ABSTRACT
SUMMARY: Systemic inflammatory response syndrome (SIRS) is a potentially fatal reaction to various forms of tissue damage and infections that cause damage to various organs. Furthermore, the brain is damaged earlier than other organs, resulting in diffuse brain dysfunction. The central clinical symptom of SIRS is delirium and emotional changes are involved in disease development. Although the amygdala is known to play a major role, the mechanisms underlying emotional changes in the early stages of SIRS have not been elucidated. Therefore, changes to dopamine levels in the amygdala were observed using an in vivo model of lipopolysaccharide (LPS)- induced SIRS to clarify the biochemical mechanisms activated in the early stages of SIRS. Extracellular dopamine was collected from the amygdala of free moving rats via microdialysis and then analyzed by high-performance liquid chromatography. In addition, emotional changes were assessed with the open field and sucrose preference tests. In the LPS group, dopamine release in the amygdala increased remarkably immediately after LPS administration, peaking at 120 min. Thereafter, dopamine release temporarily decreased, but then significantly increased again after 180 min. The present results suggest that diffuse brain dysfunction in the early stages of SIRS may involve altered dopamine levels in the amygdala.
El síndrome de respuesta inflamatoria sistémica (SRIS) es una reacción potencialmente fatal a diversas formas de daño tisular e infecciones que causan injuria a varios órganos. Además, el cerebro se daña antes que otros órganos, lo que provoca una disfunción cerebral difusa. El síntoma clínico central del SIRS es el delirio y los cambios emocionales están involucrados en el desarrollo de la enfermedad. Aunque se sabe que la amígdala desempeña un papel importante, no se han dilucidado los mecanismos que subyacen a los cambios emocionales en las primeras etapas del SRIS. Por lo tanto, en el estudio se provocaron cambios en los niveles de dopamina en la amígdala utilizando un modelo in vivo de SRIS inducido por lipopolisacáridos (LPS) para dilucidar los mecanismos bioquímicos activados en las primeras etapas del SRIS. La dopamina extracelular se recogió de la amígdala de ratas en movimiento libre mediante microdiálisis y luego se analizó mediante cromatografía líquida de alta resolución. Además, se evaluaron los cambios emocionales con las pruebas de campo abierto y de preferencia de sacarosa. En el grupo de LPS, la liberación de dopamina en la amígdala aumentó de manera notable inmediatamente después de la administración de LPS, alcanzando un máximo a los 120 minutos. A partir de entonces, la liberación de dopamina disminuyó temporalmente, pero luego volvió a aumentar significativamente después de 180 min. Los resultadosactuales sugieren que la disfunción cerebral difusa en las primeras etapas del SIRS puede implicar niveles alterados de dopamina en la amígdala.
Subject(s)
Animals , Male , Rats , Dopamine , Systemic Inflammatory Response Syndrome , Amygdala , Lipopolysaccharides/toxicity , Cytokines , Rats, Sprague-Dawley , Systemic Inflammatory Response Syndrome/chemically inducedABSTRACT
AIM:To improve the success rate of experimental modeling of non-alcoholic fatty liver(NAFLD)in rats by high-fat diet through comparing three different formulations of high-fat diets in con-structing non-alcoholic fatty liver rats model,so as to provide a reliable animal model for the study of non-alcoholic fatty liver disease.METHODS:SPF-grade male SD rats were divided into four groups randomly:control group,high-fat diet group1(HFD1),high-fat diet group2(HFD2),high-fat diet group3(HFD3).Each group was given the corre-sponding feed for 8 weeks while modeling.The da-ta on general observation,body weight changes,and ingestion of the rats were recorded during the modeling period.After 8 weeks'feeding,liver ultra-sound,CT and MRI examination were performed for the rats of each group to check the status.Blood and liver samples were collected.Changes in liver function(ALT,AST),blood lipids(TC,TG,HDL-C,LDL-C),and inflammatory indexes(IL-1β,IL-6,TNF-α)were detected.The morphology of the liv-ers was observed with the naked eyes,and the liv-er index and Lee's index were calculated at the end of 8 weeks.The effects of different high-fat diets on the establishment of NAFLD model in SD rats were comprehensively evaluated by comparing the difference of above indexes among the groups.RE-SULTS:Compared with the control group,rats in the HFD1,HFD2 and HFD3 groups showed poor mental deterioration,decreased activity,severe hair loss,decreased food intake,increased body weights,and significantly increased liver index and Lee's in-dex,along with increased liver volume,blunt edge,steatosis and lipid deposition,and the trend was even more pronounced in the HFD3 group.Com-pared with the control group,the serum levels of ALT,AST,TC,TG,LDL-C,IL-1β,IL-6 and TNF-α were significantly increased,while the contents of HDL-C was significantly decreased in the HFD1,HFD2 and HFD3 group,especially in the HFD3 group.Com-pared with the control group,the B ultrasonogra-phy showed an enlarged liver with enhanced paren-chymal echo and pipe unsharpness,CT showed that the liver and spleen CT ratio decreased obvi-ously,and the MRI images showed obvious differ-ence of liver signal intensity between in/out of phase image in the HFD1,HFD2 and HFD3 group,and the most significant imaging changes was ob-served in the HFD3 group.CONCLUSION:The above three kinds of high-fat diets can establish NAFLD model in SD rats after 8 weeks'feeding,the models induced by HFD3 was better than those in-duced by the other two groups.NAFLD lesion is rel-atively serious and expected to last longer in HFD3 group,which are more suitable for investigating the underlying mechanisms of non-alcoholic fatty liver disease and development of lipid-lowering drugs.
