Table of Content

2022, Volume 53 Issue 1
06 February 2022 Previous Issue    Next Issue
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Technology and Methodology
Cell and Molecules Biology
Anatomy
Anthropology
Histology,Embryology and Developmental Biology
Cancer Biology

Cancer Biology



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Technology and Methodology

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Construction of aquaporin 9 gene knockout mice using CRISPR/Cas9 gene editing system CHENG Quan-cheng FAN Jing LIU Huai-cun DING Hui-ru FANG Xuan WANG Jian-wei CHEN Chun-hua ZHANG Wei-guang 2022, 53 (1):  126-131.  doi: 10.16098/j.issn.0529-1356.2022.01.018 Abstract ( )   PDF (5078KB) ( )   Objective  To construct homozygous aquaporin 9(AQP-9)-/- mice using the CRISPR/Cas9 system.    Methods  According to the design principle of CRISPR/Cas9 target, the exon region of the AQP-9 gene sequence was found in the Ensembl database. AQP-9-202 exon was selected through comprehensive analysis. In the early stage, 7 small guide RNA(sgRNA) targets were designed on both sides of it, appropriate targets were selected and AQP-9 was knocked out. The knockout result  were detected by PCR and gene sequencing. After the F1 heterozygous mice were obtained, 10 wild-type mice (5 males and 5 females) were provided for breeding in order to obtain homozygous AQP-9-/- mice.    Results   After injection of 74 fertilized eggs and transplantation of 60 pieces, 6 F0 generation positive mice were obtained.After breeding and identification, the homozygous AQP-9-/- mice were finally obtained.    Conclusion  Homozygous AQP-9-/- mice with stable inheritance could be obtained by using the CRISPR/Cas9 system.  References | Related Articles | Metrics

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N-butanol combined with turpentine instead of xylene for paraffin embeddin YU Jing JIANG Si-yu TIAN Jun-jie REN Li-qun XU Lu-dan LI Bao-qun WU Xiao-guang 2022, 53 (1):  132-136.  doi: 10.16098/j.issn.0529-1356.2022.01.019 Abstract ( )   PDF (25199KB) ( )   Objective  To look for a tissue paraffin embedding method  that can replace xylene transparent agent.    Methods  The mixture of N-butanol and turpentine was used to replace the role of anhydrous ethanol and xylene in the process of conventional paraffin embedding. The brain, kidney, stomach, liver and duodenum of multiple cerebral infarction model rats were embedded in paraffin. Finally, the new dehydration procedure was evaluated according to the effect of paraffin section, HE staining and immunohistochemical result .    Results  The mixture of N-butanol and turpentine could replace not only the dehydration effect of anhydrous ethanol but also the transparency effect of xylene in the conventional paraffin embedding process. The tissue sections treated with the mixture of N-butanol and turpentine were smooth, and the tissue did not become brittle or hard; After HE staining, the nucleus and cytoplasm of the new dehydrated tissue were distinct, and the chromaticity, color and transparency of the tissue were not different from those of the conventional dehydration procedure; Immunohistochemical staining was performed on different tissues of rats, and the comparison result  were no different from conventional embedded tissue immunohistochemical staining.    Conclusion  N-butanol combined with turpentine for tissue dehydration can not only avoid the toxic effect of xylene on human, but also reduce the tissue damage caused by excessive dehydration of anhydrous alcohol. References | Related Articles | Metrics

Cell and Molecules Biology

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Effects of CD73 on proliferation and function of rat cardiomyocytes in vitro WANG Yan ZHAO Yu-jie LI Zhi-ying WANG Qian GUO Zhi-kun 2022, 53 (1):  28-34.  doi: 10.16098/j.issn.0529-1356.2022.01.004 Abstract ( )   PDF (7856KB) ( )   Objective  To investigate the effect of CD73 on the proliferation and function of rat cardiac myocytes (CMs) in vitro.    Methods  The primary rat CMs and H9C2 cells were cultured in vitro, and lentivirus CD73 overexpression vector and empty vector group were constructed and transfected into CMs and H9C2 respectively. Experimental groups: CD73 overexpression group (OE group) including CMs-OE group and CMs-NC group, and negative control group (NC group) including H9C2-OE grop and H9C2-NC group, 5 in each group. After transfection, the expression of CD73 gene was detected by Real-time PCR method , and the proliferation ability was detected by MTT method , and the proliferation curve, doubling time and pulsatility of CMs were detected by non-destructive cardiomyocyte function analyzer.    Results  After 72 hours of lentivirus transfection, CD73 mRNA level in the over expression group was significantly higher than that in the control group (P<0.05); the proliferation ability of CMs and H9C2 at 3-4 days after transfection in OE group was larger than that in NC group (P<0.05); the proliferation curve of CMs and H9C2 cells in OE group was higher than that in NC group, and the doubling time of CMs and H9C2 in OE group was lower than that in NC group 72 hours after CMs transfection, the beating rate of OE group was higher than that of NC group, the expression of T-box 5(TBX5) and the secretion of vasuular endothelial growth factor VEGF) in OE group were higher than those in NC group, while the secretion of hypoxia iducible factor-1α(HIF-1α) and tumor necrosis factor-α(TNF-α) was lower than that of NC group.    Conclusion  CD73 can promote the proliferation of CMs and H9C2, promote CMs pulsation, TBX5 expression and VEGF secretion, and inhibit the secretion of HIF-α and TNF-α.  References | Related Articles | Metrics

