes were collected. The changes in the germinal center, lymphatic vessels, and blood vessels within the lymph nodes as well as structural alterations of lymph nodes associated with tumor metastasis and infiltration of tumor cells were observed during the progression of lung cancer using HE staining. Immunohistochemistry showed an increase in the density of B cells in the lymph nodes of the model group. Conclusions　The establishment of this model lays a foundation for the future study of immune cells in the microenvironment of lung cancer before lymph node metastasis occurs.

Abstract: Objective　To investigate the role and related mechanism of the soluble guanylate cyclase (sGC)cGMP-protein kinase G (PKG) signaling pathway in the amelioration of isoproterenol (ISO)-induced cardiac hypertrophy in mice by aqueous extract of Curcuma kwangsiensis root tubers (GYJS). Methods　72 KM male mice were divided randomly into 6 groups: normal, model, propranolol control (40 mg/kg), and GYJS low- (1 g/kg), medium-(2 g/kg), and high-dose (4 g/kg) groups. Mice in the experimental groups were injected subcutaneously with ISO 10 mg/kg on days 1~3 and ISO 5 mg/kg on days 4~14 to establish a mouse cardiac hypertrophy model. 4h after each subcutaneous injection of ISO, the mice in each group were administered the corresponding drugs orally for a dosing cycle of 14 days. The hearts were then removed and whole heart and left ventricle weight were measured. Myocardial tissue pathology was observed using hematoxylin and eosin and Masson staining, and sGC subunit beta-1 (GUCY1B3) and transforming growth factor (TGF β1) were detected by immunohistochemistry. Serum lactate dehydrogenase (LDH), creatine kinase (CK), and Nitric Oxide (NO), and myocardial superoxide dismutase (SOD) activity and malondialdehyde (MDA) content were measured using respective kits. Serum cGMP was detected by enzyme-linked immunosorbent assay and quantitative reverse transcription PCR (RT-qPCR), and atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), GUCY1B3, PKG Ⅰ, and phosphodiesterase (PDE) 5A mRNA expression levels were also determined by RT-qPCR. Results　Compared with the model group, whole heart and left ventricle weights were significantly reduced in mice treated with propranolol or GYJS (P<0.001 or P<0.0001) and myocardial hypertrophy and myocardial fibrosis were significantly reversed. All the treatments significantly elevated myocardial expression of GUCY1B3 (P<0.05 or P<0.01) and significantly reduced expression of TGF-β1 (P<0.05 or P<0.01). The myocardial damage markers LDH and CK were significantly reduced (P<0.05 or P<0.01) while NO and cGMP were significantly elevated (P<0.05 or P<0.01), the myocardial oxidative stress indicator MDA was significantly reduced (P<0.05 or P<0.01) and SOD activity was significantly increased (P<0.05 or P<0.01). mRNA levels of the myocardial hypertrophy markers ANP, BNP, and PDE5A were significantly reduced (P<0.05, P<0.01, or P<0.001) and the mRNA levels of GUCY1B3 and PKGⅠ were significantly increased (P<0.01 or P<0.001). Conclusions　GYJS may improve cardiac hypertrophy by modulating the sGC-cGMP-PKG signaling pathway.