ABSTRACT
Objective To overcome the limitations of existing human respiratory syncytial virus(hRSV)animal models,such as semi-permissiveness and short duration of infection,this study established an IL2rg gene knockout(IL2rg-/-)rat model using TALEN gene editing technology.Methods The animal model was infected with hRSV intranasally.Clinical characteristics,body weight,and temperature changes were observed over the infection period(0～35 days).The total viral loads in respiratory organs,such as the nasal tissue,trachea,and lungs,were measured at various time points(4,11,20,and 35 days post-infection).Pathological analysis was conducted on target organs at the endpoint of observation(35 days post-infection).Changes in peripheral blood T,B,NK,and NKT cells and various cytokines were assessed at various time points(4,20,and 35 days post-infection).Results(1)IL2rg/-knockout rats sustained high viral loads in the nasal cavity upon intranasal inoculation with hRSV.The average peak titer rapidly reached 1 × 1010 copies/g in nasal tissue and 1 × 107 copies/g up to 5 weeks post-infection.(2)However,no significant pathological changes were noted in nasal,tracheal,or lung tissues.(3)An increase was observed in the content of peripheral blood B cells in hRSV-infected IL2rg--rats.(4)IL-6 and MCP-1 were increased in the early stage of infection and then decreased at the end of the observation period.Conclusions This study established a new IL2rg-/-rat model using TALEN technology and found that this model effectively supported high-level replication and long-term infection of hRSV,providing a good basis for antiviral drug screening and in vivo efficacy evaluation of anti-hRSV antibodies.
ABSTRACT
Objective To construct plasmids and knock out HIF-1α gene expression in an naked mole rat skin fibroblasts(NSF)cell line using CRISPR/Cas9 genomic editing technology,to provide an in vitro cell model for studying the mechanism of hypoxia tolerance and the occurrence and development of hypoxia-related diseases in naked mole rats.Methods We designed four pairs of single guide RNA(sgRNA)sequences targeting exons 1～4 of the NSF HIF-1αgene and successfully constructed an expression plasmid.The plasmid with the optimal sgRNA was identified and transfected into 293T cells,and the supernatant was used for detecting the virus titer.Lentivirus particles carrying sgRNAs of HIF-1α were transfected into NSF cells which express Cas9 protein,based on a previous protocol.After transfection,fluorescence signals were observed under a fluorescence microscope,and HIF-1α expression in NSF cells was detected by Western Blot and T7 endonuclease 1(T7E1)analysis.Results Sanger sequencing showed that the designed sgRNA was successfully inserted into pX459 and pKLV2-U6-sgRNA2 vectors,demonstrating successful construction of a recombinant plasmid for transfection.T7E1 digestion successfully removed three bands and the target efficiency of sgRNA was 54％.Western Blot showed that the HIF-1α gene was successfully knocked out and its protein level was significantly reduced in NSF cells from naked mole rats(P=0.0019).There were no obvious morphological changes in HIF-1α-knockout cells under the microscope,and gene knockout had no obvious effect on cell proliferation.Conclusions We successfully constructed an HIF-1α-knockout cell line using CRISPR/Cas9 technology,to provide an experimental basis for further studies of the biological function of HIF-1α,as well as the mechanism of hypoxia tolerance in naked mole rats.The result also provide a theoretical foundation for the prevention and treatment of hypoxia-related diseases.
ABSTRACT
Objective A model for studying oral ulcers induced by betel nut-extract was constructed in rats.Changes in the structure and diversity of oral flora were observed to explore the involvement of oral flora and local inflammatory factors in the pathogenesis of oral ulcers induced by betel nut-extract and to provide theoretical support for the prevention and treatment of oral ulcers in the clinic.Methods Thirty SD rats were randomly divided into normal,model and intervention groups(Guilin watermelon cream,8 mg/d for 7 days),with 10 rats/group.The oral mucosa of rats was subcutaneously injected with 10 g/mL of betel nut-extract to generate an oral ulcer model.The histomorphological changes were observed,and ulcer area and ulcer scores were assessed.Local oral tissue tumor necrosis factor-α(TNF-α),interleukin(IL)-2 and IL-8 levels were determined.Oral mucosal tissues were sampled for HE staining and analyzed for the structural distribution of oral flora and the diversity of microbial communities using high-throughput sequencing method.Results Compared with rats in the normal group,those in the model group had an increased ulcer area,significantly increased ulcer scores(P＜0.01),and significantly increased levels of TNF-α,IL-2 and IL-8 in the oral mucosal tissues(P＜0.01).The amount Streptococcus(P＜0.05)and Veillonella(P＜0.001)in the oral saliva of the model group rats was significantly reduced.The model group rats showed oral mucosal epithelial cell hyperplasia or focal necrosis,mucosal lamina propria edema,and hemorrhage accompanied by mass neutrophil and monocyte infiltration.Compared with the model group rats,the intervention group rats had significantly reduced ulcerated area(P＜0.05,P＜0.01)and ulcer scores(P＜0.05).And oral mucosal tissue levels of TNF-α(P＜0.01),IL-2(P＜0.05)and IL-8(P＜0.05),as well as significantly increased Streptococcus(P＜0.001)and Veillonella(P＜0.01)and significantly reduced Staphylococcus(P＜0.01)in the oral saliva.The degree of lesions in the oral mucosal tissues was significantly improved in the intervention group.Conclusions Betel nut-extract can be used to successfully reproduce a rat model of oral ulcer,and it is speculated that the development of oral ulcers after exposure to betel nut-extract may be related to an imbalance in the oral flora and local tissue inflammatory mediators.
