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05 March 2021, Volume 41 Issue 3
Previous Issue   
Anti-tumor drug FTY720 regulates p53-dependent transcription
2021, 41(3):  311-317. 
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Objective To explore whether anti-tumor drug FTY720 participates in the regulation of p53-mediated gene transcription and evaluate the potential roles of FTY720-p53 axis in the modulation of cellular biological processes. Methods In human osteosarcoma cell line (U2OS) cells that express the wildtype p53, the CRISPR/Cas9 technique was used to generate a p53-knockout (p53-KO) cell strain. Control cells or p53-KO cells were treated with or without FTY720 and the transcriptional profiles were then assessed by RNA-sequencing (RNA-seq) analysis. The differentially expressed genes as well as their potential functions were annotated by using bioinformatic analysis. Results The U2OS p53-KO cell strain was successfully established. We identified 220 p53-dependent downstream genes in response to FTY720 treatment. These genes were functionally enriched in the regulation of multiple biological processes including negative regulation of platelet activation, cellular response to chemokine, positive regulation of Rho protein signal transduction, multicellular organismal homeostasis, mucin type O-glycan biosynthesis, ECM-receptor interaction, viral protein interaction with cytokine and cytokine receptor. Conclusions p53 activation is involved in FTY720-mediated tumor suppressive functions.
Construction of B16 cell strain expressing CSDE1-EGFP using CRISPR-PITCH system
2021, 41(3):  318-324. 
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Objective To construct the melanoma cell line expressing endotopic CSDE1-EGFP by using the new CRISPR-PITCH system, then to select the cell line of low expression CSDE1 and high expression CSDE1. Methods The EGFP coding sequence was inserted into the last exon of the CSDE1 gene using a micro-homology-mediated end joining method ; Use flow cytometry sorting, PCR, immunofluorescence and live cell imaging to detect whether CSDE1-EGFP is inserted successfully, and whether the fluorescence intensity of EGFP is consistent with the expression level of CSDE1; Use transwell experiment and mouse tumor-bearing experiment were used to observe the functional differences between the B16 cell lines with low and high expression of CSDE1. Results The B16 cell line of low expression CSDE1 and high expression CSDE1 were selected, and the strength of EGFP expression was detected to be consistent with CSDE1 expression. It was observed that melanoma cells with different expression of CSDE1 protein have different functions. Conclusion The new CRISPR-PITCH system provides a fast and effective intuitive method for studying the dynamic positioning of CSDE1 in other tumor cells and the functional differences due to different expressions.
Establishing of a CRISPR/Cas9 system targeting at mouse Krt14 that stimulates pathogenic site mutations of human
2021, 41(3):  325-332. 
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Objective To establish effective CRISPR/Cas9 system targeting mouse Krt14 that can mimic the strong pathogenic site mutation of human KRT14. Methods In this study, databases such as OMIM, ExAC and ClinVar were searched to screen the strong pathogenic sites of human KRT14, the causative gene of human epidermolysis bullosa simplex. Then, the human and mouse KRT14 protein sequence alignment analysis was used to locate the strong pathogenic sites on the mouse genome. Based on the gene targeting principle of the CRISPR/Cas9 system, we designed 4 sgRNAs targeting the mouse Krt14 and constructed 4 expression plasmids. Then, sgRNA expression plasmids and Cas9 expression plasmid were co-transfected into mouse 3T3 cells. PCR product sequencing, TA clone sequencing of the positive cells screened by drugs were performed to verify the target efficiency of 4 sgRNAs. Results According to the screening results, the p.Arg125 site of human KRT14 protein is determined to be a strong pathogenic site, which corresponds to the p.Arg131 site of mouse KRT14 protein. According to the DNA sequence of the mouse p.Arg131 site, 4 sgRNA expression plasmids targeting this site was successfully constructed. Sequencing of PCR amplification products of the target site of drug screen positive cells showed that 4 sites were all mutated. The results of TA cloning sequencing showed that the mutation efficiency of the four sites was respectively 70%, 90%, 65% and 100%. Conclusions The above results indicate that we have successfully constructed efficient CRISPR/Cas9 system targeting the mouse Krt14 based on the highly pathogenic sites of the human KRT14 screened. This study lays the foundation for the in-depth study of Krt14 gene function and the establishment of Krt14 gene editing mouse model to explore the mechanism and treatment of disease.
miR-29b inhibits LPS-induced apoptosis in human bronchial epithelial cell lines 16HBE
2021, 41(3):  333-339. 
