Abstract:Objective A rat model of damp heat and cold damp diarrhea was established and treated with Baitouweng and Lizhong decoctions to analyze differences in clinical symptoms, cell density, and oxidative stress indexes of diarrhea rats with different syndrome types of damp heat and cold damp diarrhea, and provide an Objective basis to explore the treatment characteristics of diarrhea with the same disease and different treatment. Methods SD rats were divided into blank (CON), dampness heat diarrhea model (DHD), dampness heat + Baitouweng decoction (DHB), dampness heat + Lizhong decoction (DHL), cold dampness diarrhea model group (CDD), cold dampness + Lizhong decoction (CDL), and cold dampness + Baitouweng decoction group (CDB) groups. The rat model of damp heat and cold damp diarrhea was established by a senna leaf + damp heat and cold damp environment. Baitouweng and Lizhong decoctions were used in the prescription and syndrome test. Clinical symptoms of rats were recorded. HE was used to observe pathological changes in spleen tissues. The densities of goblet and Paneth cells in the colon and ileum were observed by AB-PAS staining. Nrf2, HO-1, and AQP-4 expression was detected by Western Blot. Results (1)The rat model of damp heat and cold damp diarrhea was successfully established, and the clinical symptoms of damp heat diarrhea rats was obvious. (2)Baitouweng and Lizhong decoctions effectively improved clinical symptoms and spleen pathological changes of damp heat and cold damp diarrhea rats. (3)The densities of goblet and Paneth cells were decreased in diarrhea rats (P<0. 01), especially in damp heat diarrhea rats. Baitouweng and Lizhong decoction regulated the densities of goblet and Paneth cells in damp heat and cold damp diarrhea rats, respectively (P<0. 01). (4)Nrf2, HO-1, and AQP-4 expression was decreased in the colon of diarrhea rats (P<0. 01). Baitouweng and Lizhong decoctions increased Nrf2, HO-1, and AQP-4 expression in the colon of damp heat and cold damp diarrhea rats, respectively (P<0. 01). Conclusions The model of damp heat and cold damp diarrhea was successfully established, and the clinical symptoms of damp heat diarrhea rats were obvious. There was no significant difference between spleen pathological changes and the densities of goblet and Paneth cells between the two models. Compared with damp heat diarrhea rats, the changes in antioxidant indexes in cold damp diarrhea rats were more obvious. Baitouweng and Lizhong decoction had significant effects on the treatment of damp heat and cold damp diarrhea, but had no effect on the treatment of cold damp and damp heat diarrhea, which provides an Objective basis for the research on the same disease but different treatments of diarrhea.

Abstract:Objective To construct and identify macrophage-conditional Cd226 gene knockout mice, and to provide an animal model for studying the effect of CD226 on the occurrence and development of diseases by regulating the phenotype and function of macrophages. Methods Cd226^{flox/ +} mice were self-crossed by male and female to obtain Cd226^{flox/ flox} mice. Cd226^{flox/ flox} were hybridized with Lyz2-Cre⁺ mice to obtain Cd226^{flox/ +}Lyz2-Cre⁺ mice, and then crossed with Cd226^{flox/ flox} mice to obtain macrophage-conditional Cd226 knockout mice(Cd226^{flox/ flox} Lyz2-Cre⁺ ). The phenotypes of the mice were identified by PCR and agarose gel electrophoresis. To verify that CD226 had been conditionally knockdown in macrophages, qRT-PCR, flow cytometry and Western Blot were used to evaluate CD226 expression. Transwell was used to dectect the effect of CD226 on the migration of macrophages. Results The successful establishment of macrophage-conditional Cd226 gene knockout mice was confirmed at gene and protein levels. Compared with Cd226^{flox/ flox} mice, the migration ability of peritoneal macrophages was significantly inhibited in Cd226^{flox/ flox} Lyz2-Cre⁺ mice. Conclusions Macrophage-conditional Cd226 gene knockout mice were successfully established, which can provide a more accurate animal model for studying the role and mechanism of CD226 regulation of macrophage in the pathogenesis of immune diseases.