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Hypoxic pretreatment enhancing the angiogenesis of aged human bone marrow mesenchymal stem cells through hypoxia inducible factor-1α SONG Hui-fang TAN Jia-yin LIU Yang ZHANG Wei-guo GUO Rui 2022, 53 (1):  35-41.  doi: 10.16098/j.issn.0529-1356.2022.01.005 Abstract ( )   PDF (9515KB) ( )   Objective  To investigate the effect of hypoxic pretreatment on the angiogenesis of aged human bone marrow mesenchymal stem cells (hBMSCs),  so to provide experimental support for more effective autologous stem cell transplantation therapy in aged patients with ischemic myocardial injury.    Methods  The aged hBMSCs were cultured in a hypoxic incubator, and then the cell morphology  was observed under inverted phase-contrast microscope, the surface markers were detected by flow cytometry, and the differentiation potential was detected by osteogenic and adipogenic differentiation. Subsequently, the conditioned medium(CM) of young hBMSCs under normoxic culture (norCM), the conditioned media of aged hBMSCs under normoxic and hypoxic culture(hypoCM) were collected respectively. They were named as norCM-young, norCM-old and hypoCM-old. The equal volume of medium, which was not treated with stem cells, was set as control group. Human umbilical vein endothelial cells (HUVECs) were treated with 4 conditioned media, the cell survival rate was detected by CCK-8 assay, and the tube formation experiment was used to detect the angiogenesis ability in vitro. The BCA method  was used to detect the total protein concentration of the conditioned medium of each group, and the Western blotting was used to detect  the expression of hypoxia inducible factor-1α (HIF-1α) in the cells and the conditioned media.    Results  There was not significant effect of hypoxic pretreatment on the morphology, surface markers and differentiation ability of aged hBMSCs (P>0.05. n≥3). Compared with the norCM-old group, hypoCM-old group significantly improved the survival of HUVECs under hypoxia-reoxygenation condition (P<0.05, n=5), and the tube formation ability of it (P<0.01, n=5). The total protein concentration of hypoCM-old group was significantly higher than that of norCM-old group (P<0.05, n=3). The expression of HIF-1α in hBMSCs of hypo-old group was significantly higher than that of nor-old group (P<0.05,n=3), while the content of HIF-1α in conditioned medium of hypoCM-old group was significantly higher than that in norCM-old group (P<0.01,n=3).    Conclusion  The aged hBMSCs pretreated with hypoxia can promote the survival and tube formation of HUVECs through paracrine in vitro, which is HIF-1α-related. References | Related Articles | Metrics

Anatomy

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Anatomic classification of the acromial angle and its clinical significance ZHOU Anping YAN Hang CHEN Yong-hong XIE Dong-bo ZHANG Yu-ke GUO Shi-tao WANG Jian-xue 2022, 53 (1):  92-95.  doi: 10.16098/j.issn.0529-1356.2022.01.012 Abstract ( )   PDF (1954KB) ( )   Objective  To study the acromial angle morphologic type and measurement analysis based on CT 3D reconstruction.    Methods  Totally 278 cases of adult CT three-dimensional reconstruction of the shoulder morphological data were collected, and the measurement data of the different types was analyzed, its statistical significance was clarified, and the morphological characteristics to division the type of acromial angle were summarized, its diagnosis and treatment under the acromion impingement of guiding significance were discussed.    Results  The acromial angle was divided into three types (C shaped acromial angle, L shaped acromial angle, and double angle shaped acromial angle). Among them, L type was the most, accounting for 48.56%, followed by C type, and double angle type was the least. In comparison of the breadth of the acromion and the length of the acromion, L type was significantly longer than C type (P<0.05). The thickness of acromion at a point of the double angle shaped acromial angle was greater than that of the other two type (P<0.05). In the comparison of ∠a, the double-angle type was greater than the C type (P<0.05), and the C type was greater than the L type (P<0.05).    Conclusion  There are significant differences in the classification and anatomical parameters of acromial angle, and the differences are statistically significant. It has certain guiding significance to the etiology and clinical diagnosis and treatment of subacromial impingement syndrome. References | Related Articles | Metrics

