Table of Content

28 December 2020, Volume 37 Issue 06

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Effect of Overexpression of miRNA-320 on Ovarian Function in Cisplatin Induced Premature Ovarian Failure Rats 

2020, 37(06):  1. 

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Abstract: Objective To investigate the effect of overexpression of miRNA-320 on ovarian function in cisplatin induced premature ovarian failure rats. Method Thirty two SD female rats were randomly divided into 4 groups: normal control group, model group, negative control group and miRNA-320 overexpression group. During the experiment, the differences of diet, activity and body hair were observed every day. The contents of E2, FSH and LH in serum of each group were detected by ELISA. HE staining was used to observe the histological changes of ovaries. The expression levels of miRNA-320, PGRMC1 and BMP15 in ovaries of rats in each group were detected by qRT-PCR. Western blot was used to detect the protein expression of PGRMC1 and BMP15 in ovarian tissue of each group. Result Compared with the normal control group, the rats in the model group were slow in activity and in response, loss of appetite and weight, dull and lusterless body hair; histomorphological analysis showed that the ovarian tissue was obviously atrophic, the cortex became thinner, the number of atretic follicles was more, the interstitial fibrous tissue was more; the serum FSH and LH were significantly increased, while the protein and mRNA levels of PGRMC1 and BMP15 were decreased in serum E2 and ovarian tissues ( P<0. 05) . Compared with the negative control group, the activity of overexpression group was increased, the response was more agile, the food intake was increased, the body hair was gradually glossy; interstitial fibrosis was significantly reduced, the

number of mature follicles was significantly increased, the number of corpus luteum was increased; the content of serum E2, the protein and mRNA expression level of PGRMC1 and BMP-15 in ovarian tissue were increased, while the content of FSH and LH were decreased ( P<0. 05) . Conclusion Cisplatin induces ovarian dysfunction in rats with premature ovarian failure; overexpression miRNA-320 improves ovarian function in cisplatin-induced premature ovarian failure rats.

A Novel Method for Recording Intracellular Calcium Ion Concentration of Primary Neurons Using Confocal Microscopy System Combined with Electrical Stimulation

2020, 37(06):  7. 

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Abstract: Objective To detect changes in intracellular calcium ion using a detection system of spinning disk confocal microscopy combined with electrical stimulation. Method Calcium ion change is one of the key factors for neuron function. In this experiment, the fluorescent probe fluo-4AM was used to label calcium ions, neurons were excited by electrode stimuli and the spinning disk confocal was used to detect the rapid changes of intracellular fluorescence intensity. Result The electrode was successfully prepared. It was detected that about 10% of the primary neurons had spontaneous calcium ion rise. After electrical stimulation, the intracellular calcium ion rose rapidly. When the stimulation ended, the calcium ion still rise, but the amplitude decreased. Conclusion
Spinning disk confocal microscopy combined with electrical stimulation system detects rapid changes of calcium ions before and after stimulation.


Establishment and Evaluation of Myocardial Infarction Model Induced by Coronary Artery Occlusion in Ovariectomized Mice

2020, 37(06):  11. 

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Abstract: Objective To establish a mouse model that simulates myocardial infarction ( MI) in postmenopausal women. Method Female mice were ovariectomized ( OVX) and occluded the left anterior descending coronary artery to induce myocardial infarction. We evaluated the changes in mouse heart function and myocardial infarction area, and the degree of myocardial fibrosis by ultrasound assessment, hemodynamic analysis, morphological observation, immunohistochemistry and TUNEL staining. Result OVX combined with MI treatment can aggravate cardiac function impairment, significantly increase myocardial fibrosis and myocardial infarction area, induce cell apoptosis and TGF-β1 expression, and significantly increase serum pro-inflammatory cytokines levels, such as TNF-α and IL-10. These mice displayed thinning of the ventricular walls, dilation of the ventricles and remodeling of the heart. All these symptoms consistent with the menopausal women after MI. Conclusion The mouse model that mimics the occurrence of MI in postmenopausal women were successfully established, its function and histological characteristics were evaluated. It provided an ideal animal model for the study of heart protection in menopausal women.

HbA1c Can’ t Indicate the Changes of Serum Glucose Concentrations in Bama-minipigs

2020, 37(06):  17. 