Abstract:Objective To establish a stable and efficient asthma model in beagle dogs. Methods The antigen of ragweed short (Ambrosia elatia, RS) pollen or mixed antigens of both house dust mite (Dermatophagoides farinae, HDM) and RS pollen were prepared. Healthy beagle dogs were sensitized by injecting mixed antigens within 24 hours after birth and at 1, 4, and 8 weeks. From week 1 to 12, RS pollen antigen or mixed antigens were inhaled every day (daily nebulization group, DN) or twice a week (intermittent nebulization group, IN) for airway challenge. From week 13, RS pollen antigen was inhaled once a week until week 36. At 36 and 48 weeks, an airway responsiveness test, differential counting of leukocytes in alveolar lavage fluid, and pathological analysis were carried out to evaluate the outcomes of asthma modeling. Results At 36 weeks, neutrophilic and eosinophilic inflammation in airways and airway hyperresponsiveness were observed in DN and IN groups. At 48 weeks, airway eosinophilic inflammation in DN and IN groups, and airway hyperresponsiveness in the DN group persisted. Asthma-related airway remodeling was observed in both groups. Conclusions An asthma model in beagle dogs can be established by sensitizing and challenging newborn dogs with mixed antigens of RS pollen and HDM, and increasing the frequency of antigen inhalation challenge.

Abstract:Objective To investigate whether airway inhalation of a PM2. 5 suspension at carious concentrations activates the NLRP3 inflammasome in rat myocardial tissue to induce cardiac damage and provide a reference to research drug treatments. Methods SD rats were randomly divided into a control group (Sham) and low (Low, 7. 5 mg/ kg), middle (Middle, 15 mg/ kg), and high (High, 30 mg/ kg) exposure groups with seven in each group. The exposure groups were instilled with the corresponding dose of suspension (1 mL/ kg) through a non-exposed trachea once every 6 days for 2 months, and rats in the Sham group were instilled with the same amount of normal saline. Physiological status was recorded daily. After the last infusion, the rats were fasted for 12 hours and then anesthetized with 1% sodium pentobarbital (50 mg/ kg). The cardiac function status was reflected by pathological sections of myocardial tissue, myocardial cell apoptosis, and changes in myocardial enzymes. Bcl-2 and Bax protein expression, and NLRP3, Caspase-1, and IL-1β protein and mRNA expression in myocardial tissue, and serum IL-1β, IL-6, TNF-α, and IL-18 levels reflected the NLRP3 inflammasome activation status in myocardial tissue. Results After instillation of various doses of PM2. 5 suspension, the rats in exposure groups became yellow, dull, and had decreased activity to varying degrees. There was no significant effect on diet or body weight. Decreased, irregular arrangement, nuclear pyknosis, edema. Compared with the Sham group, CK, LDH, and AST levels in myocardial tissue, Bcl-2 and IL-1β protein expression, NLRP3, IL-1β, and Caspase-1 mRNA expression, serum of IL-1β and IL-6. IL-18 and TNF-α levels were significantly increased, Bax protein expression, and the Bcl-2/ Bax ratio were decreased significantly. Cardiomyocyte apoptosis in Middle and High groups was significantly increased, and NLRP3 and Caspase-1 protein expression was significantly increased. Significantly increased; myocardial tissue apoptosis in the Low group and NLRP3 and Caspase-1 protein expression were not significantly different from those in the Sham group. Conclusions PM2. 5 exposure causes structural damage in myocardial cells, activates the NLRP3 inflammasome, promotes expression of proinflammatory factors and apoptotic proteins, and disrupts myocardial tissue functions.

Abstract:Objective To compare the effects of yeast, lipopolysaccharide (LPS), and 2,4-dinitrophenol on fever and respiratory functions in mice by a real-time temperature monitoring technique. Methods Twelve SPF C57BL/6J female mice were implanted intraperitoneally with animal core body temperature monitoring capsules. The mice were randomly divided into blank control, yeast, LPS, and 2,4-dinitrophenol groups by their body weight and temperature. After injection of the corresponding heating medium, body weight was recorded every day, and the recording frequency of the animal core body temperature monitoring capsule was times 15 min. Three days later, non-invasive respiratory monitoring was performed in mice, and specific airway resistance (sRaw), tidal volume (TV), respiratory rate (F), and airway conductivity (sGaw) were measured. At the end of the experiment, the mice were dissected, their thymus, spleen, and lungs were weighed, and organ indexes were calculated. Results During the experiment, body weight of the blank control group continued to increase, body weight in yeast and 2,4-dinitrophenol groups continued to decrease, and body weight in the LPS group increased within 48 h and decreased after 48 h. There was a hightly significant difference in the spleen index between normal and yeast groups (P<0. 01), and there were significant differences in thymus index between normal and yeast groups, and between normal and 2,4-dinitrophenol groups (P<0. 05). Body temperature in the yeast group began to increase at 3 h after injection, remained high within 48 h, and reached the maximum during 24 ~ 48 h, which was statistically different from that in the blank control group (P<0. 01). From 48 to 72 h, body temperature in the yeast group began to decrease gradually, and there was no statistical difference between yeast and blank control groups (P>0. 05). Body temperature in the LPS group increased at 4 h after injection and decreased at 8 h after injection, but it remained higher than that in the blank control group. Non-invasive respiratory monitoring showed that sRaw in yeast, LPS, and 2,4-dinitrophenol groups was increased, while TV, F, and sGaw were decreased. Conclusions From comparisons of the febrile duration, yeast as the heating medium caused the longest febrile duration, followed by LPS and 2,4-dinitrophenol. By comparing fever trends, yeast induced a state of first decreasing and then increasing temperature, reaching the peak at 10 h after injection, LPS induce a state of continuous warming, reaching the peak at 4 h after injection, and 2,4-dinitrophenol induced a state of first decreasing and then increasing temperature, reaching the peak at 8 h. From the comparison of respiratory functions, mice injected with the heating medium showed increased airway resistance, an inhibited respiratory rate, reduced tidal volume, and weakened airway conduction. By comparing the effects of the three heating media on respiration, yeast had strongest effects on the respiratory rate, tidal volume, airway resistance, and airway conduction. Therefore, yeast has a more significant effect on the duration and peak of fever and inhibits respiratory functions.