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Anatomy and imaging of abdominal aorta and its branches in Southwestern China population#br# #br# LIU Jian XU Li-jun ZHOU Yan-ping LI CHun-yan JIN Kai-yuan ZHOU Bo-jiang 2022, 53 (1):  96-102.  doi: 10.16098/j.issn.0529-1356.2022.01.013 Abstract ( )   PDF (3606KB) ( )   Objective  To investigate the normal reference values of abdominal aorta and its branches in Southwest China, so as to provide basis for clinical diagnosis and treatment of abdominal aortic aneurysm.    Methods  The length, diameter and angle of the abdominal aorta and its branches were measured in 20 cadavers. At the same time, 300 cases without obvious pathological changes in four urban populations were selected for abdominal CT angiography(CTA) to measure the length, diameter and angle of abdominal aorta and its branches.    Results Display rate of abdominal aorta was 100%,the average length of abdominal aorta was (12.96±0.81) cm, the diameter of abdominal aorta was (1.93±0.28) cm, (1.55±0.23) cm, (1.61±0.19) cm in renal artery plane (Ⅰ), inferior mesenteric artery plane (Ⅱ) and lower edge plane of lumbar vertebrae (Ⅲ), and the average value on image was (2.17±0.42) cm, (1.81±0.40) cm, (1.99±0.53) cm; Display rate of common iliac artery was 100%, the angle between left and right common iliac artery was (59.80±4.66) ° and the diameter of the end was (1.16±0.33) cm and (1.12±0.11) cm; Display rate of celiac trunk on cadaver was 100%, the average length was (1.61±0.27) cm, the average diameter on cadaver was (0.78±0.71) cm, the image was (0.88±0.31) cm; Display rate of renal artery was 100%, the average length of the left renal artery was (2.58±0.50) cm, that of the right was (4.26±0.65) cm, the initial average diameter of the left renal artery was (0.55±0.24) cm, and the right renal artery was (0.62±0.20) cm; Display rate of the superior mesenteric artery was (100%), that of the on cadaver was (4.56±0.29) cm, that of the image was (4.93±0.84) cm, and that of the angle between the abdominal aorta was (38.05±5.99)°; Display rate of the inferior mesenteric artery was (100%),the average length of the inferior mesenteric artery was (6.57±0.79) cm on the cadaver, the image was (6.70±0.76) cm, and that of the angle between the abdominal aorta was (73.79±9.62) ° and the initial diameter was (0.48±0.29) cm.    Conclusion  The normal reference values of abdominal aorta and its visceral branches in Southwest China population were measured. CTA has a special advantage in showing the lesions of abdominal aorta and its branches in lumen, extraluminal and vascular wall, which can provide anatomical data of patients before operation. References | Related Articles | Metrics

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Morphology and tissue structure of acromioclavicular joint ZHOU Ri XIE Xiao-xiao WANG Shuang REN Shi-you ZHANG Wen-tao 2022, 53 (1):  103-107.  doi: 10.16098/j.issn.0529-1356.2022.01.014 Abstract ( )   PDF (2643KB) ( )   Objective  To explore the morphology and the tissue structure of acromioclavicular joint.    Methods  Anatomical analysis was performed on 27 fresh adult cadavers and the morphology of the acromioclavicular joint was observed. The relevant bone structure and ligament parameters were measured, and the specimens were randomly divided into group A and group B. Group A reserved the acromioclavicular ligament and coracoclavicular ligament, and group B reserved only the acromioclavicular ligament. The difference in tension between the two groups was compared.    Results  The distance from the midpoint of the conical ligament to the distal end of the clavicle was (42.68±6.34) mm, the width of the end point was (16.97±4.28) mm, and the thickness of the center point was (5.39±0.34) mm; the distance from the midpoint of the trapezoidal ligament to the clavicle was (20.35 mm±4.18)mm, the width of the end point was (10.35±1.31)mm, the thickness of the center point was (5.19±0.342)mm; the average vertical distance from the base of the coracoid process to the surface of the clavicle was 30.75 mm, and the mean coracoclavicular gap was 12.02 mm; the length of the central axis of the conical ligament was (15.68±3.30)mm and the angle was (117.25±10.80)°, while the length of the central axis of the trapezoidal ligament was (9.67±2.25)mm, and the angle was (75.42±11.37)°. The distance between the start joint of the trapezoidal ligament and the trapezium was (8.96±3.00) mm, and the distance between the end points (13.09±3.50) mm. The average tensile force of group A was higher than that of group B ［(610.04±51.24) N vs (560.41±44.63) N, P<0.05］.    Conclusion  During distal clavicular resection, the resection of the distal clavicle shall be within 10-30 mm. The depth shall not exceed 42 mm when drilling under the coracoid process. The reconstruction of the coracoclavicular ligament during acromioclavicular joint dislocation has an anatomical and biomechanical basis. References | Related Articles | Metrics