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Objective To study the effect of miR-29b on lipopolysaccharides(LPS)-induced human bronchial epithelial cell 16HBE apoptosis. Methods 16HBE was divided into Control group, LPS group (LPS treatment), transfection mimics control group, transfection miR-29b mimics group, MTT assay for proliferation, flow cytometry for apoptosis, and Western blot for cleaved cysteinyl aspartate specific proteinase 3(c-caspase-3), cleaved cysteinyl aspartate specific proteinase 12(c-caspase-12), Glucose tegulated protein 78(GRP78) and CCAAT/enhancer-binding protein C/EBP(CHOP)protein expression. Human bronchial epithelial cells 16HBE were co-transfected with miR-29b mimics and pcDNA-CHOP, the proliferation and apoptosis were also determined using the above method. Results Compared with the Control group, the cell proliferation activity and apoptosis rate of the LPS group decreased, and the protein expression levels of c-caspase-3, c-caspase-12, GRP78, and CHOP increased in the cells. Compared with the LPS+NC group, the LPS+miR-29b group increased cell proliferation activity, decreased apoptosis rate, and decreased c-caspase-3, c-caspase-12, GRP78, and CHOP protein expression levels. pcDNA-CHOP could reverse the effects of miR-29b mimics on the proliferation and apoptosis of bronchial epithelial cells under LPS conditions. Conclusions miR-29b inhibits 16HBE apoptosis induced by LPS, the mechanism was related to the inhibition of endoplasmic reticulum stress.
Macrophage polarization imbalance and metabolic changes in autoimmune thyroiditis mice
2021, 41(3):  340-345. 
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Objective To investigate the significance of macrophage polarization imbalance and metabolic pathway changes in the occurrence and development of autoimmune thyroiditis (AIT).Methods 4 weeks old NOD.H-2h4 female rats were randomly divided into control group and AIT group(0.05% sodium iodide water). Hematoxylin-eosin staining was used to observe the lymphocytic infiltration of thyroid tissue. TgAb were measured by Enzyme Linked Immunosorbent Assay (ELISA). The ratio of M1 type and M2 type macrophages in thyroid tissue was analyzed by immunofluorescence. Seahorse was used to analyze the energy metabolism level of cells, and the extracellular acidification rate (ECAR) and O2 consumption rate (OCR) were measured. Results The thyroid tissue of mice in AIT group had obvious lymphocyte infiltration, and the incidence of AIT is significantly higher than the control group(P<0.05). Compared with the control group, the ratio of M1 and M2 macrophages in the thyroid tissue of mice in the AIT group increased, while the proportion of M2 type macrophages decreased (P<0.05); the ECAR and OCR values of mice in the high iodine drinking water group were significantly higher than those of the control group (P<0.05).Conclusion The change of macrophage metabolism and the polarization imbalance affected the occurrence and development of AIT.
Analyses of factors influencing myocardial fibrosis in non-ischemic heart failure patients
2021, 41(3):  346-351. 
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Objective To investigate the potential factors influencing myocardial fibrosis (MF) in nonischemic heart failure patients with an emphasis on the role of right heart catheter (RHC) parameters. Methods We assessed the clinical data from 33 nonischemic heart failure patients admitted to China-Japan Friendship Hospital who receiving endomyocardial biopsy (EMB) and right heart catheter (RHC) examination. All the patients were divided into two groups: mild MF (n=17) and severe MF (n=16), by means of K-means cluster analysis. MF was quantitatively evaluated by MASSON staining. Potential factors influencing MF were analyzed statistically. Results Compared with that in mild MF group, patients in severe MF group had aggravated cardiac function (cardiac index, 2.15±0.76 vs. 2.94±1.10,P<0.05), accompanied by higher systolic right ventricle pressure (RVP), systolic pulmonary arterial pressure (PAP), pulmonary arterial wedge pressure (PAWP) and total pulmonary resistance (TPR) (P<0.05). Hypertension and TPR were independently correlated with degree of MF in multiple linear regression analysis (Adjusted R2=0.454, P<0.001). Conclusions Hypertension and TPR, reflecting afterload of left and right ventricle, are found to be independently correlated with degree of MF in nonischemic heart failure patients. Cardiologists should pay more attention to high-risk patients in order to retard MF progression through timely intervention.
Whole-exome sequencing from a case of pediatric aldosterone- and cortisol-coproducing adrenal adenoma
2021, 41(3):  352-357. 