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Abstract: Objective Glycosylated hemoglobin ( HbA1c) is an indicator commonly applied to clinical medicine to monitoring blood glucose control status. To investigate whether HbA1c can be used to monitoring the blood glucose control status of miniature pigs, we detected the blood HbA1c ratio in diabetes mellitus research in Bamaminipigs, so as to provide fundamental data for the minipigs using in diabetes mellitus researches. Method Fourteen Bama minipigs were divided randomly into 3 groups, which were control, HFD+STZ and HCFD+STZ group respectively. The animals in control group were fed with standard diet, and that in HFD+STZ and HCFD+STZ groups were fed with HFD diet or HCFD diet separately. After 3 months, STZ of 90 mg / kg were administered by intravenous injection. Then the pigs continued to be fed with HFD or HCFD diet. During the test, serum GLU, INS, HbA1c, GST, GLT, BUN and CRE were detected monthly. Result Four fifth of minipigs in HFD+STZ group and 4 / 5 of minipigs in HCFD + STZ group got significantly high serum glucose concentrations after injection of STZ, and sustaining to the end of the research. Nevertheless, their HbA1c ratios were not changed significantly during the
research and no apparently differences among the groups. The reason that the HbA1c ratios did not raised with the increase of serum glucose is presumed the badly permeability of pig erythrocyte for glucose. Conclusion To detect HbA1c ratio is not useful to monitoring serum glucose levels in minipigs.

PTEN Overexpression Mediated by Celastrus Orbiculatus Extracts Enhanced the Chemical Sensitivity of Cisplatin to Esophageal Cancer Cells

2020, 37(06):  22. 

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Abstract: Objective To explore the effect of PTEN over expression mediated by Celastrus orbiculatus extracts on the chemical sensitivity of esophageal cancer cells. Method The model mice of esophageal cancer were constructed, and they were treated intratumoral with Cel. orbiculatus extracts ( 1, 5 mg / kg ) and cisplatin (5 mg / kg) alone or in combination. The tumor volume was observed and the tumor weight was detected at 30 days. Immunohistochemistry detected the expression of Ki67, VEGF and Caspase 3. HET-1 A was treated with different doses of Cel. orbiculatus extracts (0. 5, 1, 2. 5, 5, 10, 20, 50, 80 and 100 μmol / L) for 24 h, and cell viability was determined by CCK8 assay. After pretreatment of EC109 / DDP cells with cisplatin ( 4 μg / mL ) , EC109 / DDP cells were treated with different doses of Cel. orbiculatus extracts ( 1, 2. 5 and 5 μmol / L ) . The control group was replaced with PBS instead of cisplatin. CCK8 was used to detect cell survival. The transwell was used to detect the invasion of cells. The expression of EMT-related protein and apoptosis-related protein was detected by Western blot. The apoptosis of cells was detected by Hoechst staining. Result After treatment with Cel. orbiculatus extracts ( 1 mg / kg and 5 mg / kg ) , the tumor weight and volume in the model mice were significantly reduced ( P< 0. 05) . The expression level of PTEN protein was significantly increased ( P < 0. 05) . Ki67, VEGF and Caspase 3 positive cells were significantly reduced ( P<0. 05) . Compared with the control group, the survival rate and invasiveness of EC109 cells were significantly decreased ( P < 0. 05 ) . Ki67, PCNA, ncadherin and Vimentin were significantly decreased ( P < 0. 05 ) . The expression levels of e-cadherin, cleaved PARP and cleaved caspase 3 protein and the apoptosis rate were significantly increased ( P <0. 05) . Cell survival rate and invasion force were significantly increased after PTEN interference ( P < 0. 05) . Apoptosis rate decreased significantly ( P < 0. 05) . Conclusion PTEN overexpression mediated by Cel. orbiculatus extracts enhanced the inhibitory effect of cisplatin on EC109 growth, invasion and EMT of esophageal cancer cells, promoted cisplatin induced EC109 apoptosis of esophageal cancer cells, and thus enhanced the chemical sensitivity of cisplatin to esophageal cancer cells.

The Effect of Compound Traditional Chinese Medicine on Lymphocyte Function and Cytokines of Mice Infected with Influenza Virus A / PR / 8 / 34( H1N1)

2020, 37(06):  30. 