Abstract:Objective A mouse model of influenza and Staphylococcus aureus co-infection was established to evaluate the efficacy of Tamiflu and its regulatory effect on lymphocytes and inflammatory factors. Methods (1) An influenza-Staphylococcus aureus co-infection mouse model was established by screening different titers of influenza A H1N1/PR8 virus and Staphylococcus aureus. (2) Male BALB/ c mice were selected and randomly divided into six groups. On day 0, mice were inoculated intranasally with 20 μL of influenza virus H1N1/ PR8 at a dose of 0. 25 TCID₅₀. On day 3, mice were inoculated intranasally with 20 μL of Staphylococcus aureus at a dose of 2. 5 × 10⁷ CFU. Twenty-four hours after infection with influenza virus, mice were given Tamiflu by gavage once daily for 7 days, and body weight changes were recorded every day. Mice were necropsied 24 h after the last dose. The organ index was calculated; the expression of influenza virus M gene was detected by RT-qPCR; and the contents of CD3⁺ , CD4⁺ , and CD8⁺ T lymphocytes were assessed. The levels of 13 inflammatory factors, including IL-6, were detected. The average and total pro-inflammatory values of inflammatory cytokines in each group were displayed in the form of bubble charts. Results (1) Weight loss in mice infected with 0. 25 TCID₅₀ influenza virus was relatively subtle, and the mice survived. Mouse body weight decreased steadily after co-infection with 2. 5 × 10⁷ CFU, and half of the group died; (2) thus 2. 5 × 10⁷ CFU was used as the dose for the subsequent co-infection group. In co-infected mice, body weight and thymus index decreased significantly (P<0. 05), and lung index increased significantly (P<0. 05). (3) Compared with the influenza group, the co-infection group had significantly higher M gene expression (P<0. 05). Compared with the Staphylococcus aureus group, the co-infection group’s bacterial load was significantly increased (P<0. 05). Compared with the influenza group and Staphylococcus aureus group, the absolute content of lymphocytes was significantly lower, and the content of inflammatory factors significantly increased (P<0. 05). Tamiflu treatment delayed weight loss and significantly increased the thymus index (P<0. 05), and M gene expression and bacterial load were significantly decreased, while lymphocyte content significantly increased (P<0. 05). The expression of inflammatory factors was significantly decreased (P<0. 05). A bubble diagram showed much higher average and total amounts of proinflammatory factors in the co-infection group than the other groups, and Tamiflu had a good effect after timely intervention. Conclusions In this study, we established a co-infection mouse model that conformed to the clinical characteristics of the mixed virus-bacteria infection, and Tamiflu intervention reduced subsequential damage to the body.