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Objective To identify the genetic mutations of aldosterone- and cortisol-coproducing adrenal adenoma from a young boy by whole-exome sequencing. Methods The clinical data of a boy with aldosterone- and cortisol-coproducing adrenal adenoma were collected.Whole-exome sequencing was performed with DNA extracted from the blood and tumor tissue to identify germline and somatic mutations. Results A 13-year-old boy with hypertension and hypokalemia was diagnosed as primary aldosteronism complicated with subclinical Cushing's syndrome according to clinical manifestations and adrenal hormone testing. CT scan found a 4.5cm ×3.6 cm mass in right adrenal gland. The mass was removed and pathologically diagnosed as adrenocortical adenoma. A somatic CTNNB1 c.133T>C (p.S45P) mutation was detected in the adenoma tissue. Somatic mutations, such as KCNJ5, ATP1A1, ATP2B3 and CACNA1D were not detected. Conclusions Somatic CTNNB1 mutation is probably the main cause of this rare case with aldosterone- and cortisol-coproducing adrenal adenoma. But the profound mechanism needs further study.
miR-205 inhibits cell proliferation of papillary thyroid carcinoma by regulating Wnt-5a
2021, 41(3):  358-363. 
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Objective To investigate whether miR-205 can regulate Wingless-Type MMTV Integration Site Family Member 5a (Wnt-5a) to inhibit the papillary thyroid carcinoma (PTC) cell proliferation. Methods RT-qPCR was used to detect the expression of miR-205 and Wnt-5a in PTC tissues and adjacent normal tissues .Further correlation analysis between miR-205 and Wnt-5a was carried out. The low expression of miR-205 related to the stage of PTC TMN and lymph node metastasis was also discussed. Meanwhile, RT-qPCR was also used to detect NTHY-OR-3-1 and PTC cell lines (K1 and BCPAP). In addition, the proliferation of PTC cells was detected by CCK-8 after over expression and inhibition of miR-205. Xenograft tumor formation test was used to observe whether inhibition of miR-205 accelerate the growth of tumor. The online target gene prediction software and dual luciferase reporter gene test were used to predict and verify whether Wnt-5a is the target gene of miR-205. Results The expression of miR-205 was significantly lower than that of surrounding normal tissues, and the expression of Wnt5a was significantly higher. TNM stage and lymph node metastasis severity of PTC patients were significantly higher in patients with low miR-205 expression than in patients with high miR-205 expression. Low expression of miR-205 has poor survival rate. Expression of miR-205 in NTHY-OR-3-1 was significantly lower than that of K1 and BCPAP. miR-205 inhibited the proliferation of PTC cells. Over expression of miR-205 also inhibited the growth of PTC. It was confirmed that Wnt5a was the target gene of miR-205 by luciferase reporter assay. Conclusions miR-205 plays an anti-cancer role in PTC, which may provide a new therapeutic target for PTC.
Over-expression of c-SKI inhibits isoprenaline-induced myocardial fibrosis in mice
2021, 41(3):  364-369. 
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Objective To investigate the effect of c-SKI on isoproterenol-induced cardiac fibrosis in Mice. Methods Mice were divided into 4 groups: the control group,the Model group,the c-SKI gene infection group (LV-SKI group) and the normal control virus infection group (LV-NC group), with 10 ones in each.In the model group, isoproterenol (ISO) was injected subcutaneously for 30d, with the first dose of 5 mg/kg followed by 2.5 mg/kg. In the LV-SKI group and the LV-NC group, c-SKI lentivirus and control virus were injected into the tail vein on days 18, 21, 24, 27 and 30 at a dose of 100μL, titer was 1×108 TU.HE staining and Masson staining were used to observe the pathological changes of myocardial tissue. ELISA was used to detect type I and type III collagen in myocardial tissue.Western blot was used to detect the expression of c-SKI,interstitial cell marker vimentin, α SMA,endothelial cell marker CD31 and the expression levels of transcription factors Snail, Twist and Slug. Results Compared with the control group, the expression of c-ski in the model group was time-dependent (P < 0.01). In the model group, the myocardial cells were disordered, the interstitium was significantly increased, and the contents of type I and type III collagen were increased (P < 0.01). Expressions of vimentin, α-SMA, Snail, Twist and Slug were all upregulated, while CD31 protein expression was down-regulated (P < 0.05). After over-expressing c-SKI, compared with the LV-NC group, the myocardial structure of the mice tended to be neat, the interstitial collagen fibers decreased, and the type I and III collagen in the myocardial tissue decreased (P < 0.01). The expression of vimentin and belt-sma was down-regulated (P < 0.05), and the expression of CD31 was up-regulated (P < 0.05). Meanwhile, the expression of transcription factors Snail, Twist and Slug was also down-regulated (P < 0.01). Conclusions c-SKI expression is down-regulated during myocardial fibrosis in mice, and overexpression of c-SKI can improve isoproterenol-induced myocardial fibrosis by inhibiting endothelial-mesenchymal transition.