ABSTRACT
Objective:To establish the rat cardiac arrest model in high-altitude hypobaric hypoxia environment, and to explore the effect of the treatment time in the hypobaric oxygen chamber on the reproduction of high-altitude rat cardiac arrest model.Methods:SPF grade healthy male Sprague-Dawley (SD) rats were used as observation subjects. The experiment was conducted in two different altitude areas. The rats from the Plateau Branch of Institute of Cardiopulmonary and Cerebral Resuscitation of Sun Yat-sen University (Xining, Qinghai) were weighed and numbered, and they were placed in a hypobaric oxygen chamber (simulated altitude of 3 000 meters, speed of ascent and descent of 15 m/min, temperature of 20 ℃, cabin pressure of 69.5 kPa, cabin oxygen pressure of 14.5 kPa). After 30 days of feeding, the rats were obtained according to random number table method, and the cardiac arrest model was established by asphyxia method as the 30-day hypobaric hypoxia group. After 60 days of feeding, rats were randomly selected again, and the cardiac arrest model was established as the 60-day hypobaric hypoxia group. Thirty rats were randomly selected from the Institute of Cardiopulmonary Cerebral Resuscitation at Sun Yat-sen University (Guangzhou, Guangdong) by the same method, and the cardiac arrest model was established as the plain control group. The differences in the body weight of rat modeling precursors and the induction time of asphyxia during the modeling process among different groups were compared.Results:Finally, cardiac arrest model was established in 16 rats in the 30-day hypobaric hypoxia group and in 22 rats in the 60-day hypobaric hypoxia group. There was no significant difference in the body weight of rats before modeling among the plain control group, 30-day hypobaric hypoxia group and 60-day hypobaric hypoxia group [g: 429.00 (389.25, 440.75), 440.00 (415.50, 486.25), 440.00 (400.00, 452.50), all P > 0.05]. The asphyxia induction time of rats in the 60-day hypobaric hypoxia group was significantly longer than that in the 30-day hypobaric hypoxia group (s: 294.59±75.39 vs. 234.31±93.86, P < 0.01), even about 1.4 times of the plain control group (s: 294.59±75.39 vs. 208.73±30.88, P < 0.01). There was no significant difference in the asphyxia induction time between the 30-day hypobaric hypoxia group and the plain control group ( P > 0.05). Conclusion:Rats treated in a hypobaric oxygen chamber for 60 days are more suitable for the preparation of high-altitude cardiac arrest model, and are also consistent with the oxygen reserve and hypoxia tolerance of high-altitude rats.
ABSTRACT
Objective:To investigate the expression of hippocampal synapse-related proteins including synaptophysin (SYN), postsynaptic density protein 95 (PSD95) and growth-associated protein 43 (GAP43) in rats with epilepsy accompanied by depression.Methods:The 3-month-old female clean grade SD rats were selected for the experiment.Lithium chloride pilocarpine was used to establish an epileptic rat model. Rats with successful epilepsy models were divided into epileptic depressive group (EWD group)and epileptic group with 10 in each group based on whether they were accompanied by depression. Furthermore, ten rats with matched body mass were taken as the depressive group and 10 were taken as control group. As for the depressive group rats, chronic unpredictable mild stress stimulation combined with orphanage was adopted to establish a model of depression.The depressive behaviors of rats were evaluated by body mass, sucrose preference test and open field test. Immunohistochemical staining and Western blot were used to detect the expression of SYN, PSD95 and GAP43 proteins in rat hippocampal tissue. SPSS 17.0 software was used for data statistical analysis, repeated measurement ANOVA was used for behavioral results, one-way ANOVA was used for inter group comparison of protein expression data, and LSD test was used for further pairwise comparison.Results:As for the body mass, there was significant interaction effect between the time and group among the 4 groups ( F=7.33, P<0.01). On the 8th day and the 29th day, the body weight of rats in the EWD group and the depressive group were lower than those in the epilepsy group (all P<0.05). The body weight of rats in the EWD group on the 29th day was lower than that on the first day ( P<0.05). As for the sucrose preference rates, there was significant interaction effect between the time and group among the 4 groups( F=2.67, P<0.05). The sucrose preference rate of EWD group on the15th and 29th day were lower than that on the first day (both P<0.05). The results of the open field test showed that the interaction effects of the number of vertical standing times( F=2.74) and the number of horizontal movement lattices ( F=1.76) both were not significant (both P>0.05), but both the time effect and group effect were significant (vertical standing times: Ftime=4.35, P<0.05, Fgroup=25.64, P<0.01; horizontal movement lattices: Ftime=12.75, P<0.01, Fgroup=21.37, P<0.01). The immunohistochemical results showed that there was a statistically significant difference in the number of positive cells expressing synaptic proteins SYN, PSD95 and GAP43 among the four groups of rats ( F=93.85, 58.66, 98.84, all P<0.05). The numbers of positive cells of SYN (11.73±4.30), PSD95 (24.47±7.58) and GAP43 (9.40±3.50) in the epilepsy group were lower than those in the control group ((51.00±15.39), (55.60±13.17) and (29.53±4.05)) (all P<0.05). The numbers of positive cells of SYN (5.80±3.53), PSD95 (12.87±4.03) and GAP43 (5.33±3.50) in the EWD group were lower than those in the depressive group ((11.33±3.22), (48.13±12.69) and (15.47±5.21) )(all P<0.05). Western blot results showed that there were statistically significant differences in the expression of synaptic proteins SYN, PSD95 and GAP43 among the four groups of rats ( F=13.19, 9.38, 16.80, all P<0.05). The expression levels of SYN, PSD95 and GAP43 in the epilepsy group were lower than those in the control group (all P<0.05). The expression levels of SYN, PSD95 and GAP43 in the EWD group were lower than those in the epilepsy group and the depressive group (all P<0.05). Conclusion:The low expression of SYN, PSD95 and GAP43 proteins in the hippocampus of rats with epilepsy accompanied by depression may be related to their pathogenesis.