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Abstract: Objective To observe the effect of compound traditional Chinese medicine on lymphocyte function and cytokines of mice infected with influenza virus A / PR / 8 / 34 ( H1N1 ) . Method Ninety BALB / c mice were randomly divided into 6 groups, 15 mice in each group, i. e. normal control group, virus control group, Amantadine hydrochloride group ( 100 mg / kg ) and compound traditional Chinese medicine high dose group (1. 5 g / kg) , medium dose group (0. 75 g / kg) and low dose group (0. 375 g / kg) ,respectively. In the traditional Chinese medicine group, the compound traditional Chinese medicine was administered intragastrically for 2 days, the normal control group was dropped nose with deionized water, the mice infection model was established by dropped nose of influenza-virus in the other 5 groups. Four hours after the model was established, each group was intragastric administrated with different dose of medicine or saline, and continuously given for 5 days. Blood sample was collected aseptically 5 days after gavage. Thymus and spleen were taken after euthanasia, then the thymus and spleen index were calculated. T and B lymphocyte transformation rates of each group were measured by MTT, and the levels of cytokines IL-6, IL-10, TNF-α and IFN-γ in lung tissue were measured by ELISA. Result Compared with normal control group, thymus index and spleen index, T and B lymphocyte A value, the level of cytokines IL-10 and IFN-γ decreased significantly, TNF-α and IL-6 increased significantly (P<0. 01) in virus control group; Compared with virus control group, spleen index and thymus index, T and B lymphocyte A values,cytokines IL-10 in medium dose group and cytokines IFN-γ in high dose groups were significantly increased,cytokines IL-6 was significantly decreased ( P < 0. 05) in medium dose group,cytokines TNF-α level was significantly decreased and IFN-γ was significantly increased (P<0. 01) in medium and low dose groups. Conclusion Compound traditional Chinese medicine can improve lung inflammation by regulating lymphocyte function and cytokine secretion level, which is one of the mechanisms of antiviral activity.

Histological Changes of Ovaries inRhesus Monkeys from Juvenile to Elderly

2020, 37(06):  37. 

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Abstract: Objective To observe the changes in the ovarian tissue of rhesus monkeys from juvenile to elderly age, and explore the mechanism of ovarian changes in rhesus monkeys with age, so as to provide a histological reference for the study of ovarian aging. Method (1) Screen 3 juvenile rhesus monkeys, 3 young rhesus monkeys, and 3 elderly rhesus monkeys. ( 2) Euthanize the rhesus monkey, take the ovarian tissue, weigh it with an electronic balance and take a picture, cut it into four horizontally, and fix it in 4% paraformaldehyde. ( 3) HE staining to observe all levels of follicles and structural changes in ovarian tissue; Masson staining to observe the degree of ovarian tissue fibrosis; Tunel staining to statistically analyze the apoptosis rate of ovarian cells; immunohistochemical staining to count the number of blood vessels. Result (1) The ovarian organ index of young rhesus monkeys was not significantly different from that of young rhesus monkeys, and both were higher than that of old rhesus monkeys. (2) Primitive, primary, secondary, and mature follicles can be seen in the ovarian tissue of juvenile and young rhesus monkeys; in the ovaries of old rhesus monkeys, only atretic follicles are seen locally, most of which are filled with adipose tissue. (3) The degree of ovarian fibrosis in young macaques is higher than that of young macaques, and the degree of fibrosis in old macaques is the highest. (4) The ovarian cell apoptosis rate of young macaques is not significantly different from that of young macaques, and both are lower than that of elderly macaques. (5) Juvenile macaques have the largest number of blood vessels, and young macaques have more blood vessels than elderly macaques. Conclusion The number of follicles in the ovarian tissue of macaques gradually decreases, the medulla is filled with a large amount of adipose tissue, and the interstitial arrangement is
scattered with age. This biological process is related to the increase of ovarian fibrosis, the increase of ovarian cell apoptosis and the decrease number of blood vessels are related.

Therapeutic Effect of Traditional Chinese Medicine on a Mouse Model of Choroidal Neovascularization

2020, 37(06):  44. 