Effects of administration of omeprazole on the esophageal microbiota composition in rat model with reflux esophagitis
2021, 41(3):  370-375. 
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Objective To investigate the effect of proton pump inhibitor (PPI) omeprazole treatment on the composition of esophageal mucosal microbiota in rats with reflux esophagitis. Methods The rat model of reflux esophagitis was established by the method of pyloric partial ligation and complete anterior gastric ligation. Rats were randomly divided into sham operation control group, reflux esophagitis control group and PPI treatment group (omeprazole). One week after the operation, distilled water / omeprazole tablets were intragastric administration once a day, and two pieces of lower esophageal mucosa were obtained 2 weeks later, one for bacteria 16S rDNA gene sequencing, and another for pathological examination. Results Six surviving rats in each group were selected. The abundance values of sham operation control, reflux esophagitis control and PPI treatment groups at the genus level were as follows: Peptostreptococcus 2.5%±0.5%, 1.7%±0.7% and 1.9%±0.5%; Helicobacter 1.5%±0.1%, 1.0%±0.5% and 1.2%±0.3%;Turicibacter 1.0%±0.2%, 0.5%±0.2% and 1.0%±0.4% (P < 0.05). The abundances of Peptostreptococcus, Helicobacter, and Turicibacter in the reflux esophagitis control group were significantly reduced than those in the sham operation control group. The pathological grades of reflux esophagitis (mild / moderate / severe) in reflux esophagitis control and PPI treatment groups were 0/4/2 and 5/1/0 (P < 0.05). Conclusion There were moderate changes in the composition of esophageal mucosal microbiota in rats with reflux esophagitis,the proportion of beneficial bacteria reduced. After the treatment of PPI omeprazole, the esophageal mucosal injury was alleviated, the proportion of beneficial bacteria increased, and the composition of esophageal mucosal microbiota returned to normal.
Salvianolic acid B alleviates LPS- induced acute lung injury in mice
2021, 41(3):  376-381. 
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Objective To study whether Salvianolic acid B can reduce LPS induced acute lung injury .Methods 48 mice were randomly divided into control group, model group (intratracheal instillation of LPS), Salvianolic acid B low / medium / high dose intervention group, and sulforaphane positive control group. The lung wet / dry weight ratio (W/D) was detected; the protein concentration of alveolar lavage fluid (BALF) was detected; lung tissue pathological damage was evaluated by HE staining; TNF-α,IL-1βand IL-6 in BALF were detected by ELISA .Western blot was used to detect the protein expression of Nrf2, NQO1, HO1, Keap1,NF-κB and p -NF-κB in lung tissue. Results compared with the control group, pathological changes were observed in the lung tissue of the model group,W/D , protein concentration in BALF and inflammatory cytokine content were significantly increased (P < 0.05). Compared with the model group, Salvianolic acid B intervention group had significant changes (P < 0.05), alleviated the oxidative stress induced by LPS, and increased the expression of Keap1 (P < 0.01), increased the expression of Nrf2, NQO1 and HO1 (P < 0.01), inhibited the activation of NF-κB and p-NF-κB induced by LPS. Conclusion Salvianolic acid B can alleviate LPS induced acute lung injury in mice.
Sophoridine alleviates hypoxia-induced injury of rat myocardial cell line H9c2
2021, 41(3):  382-387. 
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Objective To investigate the inhibitory effect and mechanism of sophoridine on cell injury caused by hypoxia in rat myocardial cell line H9c2. Methods H9c2 cells were exposed to 1%O2 and cultured with different concentrations (1, 4 and 16 mmol/L) of sophoridine for 48 h. Cell viability was measured by MTT assay. The release level of LDH and the content of MDA were determined by the kit methods. Cell apoptosis was detected by flow cytometry. The protein expressions of cleaved caspase-3, p-PI3K and p-AKT were detected by Western blot. Results Sophoridine alleviated the cell viability, LDH release, MDA content and hypoxia-induced apoptosis of H9c2 cells in a concentration-dependent manner (P<0.05, P<0.01, P<0.001). Sophoridine decreased the cleaved caspase-3 expression and increased the ratios of p-PI3K/PI3K and p-AKT/AKT in a concentration-dependent manner (P<0.05, P<0.01, P<0.001). Conclusions Sophoridine can reduce the hypoxia-induced cell injury and has a protective effect on H9c2 cells, which may be achieved by enhancing the activity of PI3K/AKT signal.