Abstract:Objective To establish in vitro isolation and culture technology of tree shrew retina-derived microvascular endothelial cells and immortalize cell lines to provide new experimental materials for in vitro study of tree shrew retinal microvascular endothelial cells. Methods Primary retinal microvascular endothelial cells were isolated by treatment with collagenase type Ⅱ, disperse and DNase I. The cells were purified by differential digestion. Morphological observation, immunofluorescence, and karyotyping were performed on cells passaged more than 50 times. Results Microvascular endothelial cells were isolated by enzyme digestion. Purified cells had an irregular polygonal shape and pavement pattern. The morphology of cells infected with a lentivirus was consistent after subculture, and cell morphology was intact at passage 50. Immunofluorescence showed that VWF, CD34, Claudin 1, ZO-1, and SV40T were positive. The growth curve showed that the cells grew vigorously and were in the logarithmic growth phase from day 2 to 4, which reached a plateau on day 4. Karyotyping showed that the chromosome number was consistent with the chromosome number of Sino-Burmese tree shrew. Therefore, the obtained cells were immortalized retinal microvascular endothelial cells. Conclusions The immortalized cell line of tree shrew retinal microvascular endothelial cells had an intact morphology, structure and functions, which provides new experimental material for the study of retinopathy and ophthalmology-related diseases.

Abstract:Objective Through a systematic literature search, we collected, organized and analyzed the existing method for preparing animal models of functional dyspepsia and commonly used assays to improve model research method. Methods The search terms “functional dyspepsia, animal model” and “functional dyspepsia, animal model” were search CNKI, WanFang, VIP and SinoMed, PubMed, Web of Science, Cochrane Library, and Embase databases, and the search time was from the establishment of the database to June 21, 2022. Results A total of 71 articles were included, including 63 articles in Chinese and 8 articles in English. The subjects of the study were all rats, with male and female selection, and the age ranged from 1 day to 84 days. There were 32 modeling method, including 7 single-factor modeling and 25 multifactor modeling. Modeling cycles ranged from 6 to 98 days. Test indicators covered the general situation, gastrointestinal function and mental state and other major aspects. Conclusions (1) For animal models of functional dyspepsia, male SD rats of 6 ~ 8 months old are mainly used for modeling, and the modeling method are mainly single-factor tailclamping stimulation and multi-factor irregular diet+tail-clamping stimulation. The most selected detection indicators are gastric emptying/ residual rate, general conditions of the experimental animals and small intestinal propulsion ratio, respectively. (2) There is no unified regulation on modeling standard, and the selection of modeling factors needs further investigation.

Abstract:Objective To explore the effects of various modeling times on the degree of colonic injury in mice subjected to water immersion restraint stress (WIRS) and compare the degree of inflammatory injury and the composition of intestinal flora between WIRS and control mice. Methods Thirty-four Kunming mice were used in this study. Eighteen mice were randomized into control, WIRS4 h, 12 h, 24 h, 36 h, and 48 h groups. After modeling, HE staining was performed to observe the degree of colonic injury at various modeling times and evaluate the modeling conditions. Sixteen mice were randomized into control and WIRS24 h groups. Tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interferon-γ ( IFN-γ) contents were measured by enzyme-linked immunosorbent assays at the end of modeling. The composition of colonic contents was assessed by 16S rRNA gene sequencing. Results All WIRS groups exhibited various degrees of diffuse bleeding and reddening in the colonic lumen. HE staining showed inflammatory cell infiltration in WIRS mice. Colonic injury of the WIRS24 h group was obvious and the survival rate was high. TNF-α, IL-6, and IFN-γ levels were significantly increased in the WIRS24 h group (P<0. 0001). 16S rRNA sequencing showed intestinal flora disorder in the WIRS24 h group. Compared with the control group, the abundance of Campylobacter, Deferribacterota, Helicobacter, Bacteroides, Roseburia, norank_f__Ruminococcaceae, and unclassified_f__Oscillospiraceae was significantly increased (P<0. 05 or P<0. 01), and that of Actinobacteriota, Desulfobacterota, norank _f __norank _o __Clostridia _UCG-014, and Odoribacter was significantly decreased (P<0. 05 or P<0. 01). Conclusions The WIRS model causes histopathological damage to the colon in mice and significant alterations in the composition of intestinal flora.

Abstract:Objective The pathogenesis and pathological characteristics of Pkd1^{f/ f}: Cre transgenic mice were studied, and the transgenic mice were used for clinical research. Methods Diseased Pkd1^{f/ f}: Cre transgenic mice in their natural state were selected, liver and kidney tissues were collected, liver morphology under Doppler ultrasound was observed, the organ coefficient was determined, and the pathological conditions of liver and kidney tissues were observed by HE staining. Expression changes of PCNA and E-cadherin proteins in the liver and kidney were analyzed by histochemistry. Results Compared with the Pkd1^{f/ f} group, the liver weight and coefficient of the Pkd1^{f/ f}: Cre group were significantly increased (P<0. 01). There was no difference in body weight between Pkd1^{f/ f}: Cre and Pkd1^{f/ f} groups. There was no difference in the kidney weight and coefficient between Pkd1^{f/ f}: Cre and Pkd1^{f/ f} groups. ALT and AST levels related to liver functions were significantly higher in the Pkd1^{f/ f}: Cre group than in the Pkd1^{f/ f}: Cre group (P<0. 01). There was no difference in the levels of function-related factors BUN, CREA, and UA between Pkd1^{f/ f}: Cre and Pkd1^{f/ f} groups (P>0. 05). Conclusions Pkd1^{f/ f}: Cre mice develop liver cysts and disrupted liver functions, but no cysts appear in the kidneys.