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Abstract: Objective To observe the therapeutic effect of hexuemingmu ( HXMM ) tablet on laser-induced choroidal neovascularization ( CNV) in mice. Method C57BL / 6 mice ( aged 8 weeks, male) were selected as experimental animals. CNV mice were induced by double frequence Nd: YAG laser ( wave length: 577 nm; power: 180 mW; duration: 100 ms; spot size: 100 μm; number of light spots: 6. After generation of choroidal neovascularization, CNV mice were randomly   divided into saline group, small dose group, moderate dose group and large dose group. The other operations of the control group were the same as those of the experimental groups except laser exposure. The control group and saline group were gavaged with 10 mL / kg per day of saline. The small dose group, moderate dose group and high dose group were given 0. 34 g / kg, 0. 68 g / kg, 1. 36 g / kg HXMM ( the drug was dissolved in saline) per day respectively. Electroretinogram, fundus fluorescein angiography and choroidal neovascularization staining were performed 1 week after intervention. Result Electroretinogram showed that the amplitude of scotopic 3. 0 response b wave of intervention groups with HXMM was higher than that of saline group ( P<0. 05) . The result of fluorescein angiography in vivo showed that the degree of fundus fluorescence leakage in moderate dose group and high dose group was significantly lower than that in saline group ( P<0. 05) . The result of area of neovascularization measurement was consistent with fundus fluorescein angiography. Conclusion This study confirmed that HXMM has a certain therapeutic effect on CNV mice from aspects of function and morphology, which laid a foundation for further research on related mechanisms.

Transcriptome Analysis Associated with Mammary Gland Morphogenesis Deficiency in iRhom2^Uncv Mice

2020, 37(06):  49. 

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Abstract: Objective Mammary gland plays an important role in animal reproduction. Female iRhom2^Uncv mice showed loss of lactation and mammary gland morphogenesis deficiency. This study aims at investigating the underlying molecular basis. Method The mammary gland tissues and blood of 12-week-old wild-type and iRhom2^Uncv mice were collected. The mammary gland morphogenesis was observed by whole mounts staining. RNAseq was conducted to select the differentially expressed genes, and the serum hormone levels were analyzed by ELISA. Result Compared with wild-type mice, mammary collateral ducts in iRhom2^Uncv were sparse. Of the 18 genes with the most significant differences, 6 genes were upregulated while 12 genes were downregulated. Nearly no expression of 3 prolactin family genes were found in iRhom2Uncv both at mRNA and protein level, while the mean serum estrogen level was higher than that of wild-type mice. Conclusion Loss of expression of Zfp281 was also demonstrated. Prolactin and Zfp281 may be associated with the mammary gland morphogenesis deficiency in iRhom2^Uncv mice. Use of iRhom2^Uncv mice in research of cancer biology or neurology should be further investigated.

Study on Germicidal Effect of Micro-acid Hypochlorous Acid Solution on Dynamic Environment of Barrier Facilities

2020, 37(06):  54. 

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Abstract: Objective Study the disinfectant efficacy of microacid hypochloric acid solution on bacteria, and evaluate its disinfection effect on the dynamic environment of barrier facilities in order to obtain the disinfection strategy for dynamic environment. Method Detection of active chlorine content and pH value in microacidic hypochloric acid liquid, Staphylococcus aureus and Salmonella were used to prepare the suspension of bacteria respectively, and directly contacted with microacid hypochloric acid for certain time, then the inhibition bactericidal rate was calculated solution. This microacid hypochloric acid solution was further sprayed on the barrier facility, and the number of bacteria was measured, the germicidal rate was calculated at different time. Result The available chlorine content and pH value of slightly acidic hypochlorite solution are 6. 84 × 10- 2 g / L and 6. 24 respectively. After treatment with microacid bactericidal solution for 15 minutes inhibition bactericidal rate could reach more than 90% for those two standard strains. When microacid solution was sprayed for 15 minutes in barrier
facilities, the disinfection rate was maintained at 90% within 2 hours; 48 - 72 hours later the disinfection rate reduced less than 30%. If sprayed for 1 hour once a day,worked for 5 minutes and rest for 5 minutes and kept for 21 days the disinfection rate reached 60%; If sprayed for 30 minutes twice a day one time and kept for 21 days the disinfection rate reached more 90%. Conclusion Micro-acid hypochloric acid solutionn show perfect sterilize effect on barrier facilities, and the disinfectant rate reach more than 90% for long time use. It may be an efficient and safe microbial control strategy.

A Survey on the Sense of Belonging and Strengthening Mechanism of Non-career Staff in Experimental Animal Industry in Public Institutions

2020, 37(06):  59. 