Diagnostic value of traditional sweat chloride test for Chinese cystic fibrosis patients
2021, 41(3):  388-392. 
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Objective To explore the accuracy of the method of the traditional sweat chloride test and its diagnostic value for Chinese patients with cystic fibrosis (CF). Methods The sweat chloride level in 118 patients who were suspected to have CF in Peking Union Medical College Hospital from Jan 2012 to Oct 2019 were measured,and the results were compared with the final diagnosis of these patients. Results The patients’ age were 25.8±13.6 years old (15 months-67 years old). 102 out of 118 patients obtained valid chloride level from their sweat chloride tests with median concentration of 43.3 mmol/L (inter-quartile range: 23.8-120.1 mmol/L). 80 were consistent with their diagnosis. 39 patients had sweat chloride values higher than 60 mmol/L, while 31 (79.49%) of them were confirmed CF diagnosis. 40 cases of sweat chloride level lower than 30 mmol/L, of which 32 (80%) cases were excluded from CF. 23 patients had sweat chloride level between 30 and 59 mmol/L, while 17 (73.91%) of them were excluded from CF diagnosis. 66 (64.71%) of our patients had reproducible results of the sweat chloride level. Conclusions The positive predictive value of the sweat chloride test is 79.49% and the negative predictive value was 80% using traditional sweat chloride test measurement. The traditional sweat chloride test has high diagnostic value for Chinese CF patients.
Beneficial effect of skin micro-needling in the treatment of alopecia areata and its clinical assessment
2021, 41(3):  393-397. 
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Objective To investigate the effects of skin micro-needling in the treatment of alopecia areata (AA) using a dermaroller of 1.5 mm sized needles and in the stimulation of mouse dorsal hair growth using a dermaroller of 0.5 mm sized needles. Methods 52 patients with AA were recruited from the outpatient department of Renmin Hospital of Wuhan University (Mar. 2018-Aug. 2019) and further divided into two group, 5% minoxidil spray and 0.05% halomethasone cream were applied topically to the control group. The micro-needling group received micro-needling treatment combined with 5% minoxidil spray and 0.05% halomethasone cream Moreover, the dorsal hair of C57BL/6 mouse was removed and then divided into model group, minoxidil group and micro-needling group. From the next day, The corresponding treatment were given to the mice for 14 consecutive days. Anagen-associated pigmentation changes and hair growth were recorded using a digital camera. The morphology and number of anagenic hair follicles were observed using HE staining, the expression of K15 in the hair follicle was detected by an immunohistochemistry staining. Result The effective rate of the micro-needling group was superior to that of the control group (P<0.05), after the treatment of 12 weeks. As compared with the model group, the skin darkening appeared earlier and the hair length longer in the micro-needling group (P<0.05), the number of hair follicles and the K15 expression level were greatly increased in the micro-needling group compared with those in the model group (P<0.05). Conclusion The hair follicle epithelial stem cells may be activated by micro-needling, by which stimulates hair growth and helps to treat AA.
Increased expression of G protein-coupled receptor 84 after cerebral ischemia-reperfusion in rats
2021, 41(3):  398-403. 