Anthropology

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Comparison of sebum development between Daur adults in Inner Mongolia and Xinjiang SUI Jie LI Xin WEN You-feng ZHANG Wen-qian LI Qiang YU Ting 2022, 53 (1):  120-125.  doi: 10.16098/j.issn.0529-1356.2022.01.017 Abstract ( )   PDF (908KB) ( )   Objective  To discuss the main reasons for the difference of sedum development between Daur adults in Inner Mongolia and Xinjiang.    Methods  Nine circumference values and six skin fold thickness of 544 adults of Daur nationality (304 in Xinjiang and 240 in Inner Mongolia) were investigated by random sampling. Circumferences, skin fold thickness and obesity indexes of Daur nationality in Xinjiang and Inner Mongolia were analyzed.    Results  There were significant differences in 8 circumference without thigh circumference, 2 skin fold thickness such as triceps skinfold, subscapularis skinfold, 3 obesity indexes such as Verv index, body fat rate, body mass index(BMI). Correspondence analysis showed that subcutaneous fat development was stronger in northern regions than in southern regions.    Conclusion  The Daur nationality in Inner Mongolia has more fat and less muscle than that in Xinjiang. The problems of overweight and obesity in both regions are serious. It is suggested to pay attention to exercise and diet. Low temperature，lack of physical exercise, low temperature, low economic and medical standards and unhealthy eating habits can thicken subcutaneous fat. References | Related Articles | Metrics

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Correlation between body composition and serum lipid and uric acid in Maonan Guangxi population LUO Xiao-wei FENG Hang DENG Qiong-ying GONG Ji-chun GONG Jian-gu XU Lin 2022, 53 (1):  114-119.  doi: 10.16098/j.issn.0529-1356.2022.01.016 Abstract ( )   PDF (816KB) ( )   Objective  To investigate the relationship between body composition and serum lipids and uric acid among adults in Maonan, and to analyze the effect of body composition changes on blood lipid and uric acid.    MethodsTotally 584 Maonan adult volunteers in Maonan village of Maonan Autonomous County in Guangxi, the age from 20 to 80 were recruited. The height was measured by the personal height tester; the body composition was measured by the ANITAMC-180 instrument; and the blood lipids and blood uric acid were measured by the Hitachi 7600 instrument. The obtained data were statistically analyzed by SPSS 20.0.    Results  The age，height, weight, free fat mass, muscle mass, presumptive bone mass, body water, proptein，extracellular water, intracellular water, and waist-to-hip ratio were greater in Maonan men than in women (P<0.05). However, whereas male fat content, body fat rate, and subcutaneous fat content were smaller than those of female (P<0.01). The total prevalence of hyperuricemia and hyperlipidemia in Maonan nationality was 13.9% and 28.4%, respectively. The prevalence of hyperuricemia and hyperlipidemia in males was higher than in females. In males, the body mass, body mass index, free fat mass, fat mass, muscle mass, presumptive bone mass, protein, extracellular water, body fat rate, visceral fat content, subcutaneous fat content and waist-hip ratio of the hyperlipidemia group were higher than the normal group (P<0.05); and in females, the age, body mass index, fat mass, body fat rate, visceral fat content and waist-hip ratio of the hyperlipidemia group were higher than the normal group. In male, The body mass, free fat mass, presumptive bone mass, body water, extracellular water of the hyperuricemia group were higher than the normal group (P<0.05); In females, the age, body mass, body mass index, fat mass, extracellular water, body fat ratio, muscle mass, visceral fat content, subcutaneous fat content, and waist-hip ratio of the hyperuricemia group were higher than the normal group.    Conclusion  The detection rate of hyperlipidemia and hyperuricemia in males of Guangxi Maonan nationality is all higher than that in females. The body composition is significant differences between the normal adults and the patients with hyperlipidemia and hyperuricemia of Maonan nationality in Guangxi. References | Related Articles | Metrics

Histology,Embryology and Developmental Biology

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Expression of connexin 26 and 30 in cochlea in a rat model of type 2 diabetes LUAN Feng ZHANG Yan CHEN Xu-zhen WANG Na ZHANG Pu QU Yan 2022, 53 (1):  108-113.  doi: 10.16098/j.issn.0529-1356.2022.01.015 Abstract ( )   PDF (10821KB) ( )   Objective  To study the expression of connexin (Cx) 26 and Cx30 in the cochlea in rat model of type 2 diabetes, and their role in the hearing loss of type 2 diabetes.    Methods  Sixty wistar rats were randomly divided into a control group(n=20) and a experimental group(n=40). Rats in the experimental group received intraperitoneally injection of 10 mg/L streptozotocin to establish model of type 2 diabetes. Auditory brainstem response (ABR) was tested before and after molding at month 1, 2, 3, 4, 5. The morphology of cochlea was observed by HE staining, and the level and pattern of Cx26 and Cx30 expression within the cochlea were detected by immunofluorescence and Western blotting.    Results  In rats in the diabetes group, wave Ⅲ and Ⅴ latency, Ⅰ-Ⅲ and Ⅰ-Ⅴ interval of Click-ABRs (60 dBSPL) prolonged at month 1, 2, 3, 4, 5 after molding compared to the control (P<0.05). The number of cells was obvious reduced in the spiral ligament and ganglion of the experimental group(P<0.05). Immunofluorescence and Western blotting results showed decreased expression of Cx26 and Cx30 in the experimental group at 2, 3, 4, 5 month（P<0.05）, and the expression of the two prote decreased gradually with the time extension.    Conclusion  Expression of Cx26 and Cx30 is reduced at the same time as the occurrence of hearing impairment in rat cochlea with type 2 diabetes.  References | Related Articles | Metrics