Abstract: Objective　To investigate the inhibitory effect of Sauchinone (Sch) on renal tissue fibrosis and to explore its mechanism in unilateral ureteral obstruction (UUO) mice. Methods　Forty male ICR mice were divided into Sham, UUO-Model, Sch Low-dose, Sch High-dose, and positive control (Val) groups. Mice in the sham group underwent isolation of the left renal ureter without ligation, and mice in the UUO-Model group underwent stripping of the left proximal ureter for double ligation. Starting on day 2 after surgery, the Sch Low-dose group received 10 mg/kg Sch, the Sch High dose group received 30 mg/kg Sch, and the Val group received 100 mg/kg valsartan by gavage once daily for 4 weeks. At the end of drug administration, serum creatinine (SCr), blood urea nitrogen (BUN), and the inflammatory factors tumor necrosis factor (TNF)-α and interleukin (IL)-6 were measured. Superoxide dismutase (SOD) activity, and malondialdehyde (MDA) and reactive oxygen species (ROS) content were detected in renal tissues. TNF-α and IL-6 mRNA levels were detected in renal tissues by reverse transcription-quantitative polymerase chain reaction. Pathological changes and collagen deposition, as well as transforming growth factor (TGF)-β1, Smad3, and connective tissue growth factor (CTGF) protein expression in mouse renal tissues were detected by hematoxylin and eosin and Masson staining, and immunohistochemistry. Collagen Ⅰ, CTGF, Smad3, nuclear factor erythroid 2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), and nuclear factor (NF)-κB protein expression in renal tissues were detected by Western Blot. Results　SCr, BUN, TNF-α, and IL-6 were all significantly reduced in Sch Low-dose and Sch High-dose mice compared with the UUO Model group (P<0.05 or P<0.01), while SOD activity was significantly higher and MDA and ROS levels were significantly lower in renal tissues (P<0.05 or P<0.01), and TNF-α and IL-6 mRNA in renal tissues were significantly reduced (P<0.05 or P<0.01). Microscopic observation of swollen renal tubules with thylakoid hyperplasia and collagen fiber deposition was significantly improved. Immunohistochemistry showed a significant reduction in TGF-β1, Smad3, and CTGF protein expression in renal tissues, and Western Blot showed significantly reduced levels of Collagen Ⅰ, CTGF, Smad3, and NF-κB protein expression (P<0.05 or P<0.01), while Nrf2 and HO-1 protein expression levels were significantly elevated (P<0.05 or P<0.01). Conclusions　The amelioration of renal tissue fibrosis by sauchinone may be mediated via anti-inflammatory and antioxidative stress effects that modulate the expression of pro-fibrotic proteins in the TGF-β1/Smad3 signaling pathway.

Abstract: Objective　To explore the effect and possible mechanism of aerobic exercise and empagliflozin (EMPA) on isoproterenol (ISO)-induced pathological cardiac remodeling. Methods　Mice were divided randomly into control (Con), ISO, exercise (EX) + ISO, EMPA + ISO, and EX + EMPA + ISO groups. Mice in the EX groups were trained continuously for 6 weeks, mice in the EMPA groups were gavaged continuously for 4 weeks, and mice in the ISO groups were injected subcutaneously with ISO for 7 days before dissection. After euthanasia, the whole heart mass index, left heart mass index, heart mass to tibial length ratio, and left heart mass to tibial length ratio were calculated by weighing and measuring. Pathological changes, collagen fiber deposition, and myocardial cell cross-sectional area in the hearts were detected by hematoxylin and eosin, Sirius red, and wheat germ agglutinin staining. The expression levels of genes and proteins related to cardiac fibrosis and hypertrophy, macrophage infiltration, ferroptosis, and the phosphoinositide 3-kinase (PI3K)/AKT pathway were examined by quantitative reverse transcription-polymerase chain reaction, Western Blot, and immunofluorescence staining. Results　(1) The whole heart mass index, left heart mass index, heart mass to tibial length ratio, and left heart mass to tibial length ratio showed downward trends in the EX + ISO group compared with the ISO group. The whole heart mass index and left heart mass index were significantly decreased in the EMPA + ISO group (P<0.01, P<0.05), and the heart mass to tibial length ratio and left heart mass to tibial length ratio were both down regulated. Mice in the EX + EMPA + ISO group had a significant decrease in whole heart mass index (P<0.05), and the other three indicators were all down-regulated. (2) Myocardial cells were more orderly in the three intervention groups compared with the ISO group, with significant reductions in inflammatory cell infiltration (P<0.01), the area of cardiac fibrosis, and the cross-sectional area of myocardial cells (P<0.001). (3) The mRNA and protein expression levels of Col 1 and Anp were significantly reduced in the three intervention groups compared with the ISO group (P<0.05, P<0.01, P<0.001). Col 3 mRNA expression significantly reduced in the EMPA + ISO and EX + EMPA + ISO groups (P<0.05), and showed a downward trend in the EX + ISO group. (4) Macrophage infiltration and IL-6 mRNA levels were significantly reduced in the three intervention groups compared with the ISO group (P<0.05, P<0.01, P<0.001).(5) Nrf2 and Gpx4 mRNA levels were upregulated in the three intervention groups compared with the ISO group, with a significant increase in GPX4 protein expression (P<0.01, P<0.001) and a significant decrease in HO-1 protein expression (P<0.01, P<0.001). (6) Pi3k mRNA levels were significantly increased in the EX + ISO group compared with the ISO group (P<0.05), and Pi3k mRNA was upregulated in the EMPA + ISO and EX + EMPA + ISO groups. Akt mRNA levels showed an upward trend in the three intervention groups. PI3K and phospho-AKT protein levels were significantly increased in the EX + ISO group (P<0.01, P<0.05), and showed an increasing trend in the EMPA + ISO and EX + EMPA + ISO groups. Conclusions　Moderate intensity aerobic exercise, the novel hypoglycemic drug EMPA, and their combination can alleviate ISO-induced pathological cardiac remodeling, possibly via a mechanism related to activation of the PI3K/AKT signaling pathway and inhibition of cardiac ferroptosis.