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Objective To determine the expression and function of G protein-coupled receptor 84 (GPR84) in rat cerebral ischemia/reperfusion(I/R). Methods The rat middle cerebral artery occlusion (MCAO) models were performed, and rats were then divided into sham group, cerebral ischemia for 2 h and reperfusion for 12 h group (I/R 12 h), 24 h group (I/R 24 h), 48 h group (I/R 48 h), 72 h group (I/R 72 h); rat primary cortical neurons were isolated and cultured in vitro, neurons were then divided into control group (Con), oxygen-glucose deprivation (OGD) for 1 h and recovery for 24 h group (OGD/R 1 h), OGD for 2 h and recovery for 24 h group (OGD/R 2 h); neurons were transfected with GPR84 small interfering RNA (siRNA), cells were divided into Con group, OGD/R 1 h group, neurons pretreated with negative control and exposed to OGD for 1 h and recovery for 24 h group (NC+OGD/R 1 h), neurons pretreated with GPR84 siRNA and exposed to OGD for 1 h and recovery for 24 h group (GPR84 siRNA+OGD/R 1 h). Western blot was used to determine the expression of GPR84 protein. Immunofluorescence assay was used to analyze the localization of the immunofluorescence signals of GPR84 in cerebral ischemic tissues after MCAO. Terminal dexynucleotidyltransferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay was used to detect the percentage of apoptotic neurons. Results Compared with the sham group, the expression of GPR84 protein was significantly up-regulated at each time point after cerebral I/R (P < 0.01). In addition, GPR84 was mainly localized in neurons and microglia, while little GPR84 was found in astrocytes. Neurons in the MCAO group exhibited higher levels of GPR84 expression than those in the sham group. GPR84 knockdown significantly inhibited the increase of GPR84 expression induced by OGD/R. Compared with the control group, neurons in the OGD/R 1 h group exhibited higher apoptotic cells percentage. Compared with the NC+OGD/R 1 h group, the percentage of apoptotic neurons in GPR84 siRNA +OGD/R 1 h group was significantly decreased. Conclusions These results suggest that the elevated GPR84 expression induced by cerebral I/R may play an important role in cerebral ischemia injury.
Lnc-MALAT1 attenuates miR-217 inhibiting LPS-induced inflammatory response in rat alveolar macrophages
2021, 41(3):  404-408. 
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Objective To investigate the regulatory mechanism of long-chain noncoding RNA metastasis-associated lung adenocarcinoma 1 (LncMALAT1) in the inflammatory response of lipopolysaccharide (LPS) -induced alveolar macrophages (AMOs). Methods LPS treated AMOs cells to establish a cell damage model, and liposome method was used to transfect pcDNA, pcDNA-MALAT1, miR-NC, miR-217 mimics, pcDNA-MALAT1 and miR-NC, pcDNA-MALAT1 and miR-217 mimics to AMOs, LPS treatment for 12 h. The levels of IL-1β and TNF-α were detected by ELISA. RT-PCR was used to detect the expression of MALAT1 and miR-217 in cells. The dual luciferase report experiment and RIP experiment were used to detect the targeting relationship between MALAT1 and miR-217. Results Compared with the AMOs group, the levels of IL-1β and TNFα in the cells of the LPS+AMOs group were significantly increased (P<0.05), the expression of MALAT1 was significantly increased, and the expression of miR-217 was significantly reduced (P<0.05). The dual luciferase report experiment and RIP experiment confirmed that MALAT1 could target and bind to miR-217. Overexpression of MALAT1 could obviously promote the secretion of IL-1β and TNF-α induced by LPS in AMOs, while overexpression of miR-217 could inhibit the secretion of IL-1β and TNF-α induced by LPS in AMOs. Furthermore, overexpression of miR-217 could reduce the secretion of IL-1β and TNF-α induced by MALAT1 in LPS-induced AMOs. Conclusions MALAT1, a long non-coding RNA, can promote the inflammatory response of AMOs cells induced by LPS, and its mechanism is related to targeting miR-217.
Constructing a risk predictive model for recurrence following nephron-sparing surgery in the treatment of clear cell renal cell carcinoma
2021, 41(3):  409-414. 
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Objective To detect factors associated with recurrence after nephron sparing surgery (NSS) in patients with clinically localized clear cell renal cell carcinoma (ccRCC), and construct a model to predict the recurrence risk in five years following NSS. Methods Data of the patients who underwent NSS for renal occupied lesions between January 2006 and July 2019 at Peking University Third Hospital were retrospectively analyzed. Univariate and multivariate logistic regression analyses were used to assess the factors associated with recurrence of ccRCC after NSS, and a nomogram was established to predict recurrence risk in 5 years following NSS. Results 795 patients who were pathologically diagnosed with ccRCC were included in the study. Recurrence occurred in 17 cases, with the recurrence rate 2.1%. The median time to relapse was 35.7 months. In the univariable analysis, BMI (HR = 4.102, P < 0.05), the artery blocking time (HR = 0.194, P < 0.01) and tumor necrosis or sarcomatoid change (HR = 5.160, P < 0.01) were associated with recurrence-free survival of the ccRCC patients after NSS. The multivariate analysis showed that these three factors were independent factors of recurrence. The accuracy of our nomogram for recurrence risk was verified using internal validation, and the value of c-index was 0.843. Conclusions BMI, artery blocking time and tumor necrosis or sarcomatoid change were independent factors for recurrence following NSS in the ccRCC patients. We constructed an accurate model to predict the recurrence risk, which may help to provide personalized managements for the aforementioned patients after NSS.