Cancer Biology

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Effects of miR-515-5p on the proliferation and metastasis of ovarian cancer cell A2780 by regulating the expression of chondroitin sulfate proteoglycan 4 and its mechanism JIANG Hong HUANG Yan-li XING Hui WANG Li-ming GUO Hong 2022, 53 (1):  42-49.  doi: 10.16098/j.issn.0529-1356.2022.01.006 Abstract ( )   PDF (11747KB) ( )   Objective  To investigate the targeted regulation of miR-515-5p on chondroitin sulfate proteoglycan 4 (CSPG4) and its effect on the proliferation and metastasis of ovarian cancer cell A2780.    Methods  The target genes of miR-515-5p were predicted by bioinformatics tools. Real-time PCR and Western blotting were used to detect the expression of miR-515-5p and CSPG4 in 65 cases ovarian cancer tissues and adjacent tissues. Ovarian cancer cells A2780 were divided into miR-NC group, miR-515-5p group, si-NC group, si-CSPG4 group, miR-515-5p + pcDNA group, miR-515-5p + pcDNA-CSPG4 group. MTT assay was used to detect cell proliferation. Transell was applied to detect cell migration and invasion, and Western blotting was selected to detect cyclinD1, P21, matrix metalloproteinase(MMP)-2 and MMP-9 protein expression. The double luciferase reporter gene experiment and Western blotting confirmed the regulation effect of miR-515-5p on CSPG4 gene.    Results  Bioinformatics analysis showed that CSPG4 was one of the potential target genes of miR-515-5p. Compared with the adjacent tissues, the expression of miR-515-5p was lower in ovarian cancer tissues, and the expression of CSPG4 was higher (P<0.05). Compared with the miR-NC group, the expression of cyclinD1, MMP-2 and MMP-9 protein was lower in A2780 cells of miR-515-5p group, the expression of P21 protein was higher, the cell viability was lower, and the number of cell migration and invasion was lower (P<0.05). Compared with the si-NC group, the expression of cyclinD1, MMP-2 and MMP-9 proteins were lower in A2780 cells in si-CSPG4 group, P21 protein was higher, cell viability was lower, and the number of cell migration and invasion was lower (P<0.05). Compared with the miR-515-5p + pcDNA group, the expression of cyclinD1, MMP-2 and MMP-9 proteins was higher in A2780 cells of miR-515-5p + pcDNA-CSPG4 group, the expression of P21 protein was lower, and the cell viability was higher, the number of cell   igration and invasion was higher (P<0.05). MiR-515-5p targeted and negatively regulated CSPG4 expression.    Conclusion  MiR-515-5p could inhibit the proliferation and metastasis of ovarian cancer cell A2780 by targeting CSPG4.  References | Related Articles | Metrics

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Wiskot-Aldrich syndrome-like lower expressed in colon cancer tissues and inhibited the stemness of human colon cancer LI Hong-dan YANG Cheng 2022, 53 (1):  50-59.  doi: 10.16098/j.issn.0529-1356.2022.01.007 Abstract ( )   PDF (15745KB) ( )   Objective  To explore the expression of Wiskot-Aldrich syndrome-like（WASL）in colon cancer tissues and its function on colon cancer stemness.    Methods  The UALCAN database was used to analyze the expression of WASL in colon cancer tissues and its correlation among the gender, lymphnode stage and metastasis in colon cancer patients. Immunohistofluorescence and immunocyte fluorescence were used to verify the date from UALCAN. Western blotting was used to detect the expression and transfection efficiency of WASL in human normal colorectal mucosal cells (FHC cells) and colon cancer cells HCT-116, SW-480. Sphere formation assay and colony formation were used to detect the effect of WASL on the stemness of colon cancer. Hypoxia assay, serum deprivation, drug resistance assay were used to detect the stemness of colon cancer cells.   Results  WASL was lower expressed in colon cancer tissues (P<0.05). WASL was lower expressed in colon cancer tissues than peri-cancer tissues. WASL was obviously lower expressed in HCT-116 and SW-480 than FHC cells. WASL overexpression stable cells were constructed suscessfully in HCT-116 and SW-480.Real-time PCR showed WASL obviously down-regulated the expression of cancer stemcells(CSCs) related markers ［octamer binding transcription factor 4(OCT4), sex determining region Y-box 2(SOX2), Lin28, krüppel-like factor(KLF)］. Sphere formation assay and colony formation assay showed WASL could inhibited the sphere formation and colony formation. Hypoxia assay and serum deprivation showed WASL inhibited the resistance of cancer cells to hypoxia and serum deprivation. Drug resistance assay showed WASL inhibit the resistance of cancer cells to 5-fluorouracil(5-FU).    Conclusion  WASL is lower expressed in colon cancer tissues and cells, and is related with poor prognosis of colon cancer. Overexpression of WASL can inhibit the stemness of colon cancer cells. References | Related Articles | Metrics