ABSTRACT
Objective:To explore the effects of activating α7 nicotinic acetylcholine receptor(α7nAChR)on cognitive function and polarization of hippocampal microglia in traumatic brain injury (TBI) rats.Methods:Totally 36 male SD rats with 6-8 weeks old were randomly divided into Sham group ( n=12), TBI group ( n=12), TBI+ α7nAChR agonist group ( n=6) and TBI+ α7nAChR antagonist group( n=6). The TBI model was established by the " free fall impact" method. From the 4th to 6th day after modeling, mice in the TBI+ α7nAChR agonist group were intraperitoneally injected with α7nAChR agonist PNU-282987 (3 mg/kg). Rats in TBI+ α7nAChR antagonist group were intraperitoneally injected with α7nAChR antagonist methyllycaconitine citrate (5 mg/kg) first, then 45 minutes later they were injected with α7nAChR agonist PNU-282987 (3 mg/kg). Rats in the TBI group and Sham group were intraperitoneally injected with an equal volume of 0.9% sodium chloride solution. Morris water maze test was used to evaluate the learning and memory function of rats. Immunofluorescence staining was used to observe the ionized calcium binding adapter molecule 1 (Iba-1)(a marker for microglia) and arginase 1 (Arg-1)(a marker for M2 microglia). Western blot was used to detect the protein level of Arg-1 in hippocampal tissue. Statistical analysis was performed using GraphPad Prism 9 software. Independent sample t test was used for comparison between two groups, one-way ANOVA was used for comparison among multiple groups, and Tukey test was used for multiple comparison. Results:The results of the water maze test showed that after 7 days of modeling, there was a statistical difference in the escape latency among the 4 groups of rats ( F=6.134, P<0.05). There was no statistical difference in the escape latency between the TBI group and the TBI+ α7nAChR antagonist group( P>0.05), but the both were higher than that of the Sham group (both P<0.05). The escape latency of the TBI+ α7nAChR agonist group((31.87±9.01)s) was shorter than that of the TBI group((56.75±2.62)s) and the TBI+ α7nAChR antagonist group((60.00±0.00)s) (both P<0.05). The results of immunofluorescence staining showed that there were statistical differences in the fluorescence intensity and cell numbers of Arg-1 + /Iba-1 + among the four groups ( F=17.37, 9.33, both P<0.05). The immune fluorescence intensity (0.27±0.03) and cell numbers (21.67±4.41) of Arg-1 + /Iba-1 + in the TBI+ α7nAChR agonist group were higher than those in the TBI group((0.14±0.03), (11.33±2.60)) and TBI+ α7nAChR antagonist group((0.10±0.03), (7.67±1.20)) (all P<0.05). The results of Western blot showed that there was a statistical difference in the level of Arg-1 protein in hippocampus among the 4 groups ( F=8.323, P=0.001). There was no significant difference in the level of Arg-1 protein between the TBI group and the TBI+ α7nAChR antagonist group( P>0.05), and the level of Arg-1 protein in the TBI+ α7nAChR agonist group(1.06±0.22) was higher than that in the TBI group(0.60±0.13) and TBI+ α7nAChR antagonist group(0.35±0.10) (both P<0.05). Conclusion:Activating α7nAChR can promote the polarization of M2 type microglia in rat hippocampal tissue and improve the learning and memory function of TBI rats.
ABSTRACT
Objective:To investigate the effects of repetitive transcranial magnetic stimulation (rTMS) on learning-memory and the expression of synaptic plasticity proteins in hippocampus of rats with post-stroke sleep deprivation.Methods:A total of 28 SPF grade healthy male Wistar rats with 8-week old were randomly divided into four groups (control group, sham operation group, model group and rTMS group) according to random number table method, with 7 rats in each group. The rats in the model group and the rTMS group were treated with middle cerebral artery occlusion and p-chlorophenylalanine intraperitoneal injection to establish the post-stroke sleep deprivation model. The rats in the rTMS group were treated with rTMS intervention for consecutive 14 days after modeling. The rats in the sham operation group were only separated arteries but not ligated and inserted. The rats in control group were fed normally. The open field test (OFT) was used to observe the autonomous behavior of rats.The water maze test(WMT) was used to observe the spatial learning and memory ability of rats.The content of tyrosine kinase receptor type B(TrkB) in hippocampus was detected by Western blot.The expressions of brain-derived neurotrophic factor(BDNF) and immediate early gene c-fos in hippocampus were detected by immunofluorescence.The morphology and structure of neurons in hippocampus were observed by optical microscopy and transmission electron microscopy. SPSS 21.0 software was used for statistical analysis, and repeated measurement ANOVA was used for the escape latency data, one-way ANOVA was used for the comparison of other data among multiple groups, and LSD test was used for further pairwise comparison.Results:(1) The OFT results showed that there were statistical differences in the numbers of crossing squares, upright times and total points of rats in the four groups after intervention ( F=27.638, 10.425, 30.690, all P<0.001). The numbers of crossing squares ((72.71±10.10)), upright times ((6.57±0.87)times) and total points ((79.29±10.03) points) of rats in rTMS group were all higher than those in model group after intervention ((43.71±6.96), (3.43±0.65)times, (47.14±6.82)points) (all P<0.05). As for the escape latency of WMT among the four groups of rats, the interaction effect was not significant( F=1.108, P=0.37), and the time main effect( Ftime=27.295, Ptime<0.01) and group main effect ( Fgroup=8.691, Pgroup<0.01) were significant after rTMS intervention.On the 3rd and 4th day, the escape latency of rTMS group rats was lower than that of the model group (both P<0.01). There were statistically significant differences in the numbers of crossing platform, swimming distance and residence time in target quadrant of rats in the four groups after intervention( F=8.569, 3.308, 3.547, all P<0.05). The numbers of crossing platform ((2.00±0.31)times), swimming distance in target quadrant ((196.95±24.57) cm) and residence time ((17.72±1.36)s) of rats in rTMS group were all higher than those in model group after intervention ((1.57±0.30)times, (146.61±4.79) cm, (13.58±0.98)s)(all P<0.05). (2)Optical microscopy and transmission electron microscopy showed that the hippocampal cells arranged irregularly, the organelles' integrity was destroyed in the model group compared with the normal control group. In rTMS group the arrangement and structure of nerve cells in the hippocampus were improved after rTMS intervention. (3) The immunofluorescence results showed that c-fos (1.49±0.09) and BDNF (0.84±0.06) in the hippocampus of rats in rTMS group were both higher than those in model group ((1.24±0.12), (0.48±0.08))(both P<0.05). The Western blot results showed that the expression level of TrkB (1.81±0.03) in the hippocampus of rats in rTMS group was higher than that in model group (0.96±0.02) ( P<0.05). Conclusion:The rTMS can improve the learning-memory ability and autonomous capacity of rats with post-stroke sleep deprivation, which may be related to promoting the expression of c-fos, BDNF and TrkB in hippocampus tissue.
ABSTRACT
A Helmholtz coil is selected to generate the alternating magnetic field,and the relevant model is established with a rat brain tumor as the research object.Based on the Pennes bioheat transfer equation,the electromagnetic field distribution and temperature field distribution are calculated in COMSOL simulation software,and the factors affecting the outcome of magnetic induction hyperthermia are analyzed.The results show that both magnetic field distribution and temperature field distribution meet the requirements for magnetic induction hyperthermia.The magnetic field distribution in the tumor treatment area is uniform,and the central magnetic induction strength is 12.847 mT.The temperature rise in the tumor area is significant,and the temperature at the tumor center is 46℃or above,basically reaching the treatment temperature.The therapeutic efficacy of magnetic induction hyperthermia is affected by the number of turns,current,radius and spacing,magnetic field frequency and other parameters.The study provides reference for the clinical application of magnetic induction hyperthermia and the coil design.
ABSTRACT
Objective:To investigate the correlation between liver stiffness and histopathological changes in a rat model of acute hepatitis using virtual touch tissue imaging quantification (VTIQ) technology.Methods:A total of 100 SPF-grade SD rats were randomly divided into 3 groups: control ( n=30), low-dose ( n=35), and high-dose ( n=35) groups. Acute hepatitis models were induced in the low-dose and high-dose groups using 400 mg/kg and 600 mg/kg of Thioacetamide (TAA), respectively. Liver stiffness parameters of the right median lobe and right lobe were measured using VTIQ technology, Mean-H and Mean-L represent the liver lobes with higher and lower liver stiffness measurments, respectively, while Mean represent the average of the measurements from both liver lobes. Comparative analyses of liver stiffness parameters were performed across three groups and between the two lobes of the liver. The correlations between the Mean values of liver stiffness and semi-quantitative histopathological data were investigated. Ten rats were randomly selected from each of the 3 groups to test the repeatability of VTIQ values before and after euthanasia with intraperitoneal anesthesia. Subsequently, 10 rats after euthanasia from each 3 group were randomly chosen to assess the repeatability of VTIQ measurements for inter-observer and intra-observer variabilities. Results:VTIQ results showed statistically significant differences in Mean, Mean-H, and Mean-L among the 3 groups (all P<0.01). The high-dose group had higher measurements compared to the low-dose and control groups, with significant intergroup differences (all P<0.01). Significant differences in Mean-H and Mean-L were observed between the two liver lobes in both low and high-dose groups (all P<0.01). The Mean value showed significant positive correlations with semi-quantitative histopathological data of hepatocellular edema, periportal inflammatory cell infiltration, macrophage proliferation, and bile duct proliferation ( r=0.391, 0.648, 0.577, 0.542; all P<0.01). Multivariate linear regression analysis indicated that hepatocellular edema, eosinophilic change, and bile duct proliferation significantly and positively predicted the Mean value (β=-0.278, -0.196, -0.333; all P<0.05). There were no significant differences of VTIQ measurements befor and after euthanasia (all P>0.05), with repeatability coefficients of 0.166, 0.182, 0.185 for Mean, Mean-H, and Mean-L, respectively. Post-euthanasia, inter- and intra-observer VTIQ differences remained non-significant (all P>0.05), with Mean, Mean-H, Mean-L coefficients of 0.114, 0.194, 0.165 and 0.206, 0.322, 0.268, respectively. Conclusions:VTIQ technology demonstrates potential clinical value in assessing a rat model of acute hepatitis, offering a new perspective for non-invasive evaluation of acute hepatitis. However, its clinical application requires further validation.