Abstract: Objective　To investigate the effect of the monoamine oxidase A (MAOA), Maoa c.1409 T > C synonymous mutation on anxiety, fear, and other emotional behaviors in mice. Methods　In this study, CRISPR/Cas9 technology was used to construct a mouse model of a single nucleotide polymorphism (SNP) synonymous mutation. We evaluated the differential effect of this gene between males and females through animal behavior and gene expression studies in animal models. In terms of animal behavior, an open field test, elevated plus maze test, defensive burial experiment, forced swimming test, and 3D behavioral analysis were used. Other method were used to evaluate behavioral differences between male and female mice with polymorphisms in Maoa synonymous mutant genes. Results　The result of the open field experiment showed that the residence time of female SNP mice in the central area was significantly higher than that of male SNP mice (P<0.001). In the elevated cross maze experiment, the EPM result showed that the time and frequency of male SNP mice entering the open arm were higher than those of female SNP mice, but there was no significant difference. The defensive burial test showed that the number and duration of excavations by female SNP mice in response to rat urine were significantly reduced (P<0.01). The FST showed that SNP females had shorter immobility time and longer swimming time (P<0.05), and thus their depression was lower than males. 3D-AI fine behavior analysis showed no significant male and female differences, except for the movement trajectory and climbing behavior of mice. The MAOA enzyme content of female SNP mice was significantly lower than that of male SNP mice (P<0.001), but there was no significant difference in enzyme activity between male and female SNP mice. Conclusions　The synonymous mutation of Maoa c.1409 T > C acts by affecting the expression of MAOA and may have different fear, anxiety, and mood effects in male and female SNP mice.

Abstract: Objective　To employ a mouse model of ABI3BP gene deletion for the detection of postnatal changes in body weight and glucose metabolism and establish a different method of creating a mouse model of low birth weight. Methods　Heterozygote mice were mated to produce ABI3BP gene knockout homozygote (ABI3BP^-/-) mice, heterozygote (ABI3BP ^{+ /-}) mice, and wild-type (WT) mice. Adult mice from all three groups were evaluated for glucose metabolism markers, including the fasting blood glucose level, glucose tolerance, and insulin tolerance. Additionally, body weight was measured at various postnatal time periods, and the weight ratio of critical organs in adulthood was calculated. Results The gene sequencing result of the polymerase chain reaction product of ABI3BP^-/- mice showed that frameshift mutations occurred in the knockout region, with quantitative reverse-transcription polymerase chain reaction analysis demonstrating significantly reduced ABI3BP expression in ABI3BP^-/- mice compared with that in WT mice. Notably, the birth weight of ABI3BP^-/- mice (1.25 ± 0.08 g) was markedly lower than that of WT mice (1.34 ± 0.12 g) (P<0.05). Conversely, the weight of adult (120 d) ABI3BP^-/- mice (27.70 ± 1.93 g) was significantly higher than that of WT mice (23.64 ± 1.34 g) (P<0.01). The ratios of key organ weights to body weight were not significantly different between the groups (P>0.05). Fasting blood glucose and insulin tolerance tests showed no significant variations between the groups. However, glucose tolerance tests indicated that ABI3BP^-/- mice had lower blood glucose levels (15.68 ± 7.04 mmol/L) than WT mice (23.01 ± 5.75 mmol/L). Conclusions　Deletion of the ABI3BP gene result in mice with low birth weight, poor growth recuperation, and inadequate glucose tolerance in adulthood, similar to the clinical growth traits of low-birth-weight human neonates. Therefore, this mouse model is a promising choice for the study of low birth weight.