Changes of peripheral blood lymphocyte subsets in patients with psoriasis
2021, 41(3):  415-418. 
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Objective To investigate the changes and immune mechanisms of peripheral blood lymphocyte subtypes in psoriasis patients with progressive, stationary and regression stages. Methods The percentage of lymphocytes in each group of peripheral blood mononuclear cells (PBMC) in 77 patients with psoriasis vulgaris including 28 patients during progression, 23 patients during stationary, and 26 patients during regression). Results The percentages of CD3+, CD4+ and CD8+ lymphocytes were significantly increased in both the progression and stationary stages compared with the regression stage (p<0.01). CD4-CD8- in the peripheral blood lymphocyte subsets of the patients showed a decreasing trend according to progression, stationary and regression stages (p<0.05). The result showed that the proportion of CD3-CD16+CD56+ increased with the stability of the disease, the proportion of CD45+CD14+ gradually decreased, and the proportion of CD3-CD19+ was negatively correlated with the disease (p<0.05). Conclusions there were significant differences in lymphocyte subsets in the three groups of patients with psoriasis, which further supported the immune pathogenesis of psoriasis.
Analysis of 2 cases of glycogen storage disease typeⅠb with Crohn’s disease in children
2021, 41(3):  419-422. 
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Objective To investigate the clinical characteristics of glycogen storage disease typeⅠb(GSDIb) with Crohn’s Disease(CD) and improve the recognition of the disease. Methods Retrospectively analyze the clinical characteristics of 2 cases diagnosed with the GSDⅠb and CD in the pediatric department of Peking Union Medical College Hospital. Results There were a boy and a girl in the study. They were genetically diagnosed as GSDⅠb at 2 years old. Gastrointestinal symptoms including abdominal pain and diarrhea were observed at 4, 5 years old respectively. They were diagnosed as CD with endoscopic characteristics and pathology results. Besides corn starch, granulocyte colony-stimulating factor(G-CSF) was used to elevate the neutrophil to above 1.0×109/L. The gastrointestinal symptoms got alleviated. 5-aminosalicylic acid agents were used as supportive treatment. Conclusion Evaluation of the gastrointestinal system is necessary as the abdominal pain or diarrhea were observed in GSDⅠb patients. G-CSF with a relatively low dose is the mainstay to elevate neutrophil with few side effects. Splenomegaly is the most common side effect. Monitor renal function is necessary in the use of 5-ASA.
Research progress of blood hypercoagulable state in liver cirrhosis
2021, 41(3):  423-427. 
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The patients with liver cirrhosis are often complicated with coagulation dysfunction, and blood hypercoagulable state is one of the manifestations. Blood hypercoagulable state is not only involved in thrombosis, the latest evidence confirms that it is also involved in liver injury, liver regeneration and liver fibrosis. This review attempts to discuss the mechanism of blood hypercoagulable state in liver cirrhosis, the role of intestinal microecological environment in it and the latest influence mechanism of blood hypercoagulable state in liver cirrhosis .To provide a new concept for the diagnosis and treatment of patients with liver cirrhosis.
Research progress on metabolism-related biomarkers of frailty in the elderly
2021, 41(3):  428-432. 
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Frailty is a common syndrome in the elderly, identified by the decline of physiological functions in multiple organ systems, which may lead to adverse outcomes such as falls, cognitive impairment, hospitalization or death. Early diagnosis of frailty is of great value in promoting healthy aging. At present, it can only be diagnosed by assessment tools in clinical without specific biomarkers. Frailty is often accompanied by metabolic disorders in many aspects, such as energy, muscle, bone, hormone and so on. The study of metabolism-related biomarkers can help early identification and intervention of frailty.
Progress in molecular mechanism and targeted drugs of bone metastases in breast cancer
2021, 41(3):  433-437. 
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Breast cancer is the most common malignant tumor that threatens the health of women around the world, and bone is the site most prone to metastasis. The molecular mechanism and targeted therapy drugs of bone metastasis in breast cancer have become the current research hotspots. This article summarizes the key molecules involved in regulating bone metastasis of breast cancer. The important targeted therapy drugs are introduced and their efficacy is compared from two categories of targeted osteoclasts and targeted osteoblasts.
Research progress of lipoedema
Liquan Wang Xiao Long
2021, 41(3):  438-441. 