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Expression and clinical significance of CC motif chemokine ligand 20 in esophageal carcinoma based on the Cancer Genome Atlas data mining QIN Bo WANG Zheng-yang YIN Fan-xiang YAN Dan ZHU Zhu 2022, 53 (1):  60-65.  doi: 10.16098/j.issn.0529-1356.2022.01.008 Abstract ( )   PDF (9503KB) ( )   Objective  To explore the expression and significance of CC mokif chemokine ligand 20(CCL20) gene in esophageal carcinoma and its influence on the progression and prognosis of the disease.    Methods  The expression data of esophageal cancer were obtained from the Cancer Genome Atlas (TCGA) database, and were collected from cBioPortal(www.cbioportal.org).The survival curve was analyzed by Kaplan-Meier method . Through the website (http://ualcan.path.uab.edu) the objective was to analyze the expression of CCL20 gene in esophageal cancer tissues, the relationship between CCL20 and obesity and the methylation level of CCL20 in esophageal cancer. cBioPortal was used to analyze the copy number and mRNA expression of CCL20 gene. Finally, the expression differences of CCL20 in esophageal cancer tissues and adjacent tissues were analyzed by mRNA and protein levels.    Results  CCL20 was highly expressed in esophageal cancer, especially adenocarcinoma. The survival rate of patients with high expression of CCL20 was reduced. The expression of CCL20 was directly proportional to the degree of obesity in patients with esophageal cancer and inversely proportional to the degree of methylation. The expression of CCL20 in esophageal cancer tissues increased significantly.    Conclusion  CCL20 gene expression level is associated with poor prognosis of esophageal cancer patients.  References | Related Articles | Metrics

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Effect of tumor necrosis factorα overexpress on the biological properties dental pulp stem cells in microoxidative stress LI Wei YANG Gan-un PENG Jian-an HU Hong-mei ZHOU Xing-chen 2022, 53 (1):  66-74.  doi: 10.16098/j.issn.0529-1356.2022.01.009 Abstract ( )   PDF (6681KB) ( )   Objective  To investigate the effect of tumor necrosis factor-α(TNF-α) on biological properties of dental pulp stem cells (DPSCs) in low-oxidation stress.    Methods  The experimental groups were as follows: normox, normox control plasmid, normox-TNF-α, 3% O2, control plasmid-3% O2、TNF-α-3% O2.DPSCs were resuscitated and cultured firstly, then lentivirus TNF-α transfected cells were cultured in a CO2 incubator at 37 ℃. The expression of TNF-α  was detected by ELISA in each group, and the expression of TNF-α under oxidative stress was detected by RT-PCR,the expressions of alkaline phosphatase(ALP) and dentin sialophosphoprotein(DSPP) were detected by Real-time PCR and Western blotting,the proliferation of DPSCs cells transfected with TNF-α was detected by flow cytometry and MTT.    Results  After lentivirus-TNF-α transfected cells for 10 hours, the green fluorescence signal could be observed under inverted fluorescence microscope, TNF-α overexpression could enhance the proliferation of DSPP in hypoxic environment,and hypoxia microenvironment can greatly increase the expression of TNF-α, Western blotting and Real-time PCR showed that there were significant differences in the expression of ALP and DSPP in DPSCs（P<0.05）.    Conclusion  TNF-α overexpression can improve the proliferation efficiency and differentiation efficiency of DPSCs in hypoxic microenvironment.  References | Related Articles | Metrics

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Effect of short hairpin RNA lentivirus-mediated silencing of pleomorphic adenoma gene like-2 on proliferation, migration and invasion of hepatocellular carcinoma cells ZHAO Ke-chang WANG Wei YANG Jin-yu FENG Peng-cai WU Shi-le 2022, 53 (1):  75-81.  doi: 10.16098/j.issn.0529-1356.2022.01.010 Abstract ( )   PDF (2158KB) ( )   Objective  To investigate the effect of short hairpin RNA(shRNA)-mediated pleomorphic adenoma gene like-2 (PALAG2) silencing on the malignant behavior of hepatocellular carcinoma cells and its mechanism.    Methods   Real-time PCR and Western blotting were used to detect the expression level of PLAGL2 in liver cancer tissues and adjacent tissues. Hepatoma cells MHCC97-L were cultured in vitro, the lentiviral vector plasmid PLAGL2-shRNA and control NC-shRNA were constructed, transfected into MHCC97-L cells, and stable transfected strains were selected with puromycin. CCK-8 and Transwell chamber assay detected the proliferation activity and the number of migration and invasion of MHCC97-L cells after silencing PLAGL2. Western blotting was used to detect the expression of p-PI3K and p-Akt proteins. The PI3K/Akt signaling pathway activator was used to treat MHCC97-L cells to detect cell proliferation, migration and invasion.    Results  The expression of PLAGL2 was significantly increased in liver cancer tissue (P＜0.05). Transfection of 9 strains of MHCC97-L cells with PLAGL2-shRNA could significantly reduce the expression level of PLAGL2, and the ability of proliferation, migration, and invasion of MHCC97-L cells was also weakened (P<0.05), and the expression levels of p-PI3K, and p-Akt were inhibited (P<0.05), PI3K/Akt activator could obviously reverse the above phenomenon.    Conclusion  shRNA lentiviral vector pathway can effectively silence the expression of PLAGL2 gene in hepatocarcinoma cells. Silencing of PLAGL2 can significantly inhibit the malignant behavior of proliferation, migration and invasion of hepatocarcinoma cells, and its mechanism may be related to the inhibition of PI3K/Akt signaling pathway activation.  References | Related Articles | Metrics