Abstract:Type 2 diabetes mellitus (T2DM) is a common metabolic disease characterized by hyperglycemia and insulin resistance. Peri-implantitis is a common complication of oral implants and is a frequent cause of implant restoration failure. In persistent hyperglycemia, T2DM patients are prone to peri-implant inflammation and aggravated bone destruction. An increasing proportion of T2DM patients with tooth loss choose oral implantation, and the probability of periimplant inflammation in such patients is increased. However, the pathogenesis and correlation between the conditions have not been the focus of in-depth studies. Rodent models can simulate T2DM and peri-implant inflammation and are widely used animal models of both diseases. Animal models of T2DM complicated with peri-implant inflammation are helpful for simulating the complex internal environment and further studying the pathologic progression, pathogenesis, interactions, and treatment. However, few such models have been constructed as yet. In this paper, we review rodent models of T2DM, periimplantitis, and T2DM combined with periimplantitis constructed in recent years, and their advantages and disadvantages, to provide a reference and help for relevant researchers.

Abstract:TGF-β1 is considered a key mediator in the formation of hepatic fibrosis and mainly acts by activating the downstream Smad signaling pathway. Smad2 and Smad3 are two major downstream regulators that promote TGF-β1mediated tissue fibrosis, while Smad7 is a negative-feedback regulator of the TGF-β1/Smad pathway and inhibits TGF-β1mediated hepatic fibrosis. A growing number of studies are showing that natural products can delay the progression of hepatic fibrosis by regulating the TGF-β1/Smad pathway, inhibiting HSC activation, and reducing ECM deposition. This article reviews the molecular mechanism of the TGF-β1/Smad signaling pathway in hepatic fibrosis, and summarizes the natural products that target the regulation of this pathway, providing a reference for research into the treatment of hepatic fibrosis.

Abstract:Monoamine oxidase A (MAOA) is a membrane-bound mitochondrial enzyme that exists in almost all vertebrate tissues, where it catalyzes the degradation of biogenic and dietary-derived monoamines. MAOA has the function of regulating neurotransmitter metabolism and is associated with anti-tumor immune responses. Most previous studies have focused on the role of MAOA in tumor cells, while more recent findings suggest that MAOA plays an equally significant role in tumor-associated immune cells. In this review, we summarize the regulatory effect of MAOA on the inhibitory tumor microenvironment. The suppressing function of MAOA on various types of tumor-associated immune cells (e.g., CD8⁺ T cells and tumor-associated macrophages) by its direct effect on monoamines and their metabolic characteristics are discussed. We propose that developing novel MAOA-inhibitor drugs and exploring multidrug-combination strategies may enhance the efficacy of immune therapy for tumors. In conclusion, MAOA may act as a novel target in tumor immunity and influence the effect of tumor immunotherapy.

Abstract:Delirium is an acute brain dysfunction syndrome characterized by confusion and difficulty concentrating, which mainly affects intensive care unit patients and elderly inpatients. Treatment is expensive and may also lead to increased risks of serious complications and death. The complex etiology and unknown pathological mechanisms of delirium mean that clinical drug treatment is largely ineffective. Animal models therefore provide a powerful tool to help understand the mechanism of delirium, screen new drugs, and study potential intervention measures. We review experimental research related to delirium animal models worldwide, and summarize the latest progress in the construction and evaluation of these models from the aspects of animal selection, model construction method , and model evaluation, to provide a reference for further experimental research based on delirium animal models.

Abstract:As a complex and refractory disease, the incidence of steroid-induced osteonecrosis of the femoral head (SONFH) is increasing year by year and showing an increasing trend in younger people. A good animal model of disease can provide important support for studying the pathogenesis of SONFH and the development of treatment method . In this paper, the latest progress made in the construction of SONFH animal models is summarized and analyzed from the aspects of animal selection, modeling method , and model evaluation. To achieve this, we review and categorize the experimental studies related to SONFH animal models conducted in China and overseas in recent years, providing a reference for related research of SONFH.

Abstract:Gene-knockout technology is increasingly used as a powerful tool for establishing animal models of hyperuricemia (HUA). HUA gene-knockout animal models are not only helpful in revealing the molecular mechanisms of uric acid metabolism but are also of great value for evaluating potential therapeutic strategies. In this paper, the application of gene-knockout technology in HUA animal models is discussed in detail by reviewing the domestic and foreign literature, focusing on the knockout of urate oxidase, glucose transporter 9, and ATP-binding cassette transporter G2. The review provides a reference and guidance for the further establishment of HUA animal models by gene knockout technology.