ABSTRACT
Objective To investigate the effect of YTH domain family protein 2(YTHDF2)on an-giotensin Ⅱ(Ang Ⅱ)-induced hypertrophy and apoptosis of primary neonatal rat cardiomyocytes.Methods The expression level of YTHDF2 was detected in the primary neonatal rat cardiomyo-cytes with or without Ang Ⅱ stimulation(AngⅡ group and normal group).The cells were divid-ed into blank group(transfected with siRNA+PBS),siYTHDF2 group(transfected with siYTHDF2+PBS),model group(siRNA+Ang Ⅱ)and experimental group(siYTHDF2+Ang Ⅱ)to investi-gate the effects of silencing YTHDF2 on the hypertrophy and apoptosis of cardiomyocytes.West-ern blotting and RT-qPCR were used to detect the expression of YTHDF2 at protein and mRNA levels,and RT-qPCR was employed to measure the mRNA levels of myocardial hypertrophic related genes atrial natriuretic peptide(ANP),brain natriuretic peptide(BNP)and beta-myosin heavy chain(β-MHC),and cardiomyocyte apoptosis related genes Bax and B lymphocytoma 2 gene(Bcl-2).The surface area of cardiomyocytes was observed by α-actin immunofluorescence staining.Cardiomyocyte apoptosis was observed by TUNEL staining,and the binding relationship between YTHDF2 and Bcl-2 was verified by immunoprecipitation.Results The expression of YTHDF2 at protein and mRNA levels were significantly higher in the AngⅡ group than the nor-mal group(1.49±0.03 vs 0.97±0.09,1.50±0.08 vs 1.00±0.07,P＜0.05).Compared with the blank group,the surface area of cardiomyocytes was notably enlarged,apoptotic rate was obvi-ously increased,the mRNA levels of ANP,BNP,β-MHC and Bax were significantly increased,and that of Bcl-2 was remarkably decreased in the model group(P＜0.05).The experimental group obtained decreased surface area and apoptotic rate of cardiomyocytes,lower mRNA levels of ANP,BNP,β-MHC and Bax,and increased mRNA expression of Bcl-2(P＜0.05).Conclusion Silencing YTHDF2 can alleviate Ang Ⅱ-induced hypertrophy and apoptosis in primary neonatal rat cardiomyocyte,and YTHDF2 inhibits the expression of Bcl-2 by binding to it.
ABSTRACT
Objective To establish an allogeneic rat model of endometriosis and to evaluate the effects of gonadotropin-releasing hormone (GnRH) agonist GenSci006 on experimental rat endometriosis. Methods Endometrium from SPF grade donor female SD rats were transplanted onto the abdominal wall of recipient female rats to construct an allogeneic endometriosis model. The rats undergoing sham surgery were divided into the sham group. Three weeks later, the length, width and height of the ectopic endometrium were measured, and the volume of the endometrium (V1) was calculated before drug administration. The modeling rats were randomly divided into four groups: model group, triptorelin group (0.25 mg/kg), GenSci006-1 group (0.125 mg/kg) and GenSci006-2 group (0.25 mg/kg). Each group had 16 rats and received a single dose of the corresponding drug. The sham group and model group were administered an equal volume of solvent. Three weeks after administration, ectopic endometrium was measured to calculate the volume V2 and inhibition rate. The effect of GenSci006 on rat uterus and ovarian tissues was assessed by comparing organ coefficients and changes in pathological sections. Enzyme-linked immunosorbent assay (ELISA) was used to measure the levels of serum estradiol (E2), progesterone (P4), follicle stimulating hormone (FSH), and luteinizing hormone (LH). Real-time fluorescent quantitative PCR was used to detect the expression of GnRH receptor (GnRHR) mRNA in the hypothalamus and pituitary. Western blot was used to detect the expression of estradiol receptor alpha (ERα), beta (ERβ) and progesterone receptor (PR) in ectopic endometrium. Results Three weeks after administration, compared with the model group, the body weight of rats in the triptorelin and GenSci006-2 groups significantly increased (P < 0.05), while the volume of ectopic endometrium significantly decreased (P < 0.05). Compared with the sham group, the model group showed no significant changes in uterine and ovarian organ coefficients or endometrial thickness (P > 0.05). Compared with the model group, the uterine organ coefficients and endometrial thickness were significantly reduced in the triptorelin and GenSci006-2 groups (P < 0.05). Compared with the sham group, the serum levels of E2, P4, FSH and LH in the model group showed no significant changes (P > 0.05). Compared with the model group, the ovarian organ coefficient and serum P4 levels of rats in the Triptorelin, GenSci006-1, and GenSci006-2 groups were significantly reduced (P < 0.05), while the serum LH levels of rats in the GenSci006-1 group were significantly increased (P < 0.05). However, there were no significant changes in serum E2 and FSH levels in each group (P > 0.05). Compared with the model group, the expression levels of GnRHR mRNA in the pituitary tissue of rats in the triptorelin and GenSci006-2 groups were significantly downregulated (P < 0.05), with no significantly changes in the hypothalamus (P > 0.05). There were no significant changes in the expression level of GnRHR mRNA in the hypothalamus or the protein levels of ERα, ERβ and PR in the ectopic endometrial tissue in any group (P > 0.05). Conclusion The allogeneic endometriosis rat model is a suitable animal model for screening and evaluating drugs for treating endometriosis. The volume of ectopic endometrium, inhibition rate, uterine and ovarian organ coefficients, and serum E2 levels may serve as indicators for detecting drug efficacy.