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Improvement of astaxanthin on learning and memory  ability of vascular dementia mice ZHANG Ming MA Wei-cheng CAO Wei-juan ZOU Jia-ling ZHU Ning-wei 2022, 53 (1):  5-10.  doi: 10.16098/j.issn.0529-1356.2022.01.001 Abstract ( )   PDF (1845KB) ( )   Objective  To study the  effect and mechanism of astaxanthin on learning and memory ability of vascular dementia (VaD) mice.    Methods  The mice were used to establish VaD model by occlusion of bilateral common carotid artery. The mice were randomly divided into sham group, model group, astaxanthin low-dose group and astaxanthin high-dose group and then given corresponding forms of drug treatments. Morris water maze was used to investigate the learning, memory and space exploration abilities of mice in each group. At the same time, the pathological morphology of brain neurons, the expression of amyloid beta-peptides 42(Aβ42) protein in brain and the content of interleukin (IL)-4 and IL-6 were respectively examined by Nissl staining, immunohistochemical staining, Western blotting and ELISA.    Results  The average escape latency of mice in the model group on the 4th and 5th days was significantly higher than sham-operated group (P<0.05), and the number of plateau crossings and the duration of stay decreased significantly (P<0.05). At the same time, Nissl staining showed that the neurons in the hippocampus of the model group were shrinking and disordered. The number of neurons in the model group was significantly reduced compared with the sham-operated group. The average escape latency of mice treated with different doses of astaxanthin on the 4th and 5th day was significantly lower than the model group (P<0.05).The number and duration of platform crossing were significantly longer than model group (P<0.05). The number of neurons in the intervention group was obviously higher than model group, and the effect was dose-dependent. In addition, immunohistochemical staining, Western blotting analysis and ELISA detection showed that the levels of Aβ42 protein and IL-6 in the astaxanthin intervention group decreased significantly compared with the model group (P<0.05), while the levels of IL-4 increased significantly (P<0.05).   Conclusion  Astaxanthin can reduce the expression of neurotoxic protein Aβ42 protein and inhibit the inflammatory response in the brain, which can protect neurons and improve the learning and memory ability of mice.  References | Related Articles | Metrics

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18α-glycyrrhetinic acid promoting neural stem cells proliferation  through antioxidant in the subventricular zone of adult mice LIU Xue-qin NIU Xiao-jie WANG Bi-hui WANG Ting-yu YANG Jing LU Li 2022, 53 (1):  11-18.  doi: 10.16098/j.issn.0529-1356.2022.01.002 Abstract ( )   PDF (7430KB) ( )   Objective  To explore the effect of 18α-glycyrrhetinic acid (18α-GA) on the proliferation of adult mice neural stem cells (NSCs) and its underlying mechanism.    Methods  One hundred 6-month BALB/c mice were randomly divided into DMSO control group and 18α-GA group (mice were intraperitoneally injected with 40 mg / kg 18α-GA every day for 2 months). The proliferation capability, oxidative status and  nuclear factor E2-related factor 2 (Nrf2) level of NSCs in the adult mice subventricular zone (SVZ) were measured through both in vivoand  in vitro experiments, including Ki-67 staining, neurosphere formation assay, BrdU incorporation, CCK-8 assay, reactive oxygen species (ROS) detection, superoxide dismutase 1 (SOD1) determination, Real-time PCR and Western blotting.    Results  Elevated Ki-67 positive cells were observed in SVZ of mice with 18α-GA application. Meanwhile, ROS level attenuated but SOD1 mRNA and protein level increased significantly in the SVZ of 18α-GA group mice, the latter of which were (3.17±0.073) and (2.12±0.02) times respectively than that of the control group (P<0.05 and P<0.001). Likewise, the similar changes were exhibited in vitro data. NSCs of 18α-GA group mice displayed higher proliferation potency confirmed by accelerated neurosphere formation and increased neurosphere number (P<0.001), as well as higher BrdU positive ratio (P<0.01) and NSCs vitality (P<0.001). NSCs of mice with 18α-GA injection exhibited decreased ROS level by 18.91%±4.33% (P<0.05) and enhanced SOD1 level, compared with those in NSCs of DMSO group mice. Furtherly, the Nrf2 expression in SVZ and NSCs of 18α-GA group mice was higher than that of the control group.  Conclution  18α-GA administration plays a vital role in the maintainence and amelioration of adult mice NSCs proliferation through activating SOD1 and diminishing ROS aggregations.  References | Related Articles | Metrics