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Abstract:The sense of belonging, as a kind of social psychological resource, has an important influence on the thought and behavior of non-career staff in colleges and public institutions. Based on the sense of belonging scale developed by Goodenow, this study investigated non-career staff in the experimental animal industry from the 3 dimensions of sense of belonging, identity and school attachment. The result show that the sense of belonging of non-career staff in the animal industry shows obvious polarization: the sense of belonging of workers with lower education level is stronger to the school, while the sense of belonging of technical staff with higher education level is weaker. Based on the analysis of the research result , 3 measures are proposed to enhance the sense of belonging of non-career personnel in the laboratory animal industry in public institutions: fairness of incentive policies, enrichment of incentive method and scientific management measures.

The Preparation and Application of the Whole Specimen of Small Rodents

2020, 37(06):  62. 

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Abstract: Objective Some new attempts and improvements have been made on the traditional method of making small animal specimens, and new production method have been formed to improve the utilization ratio of experimental animals. Method Mice were used as an example, removed the contents of the skull cavity, thoracic cavity and abdominal cavity, retained the fur, muscles and bones. After carefully plastic and fixed, 0. 4% zinc sulfate and 14% formalin are mixed into a mixed solution, and potassium chloride is added to the saturated solution for 20 days, which completed the whole animal specimen. Result The specimens of KM mice produced by this experimental animal specimen were natural, non-toxic and odorless, and the specimens were in line with the typical morphology of rodents. Conclusion The small rodent experimental animal specimen making method is more convenient than the traditional dipped specimen or exfoliation specimen, and is environmentally friendly. The experimental material is easy to get, and the cost is low. It can be popularized in the teaching and scientific popularization.

Comparison of the Transfection Effect of Different AAV Serotypes in Rat DRG Neurons

2020, 37(06):  66. 

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Abstract: Objective To compare adeno-associated virus serotypes ( AAV5, AAV6, AAV8 and AAV9 ) on efficiency of transduction of primary sensory neurons. Method Twenty four male SD rats were randomly divided into 8 groups and the dorsal root ganglion ( DRG) in rats was observed by Olympus-FV1000 after 3 weeks and 4 weeks following direct injection of AAV serotypes ( AAV5, AAV6, AAV8 and AAV9) into the DRG respectively. Result AAV9 was the most effective serotype compared with other AAV serotypes ( AAV5, AAV6 and AAV8) by immunohistochemistry staining. Conclusion Our findings suggest that AAV9 is a highly effective gene delivery vector for primary sensory neurons following direct injection into the DRG

An Overview of Osteoarthritis Animal Models

2020, 37(06):  70. 

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Abstract:Osteoarthritis is a common and degenerative disease of the joints that lead to disability frequently, which seriously threat individual health and burden social public health. There is no effective radical cure for osteoarthritis clinically due to the complex and various pathogenic factors. Developing and establishing animal models of steoarthritis are necessary and significant for studying its pathogenesis, prevention and treatment. The modeling method of OA animal model mainly includes 3 categories: spontaneous modeling, induced modeling and transgenic modeling. In recent years, more and more transgenic OA models have been used in OA study. This review mainly comprehensively describes and analyzes the existing animal models of osteoarthritis, especially transgenic OA animal model, providing clues and ideas for the further development of more ideal animal models of osteoarthritis and OA related research.

Human Angiotensin Converting Enzyme Two Transgenic Mice #br#

2020, 37(06):  70. 

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Abstract:Angiotensin-converting enzyme 2 ( ACE2) has been suggested to be the cellular receptor for 2019 novel coronavirus (2019-nCoV) . ACE2 is also a target for 2019-nCoV’ s drug therapy and vaccine development. Drug therapy and vaccine research rely on the support of animal models, and mice is the most widely used model in all kinds of disease research. As early as the attack of SARS-CoV in 2003, researchers found that the infection rate of coronavirus to mice was low due to the structural differences between mouse and human ACE2 proteins. In order to better study the pathogenic mechanism of coronavirus, scientists have established several human ACE2 ( hACE2) transgenic mice. However, being their different application techniques, integration method and mouse strains these mice display different genetic background and different phenotypes. In this paper, we introduce different of hACE2 transgenic mice, which is beneficial to the follow-up research of the disease.