Abstract:Maternal immune activation ( MIA) result from immune system stimulation during pregnancy. In addition, it can cause the offspring to develop autism spectrum disorder (ASD), schizophrenia (SZ), neural tube defects (NTD), and other mental illness, according to epidemiological research, which have indicated that MIA increases the chance of neurodevelopmental abnormalities in kids. To better understand the pathophysiology of linked mental illnesses and investigate efficient preventative and treatment options, it is crucial to construct animal models of MIA. In order to provide theoretical guidance for the pertinent experimental research and clinical therapy, this paper analyzes the selection of animals used to test MIA, the choice of modeling medications, the pathological alterations of offspring, and the applicable therapeutic options.

Abstract:The novel tank diving test is a drug screening paradigm between in vivo and cellular assays, which is specially designed for the anxiety behavioral response of zebrafish or other small fish. This paradigm was derived from the maze model in rodents and has advantages in terms of the convenience of behavior assays. Zebrafish shares high gene homology with humans. Chemically triggered anxiety and aggression in zebrafish are closely related to the hypothalamicpituitary-renal gland axis in the endocrine system, which is parallel to the effect of the hypothalamic-pituitary-adrenal axis in humans. The novel tank diving test is an efficient and reliable high-throughput behavioral screening model mainly used in studying animal sociality, addiction, sleeping, learning, and memory. The novel tank diving test has been widely used in various scenarios from initiative to continuous improvement in the previous 15 years. This article reviews the origin, measurement parameters, usage, existing problems, and perspectives of this paradigm compared with the light/ dark test to broaden the anxiety behavior evaluation paradigm and application scope.

Abstract:Zebrafish have been widely used to investigate various fields as a model organism, but relatively little attention has been focused on exploring development of its spinal region. Here, we review the development of the zebrafish spinal region from five aspects: the central nervous system (CNS), skeletal system, skeletal muscle system, associated connective tissue, and movement of the spinal region, to further our understanding of this excellent animal model and contribute to its basic research.

Abstract:Zebrafish are widely used in liver disease research because of the optical transparency of embryos, rapid development, and drug penetration into the body through the skin and gills. Hepatic fibrosis ( HF) is a pathophysiological change in abnormal proliferation of connective tissue in the liver caused by various pathological factors. HF can be caused by many chronic liver diseases. Because the signaling mechanism involved in the occurrence of HF in zebrafish is similar to that in humans, a zebrafish liver fibrosis model has been successfully established. This article discusses the relevant result of zebrafish HF model research and the current status of drug screening for liver fibrosis therapy domestically and abroad to provide guidance for research into the pathogenesis of liver fibrosis, screening for drug treatments of HF, and rational application of the zebrafish HF model.

Abstract:The zebrafish has many advantages. The individuals are small, easy to feed, have transparent embryos, and are easy to observe. The zebrafish is a model organism widely used in medical and toxicological research. Recently, because of its high throughput screening, short testing life cycle, and the high credibility of the experimental result, the zebrafish is beginning to be used to evaluate functional cosmetics. It has become an approved evaluation method in the cosmetics industry and is a hot spot for scholars. At present, the efficacy of functional cosmetics is variable, but zebrafish model evaluation criteria have not yet been finalized. In this paper, we describe the progress made in the application of zebrafish models for evaluating the efficacy of functional cosmetics, such as their whitening or soothing properties.

Abstract:Zbrafish is one of the most common model organisms. Because of its sensitivity to the external environment, zebrafish is commonly used as an ecotoxicology model and a good living tool for environmental assessment. This article reviews the research progress of zebrafish in the field of radiobiology, including the accumulation and discharge of 137Cs in water by zebrafish, the effects of γ-rays, tritium, and X-rays on growth and development of zebrafish larvae, population effects of ionizing radiation on multigenerational and intergenerational zebrafish, and the research of zebrafish in radiation protection agents, DNA damage repair proteins, and radiation sensitizers. Ultimately, this review aims to provide reference for further research of zebrafish in the field of radiobiology.