ABSTRACT
【Objective】 To analyze the effect of fisetin against venous thrombosis in rats. 【Methods】 Seventy SD rats were randomly divided into the following groups: sham-operation group, model group, fisetin 45 mg/kg, 15 mg/kg, 5 mg/kg groups, and aspirin group (47 mg/kg). The corresponding medication was administered by gavage once a day consecutively (the sham-operation group and the model group were given 0.5% carboxymethyl cellulose sodium solution with 10 mL/kg, respectively) for 7 consecutive days. One hour after the last administration, the rats were anesthetized, the lower part of the intersection of inferior vena cava and left renal vein was ligated with silk thread (no ligation in the sham-operation group), and the abdominal wall was sutured. Two hours later, the abdominal cavity was reopened, the other venous branches 1.5 cm away from the ligation site were closed with the artery clamp, and blood was collected from the abdominal aorta. The anticoagulant ratio of 3.8% sodium citrate∶whole blood was 1∶9.The venous thrombus 1 cm down from the ligation point of the intersection of inferior vena cava and left renal vein was cut and the thrombus was separated. The residual blood was dried with filter paper, weighed and recorded. Plasma was taken after anticoagulant blood centrifugation. The levels of plasma antithrombin-Ⅲ (AT-Ⅲ), protease C (PC), plasminogen (PLG), and plasminogen activator inhibitor (PAI-1) were detected by ELISA kits. 【Results】 Compared with the model group, the weight of thrombus in fisetin 45 mg/kg group and aspirin 47 mg/kg group decreased (P<0.01). The content of AT-Ⅲ in three fisetin groups increased (all P<0.05). The content of PC in fisetin 45 mg/kg increased (P<0.05). The content of PLG and PAI-1 in fisetin 45 mg/kg group decreased (both P<0.05). 【Conclusion】 Fisetin has the effect against venous thrombosis in vivo, and the effect is related to the upregulation of AT-Ⅲ and PC and the downregulation of PLG and PAI-1.
ABSTRACT
Establishment of rat models of liver transplantation provides an ideal animal model for resolving the problems of postoperative complications and perioperative treatment of liver transplantation. With in-depth study of the establishment of rat models of liver transplantation, classic "two-cuff" technique has been gradually employed. However, poor surgical field, vascular torsion, biliary tract injury and long anhepatic phase remain unresolved in the process of liver transplantation using traditional techniques. At present, the rat models of liver transplantation at home and abroad are modified mainly from the reconstruction of four vital anatomic structures including the suprahepatic inferior vena cava, portal vein, infrahepatic inferior vena cava and bile duct. Therefore, the latest progress in the reconstruction of the suprahepatic inferior vena cava, portal vein, infrahepatic inferior vena cava and bile duct was reviewed, aiming to provide reference for the establishment of rat models of liver transplantation and promote further development of liver transplantation techniques.
ABSTRACT
Objective:Glaucoma is a multifactorial optic neuropathy with a high rate of irreversible visual loss,and its pathogenesis is complex and still unclear.Elevated intraocular pressure(IOP)is well recognized as the sole modifiable risk factor for the development of glaucoma in the majority of cases.This study aims to compare 2 different methods of inducing chronic ocular hypertension by circumlimbal suture or by laser burns in degree and lasting time of the IOP,different status of the retina and retinal ganglion cells(RGCs),and changes of the microstructure of neurons. Methods:The chronic ocular hypertension models were induced by 2 different ways.One kind of the models was built by unilateral circumlimbal suture(10/0)implantation(suture group),another kind of model was built by laser burns at trabecular meshwork and episcleral veins(laser group).The untreated contralateral eye served as the control group.Changes in IOP were observed and regularly monitored in the 2 groups of rats.HE staining was applied to observe the retinal and optic nerve pathology.Transmission electron microscope(TEM)was used to observe the mitochondrial morphology.RGCs were specifically labeled with Brn3b antibody and counted.The expression of caspase-3 was detected by Western blotting to clarify the apoptosis of RGCs. Results:Compared with the control group,IOP were significantly increased in the suture group and the laser group(both P<0.05).The suture group induced a 1.5-fold elevation of IOP,and sustained for 8 weeks.The laser group induced a 2-fold elevation of IOP for 12 weeks.Both methods could cause RGCs loss(both P<0.05),which were verified by pathology and immune staining of Brn3b.The expressions of caspase-3 were also increased(both P<0.05).The mitochondrial morphology became more fragment,which changed from long shape to round and small one under TEM in 2 models.For comparison,the pathology changes of retinal structure in suture group were not obviously than those in the laser group. Conclusion:Circumlimbal suture can build an effective model of chronic elevated IOP and induce glaucomatous pathologic changes similar to those in the laser photocoagulation,but the pathologic changes are milder than those in laser photocoagulation.Compare with translimbal laser photocoagulation,equipment and skill demand for circumlimbal suture is less.