Cancer Biology

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Relationship between miR-26a and metadherin and the verification of tumor-bearing experiment in nude mice YANG Chen-chen FAN Yu-qin 2022, 53 (1):  82-91.  doi: 10.16098/j.issn.0529-1356.2022.01.011 Abstract ( )   PDF (6739KB) ( )   Objective  To explore the relationship between miR-26a and metadherin (MTDH), and to verify the relationship between miR-26a and MTDH in vivo in nude mice.    Methods  Immunohistochemical SP staining method  was used to detect the expression of MTDH and in situ hybridization was used to detect the expression of miR-26a in 86 cases of esophageal cancer, and the correlation between the expressions was analyzed. The bioinformatics prediction Targetscan Human 7.2 software could predicte the binding fragment of MTDH on the miR-26a sequence, and the luciferase report experiment was used to verify the targeted regulatory relationship between MTDH and miR-26a. Nude mice were injected esophageal cancer cell lines subcutaneously which were lentiviral interferenced and overexpressed miR-26a to observe the formation of tumors. The tumors from nude mice were made into paraffin. and each was detected. The expression of MTDH in miR-26a in the tumor groups was detected by immunohistochemical staining and in situ hybridization and the relationship was analyzed.    Results  The expression of miR-26a in esophageal cancer tissues was significantly lower than that in paired normal esophageal tissues, and the expression of MTDH in esophageal cancer tissues was significantly higher than that in paired normal esophageal tissues. The expression of miR-26a was related to the patient’s pathological grade (P<0.05), N stage (P<0.05), and tumor volume (P<0.01). The expression of MTDH in esophageal cancer was related to the N stage (P<0.05) and the degree of differentiation (P<0.01). Targetscan Human7.2 bioinformatics software predicted that the MTDH gene contained a target sequence of hsa-miR-26a.The luciferase reporter gene experiment also verified the targeted regulation relationship between miR-26a and MTDH. The expression of miR-26a was the highest in KYSE-450 cells and the lowest in Eca109 cells. The mRNA expression of MTDH in the lv-miR-26a group was significantly lower than that in the lv-NC group, and the mRNA expression in the lv-miR-26a-inhibitor group was significantly  higher than that in the lv-NC group. After tumor formation, miR-26a expression increased and MTDH expression decreased in miR-26a group. After tumor formation, the expression of miR-26a decreased and the expression of MTDH increased in miR-26a inhibitor group.    Conclusion  MiR-26a can inhibit the expression of MTDH in esophageal cancer cells. Both in vitro and in vivo experiments can verify the targeted regulatory relationship between miR-26a and MTDH. MiR-26a may play a role in the occurrence and development of esophageal cancer through the MTDH.  References | Related Articles | Metrics

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Protective effect and mechanism of ginsenoside Rb1 on repairing sciatic nerve injury in mice YAO Yuan DU Jing-yi ZHOU Wen-juan 2022, 53 (1):  19-27.  doi: 10.16098/j.issn.0529-1356.2022.01.003 Abstract ( )   PDF (11037KB) ( )   Objective  To explore the effect and mechanism of ginsenoside Rb1 on the repair of sciatic nerve injury (SNI) in mice.    Methods  Seventy-eight adult male Kunming mice were randomly divided into sham group (26), SNI group (26), SNI+Rb1 group (26). The SNI+Rb1 group was given 10 mg/kg ginsenoside Rb1 (i.p.), and the SNI group and the sham group were given the same volume of normal saline. The injury method  was established by squeezing the sciatic nerve. Sciatic functional index (SFI) was used to evaluate sciatic nerve function. Growth associated protein 43(GAP43) immunofluorescent staining was used to detect neural regeneration and repair on day 14, and the structure changes of the myelin sheath of the injured segment were observed under transmission electron microscope. Ki67 and S100β were used to detect the proliferation and migration ability of Schwann cells, and Real-time PCR was used to detect the mRNA expression levels after crush on day 3 and day 7.   Results SFI of SNI+Rb1 group was higher than SNI group. The HE result  showed that the sciatic nerve was uniform in the SNI+Rb1 group. The result  of immunofluorescent staining displayed that Rb1 enhanced GAP43+ axon elongation, and the expression of brain derived neurotrophic factor (BDNF), nerve growth factor (NGF) was elevated. Transmission electron microscopy study suggested that, compared with the model group, the myelin structure and thickness of the SNI+Rb1 group were improved. Further testing revealed that Ki67+ cell proliferation and S100β+ Schwann cell migration in the vicinity of the injured segment increased after Rb1 was administered. The result  of Real-time PCR showed that the mRNA expression levels of tumor necrosis factor (TNF)-α and interleukin (IL)-1β in the SNI+Rb1 group decreased rapidly, and the expression levels of transcription factors related to Schwann cell activation increased (P<0.05).    Conclusion  Rb1 10 mg/kg can reduce the expression level of inflammatory factors, and increase the expression level of BDNF and NGF, and promote Schwann cell activation, which is beneficial for the nerve regeneration after sciatic nerve injury in mice. References | Related Articles | Metrics