ABSTRACT
Objective To investigate the effect of rats'injuries and its mechanism caused by specific dose of radiation combined with decompression exposure.Methods 81 male SD rats were randomly divided into control group(n=9),radiation group(n=18),radiation+low-load decompression group(n=18),radiation+medium-load decompression group(n=18),and radiation+high-load decompression group(n=18).In addition to control group,the rats were irradiated with 60Co γ rays at 4 Gy and then underwent rapid escape experiments.The high-pressure exposure schemes were to stay underwater 57 m for 30 min,45 min or 60 min and reduce to normal pressure within(30±5)s,respectively.The high-pressure exposure was not carried out in radiation group.The behavior and death of rats in each group were observed 0.5 h after leaving the cabin.Blood(abdominal aorta)and lung tissues were collected at 3 h and 72 h,respectively.The changes of lung wet-dry weight ratio(W/D),lung pathology and serum levels of interleukin(IL)-1β,IL-6,tumor necrosis factor-α(TNF-α),superoxide dismutase(SOD),malondialdehyde(MDA),nitric oxide(NO),intercellular adhesion molecule-1(ICAM-1)and thromboxane B2(TXB2)were analyzed.Results Compared with control group and radiation group,radiation+low-load decompression group showed no significant difference in the injury and death rate of rats(P＞0.05),while radiation+medium-load decompression group and radiation+high-load decompression group showed significantly increase of the injury and death rate of rats(P＜0.05).Compared with control group,other groups showed no significant change in pulmonary W/D at 3 h(P＞0.05),and increased at 72 h(P＜0.05).HE staining showed that compared with control group,radiation group showed mild lung interstitial edema,while radiation+low-load decompression group showed obvious pulmonary tissue edema and a small number of red blood cells exudated in the alveolar cavity.The edema,congestion and inflammatory cell infiltration of lung tissue were more serious in radiation+medium-load decompression group and radiation+high-load decompression group.Compared with control group and radiation group,all radiation+decompression groups showed an increase in serum levels of IL-1β,IL-6,TNF-α,MDA,NO,ICAM-1 and TXB2(P＜0.05),and a decrease in SOD activity(P＜0.05).Compared with radiation+low-load decompression group,radiation+medium-load decompression group and radiation+high-load decompression group showed increase in serum levels of IL-1β,IL-6,MDA,ICAM-1 and TXB2(P＜0.05),and decrease in activity of SOD(P＜0.05).Except for control group,serum levels of IL-1β,IL-6,TNF-α,MDA,NO,ICAM-1 and TXB2 were decreased at 72 h compared with 3 h(P＜0.05),and SOD activity was increased at 72 h in all groups(P＜0.05).Conclusions High-load decompression can increase the injury and death rate of rats exposed to radiation and high pressure.The potential mechanism of the combined injury effect of radiation and decompression was related to inflammation,immune stress,oxidative damage,vasomotor activity and coagulation mechanism.
ABSTRACT
Objective To investigate the effects of neuroligin-1,-2(NLGN-1,-2)on oligodendrocyte(OLs)differentiation and myelination in the central nervous system.Methods OLs were cultured in vitro in the presence of different concentrations of NLGN-1 and NLGN-2.Morphological differentiation of OLs was observed by immunofluorescent staining and mRNA expression levels of myelin-associated genes were detected by Real-time PCR.Western blotting was used to detect the expression of myelin-related proteins.Results NLGN-1,-2 accelerated the differentiation of oligodendrocyte precursor cells(OPCs)into mature OLs,and promoted the ability of myelin sheath formation.In vitro culture conditions,the dosage of 500 μg/L had the best promotion effect on OLs differentiation and maturation,and NLGN-2 had better promoting effect than that of NLGN-1.Furthermore,the mRNA expression levels of myelin-associated genes myelin protein P0(MPZ),myelin basic protein(MBP)increased after the neuroligins treatments detected by Real-time PCR.Western blotting result showed that the expressions of MBP and MPZ increased significantly after 500 μg/L treatment with NLGN-1 and NLGN-2 for 12 hours.Conclusion NLGN-1,-2 promote OLs differentiation and myelination.The positive effect of NLGN-2 is greater than that of NLGN-1 significantly,suggesting that the treatment with inhibitory synaptic-associated cytokines may improve the ability of myelin sheath formation in the central nervous system.
ABSTRACT
Objective To investigate the protective effect and mechanism of acellular nerve allografts(ANA)combined with electroacupuncture on spinal ganglia in rats with sciatic nerve injury(SNI).Methods Totally 50 male adult SD rats were randomly selected for this experiment.Ten rats were prepared for the ANA.Forty male SD rats were randomly divided into normal group,model group,ANA group and combinational group,with 10 rats in each group.The SNI model was established by cutting off the nerves 10 mm at the 5 mm on the inferior border of piriformis after separating the right sciatic nerves.The rats in the ANA group were bridged with ANA to the two broken ends of injured nerves.The rats in the combinational group were treated with electroacupuncture 2 days after ANA bridging,Huantiao(GB30)and Yanglingquan(GB34)were performed as the acupuncture points,each electroacupuncture lasted 15 minutes and 7 days as a course of treatment,4 courses in all.Sciatic nerve conduction velocity was measured by electrophysiology to evaluate the regeneration of damaged axons.Morphology of spinal ganglia was observed by Nissl staining.The expression of nerve growth factor(NGF)and brain-derived neurotrophic factor(BDNF)were detected by Western blotting and immunofluorescent staining.Results Compared with the normal group,the sciatic nerve conduction velocity in model group decreased significantly(P<0.01),Nissl bodies in neurons of spinal ganglia were swollen and dissolved,with incomplete structure and the number decreased dramatically(P<0.01),while the level of NGF and BDNF also decreased significantly(P<0.01).Compared with the model group,the sciatic nerve conduction velocity in ANA and combinational groups strongly increased(P<0.01),the damage of Nissl bodies in neurons of spinal ganglia reduced and the number obviously increased(P<0.01),the level of NGF and BDNF increased considerably(P<0.01).Compared with the ANA group,the sciatic nerve conduction velocity in combinational group increased significantly(P<0.01),the morphology of Nissl bodies in neurons of spinal ganglia were more regular and the number increased(P<0.01),moreover,the level of NGF also increased significantly(P<0.01).Conclusion ANA combined with electroacupuncture can enhance the sciatic nerve conduction velocity,improve the morphology of neurons in spinal ganglia and play a protective effect on spinal ganglia.The mechanism can be related to the higher expression of NGF and BDNF proteins,especially the expression of NGF protein.