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Lipoedema is an uncommon disorder. It is characterized by bilateral enlargement of the legs due to abnormal depositions of subcutaneous fat and essentially affects females. it is proved that the disease has a genetic tendency and can be also affected by hormone levels. Lipoedema has characteristic clinical manifestations, like pain, easy bruising and “cuff sign”. In clinical, main diseases considered for differential diagnosis are lymphedema, obesity, Dercum’s disease and chronic venous disease. Management of lipoedema includes psychological health education, weight loss, conservative treatment and operative treatment. There are increasing reports supporting liposuction using super tumescent local anesthesia and vibrating cannulas as the preferred surgical option.
Progress of ferroptosis mechanism and application in lung cancer therapy
2021, 41(3):  442-447. 
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Ferroptosis is a newly discovered cell death form characterized by accumulation of iron dependent lipid peroxidation and reactive oxygen species, which is different from apoptosis, necrosis and autophagy in morphology and biochemical. It is involved in the occurrence and development of lung cancer, and inducing ferroptosis seems to become a new strategy for anti-lung cancer. This review includes the followings: mechanism of ferroptosis from the aspects of iron metabolism, amino acid and glutathione metabolism and lipid metabolism; its effects on lung cancer occurrence and progression, chemotherapy resistance, radiation resistance and immunotherapy respectively, as to advance our understanding of ferroptosis and provides a new direction and new idea for lung cancer therapy.
Advances on the roles of tumor microenvironment in bone cancer pain
Jia-Hong ZHOU LI Min
2021, 41(3):  448-451. 
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Tumor microenvironment is the microenvironment in which tumors live in human body. Cancer-induced bone pain is one of the most common symptoms of tumor bone metastasis, . The mechanisms of bone cancer pain are complex, and the tumor acidic microenvironment, the related ion channels and inflammatory mediators released by cancer, play a role in bone cancer pain.
Analysis of the needs and feedbacks of diagnostic ultrasound teaching based on questionnaire at Peking Union Medical College
2021, 41(3):  452-455. 
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Objective This study aims to investigate the current situation of ultrasonographic education and curriculum demands of eight-year program clinical medical students in Peking Union Medical College. Methods Questionnaires of the current situation of ultrasonographic education and curriculum demands of eight-year program clinical medical students in Peking Union Medical College were collected and compared with feedbacks of the 4+4 pilot class medical students who received the new pattern ultrasonographic education. Results The eight-year program medical students showed poor knowledge of ultrasound discipline, basic ultrasonographic knowledge and ultrasonographic technology development. They were not satisfied with the current ultrasonographic education and hoped to improve the curriculum content, pattern and arrangement. Conclusion Based on these results, we suggested that the ultrasonographic curriculum design and teaching patterns should be adjusted according to students’ demands and emphasize basic knowledge such as ultrasonographic manifestations of normal human anatomy and common diseases.
Application of virtual simulation experiments in physiological experimental teaching
2021, 41(3):  456-458. 
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Objective To evaluate the teaching effects of physiological virtual simulation experiments. Methods In the spring semester of 2020, virtual simulation experiments were applied in the physiological experimental course in Peking University Health Science Center. The teaching effect was evaluated thereafter by a questionnaire based on the network tool “SOJUMP”. Results The results from the analysis of the 278 questionnaires showed that virtual simulation experiments based on the internet platform improved the students’ interest in learning, promoted the integration of theoretical and experimental teaching, and helped training the students’ independent learning ability. Virtual simulation experiments were recognized by the vast majority of the students. Conclusion The physiological virtual simulation experiment teaching platform can be a new teaching model and a complement to the traditional physiology experimental course. The combined application of virtual simulation experiments and animal experiments may achieve the integration of virtual and actual experiments so as to promote the quality of teaching and help training the students’ independent learning ability.
Application and evaluation of inquiry teaching mode for graduated students in large-scale biomedical equipments courses
2021, 41(3):  459-462. 
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Objectve To test and evaluate the inquiry teaching (IT) mode for graduated students in course named with Theory and Application of Large-scale Biomedical Equipment and provide the basis of its routinely application in the future. Methods Design the experiments basing on the IT mode. Summarize the strategy, performance and results. Evaluate the effects with the education monitoring theory and the statistic approaches. Analyze the questionnaire data quantitatively according to reliability, correlation coefficient and discrimination. Results Experiments derived from inquiry teaching (IT) mode were employed in grade 2017 and 2018 graduated students who choose this course. The primary results are satisfied. There are 23 students involving these tests. The reliability of questionnaire is good with α value 0.852. Twenty-three questions in 27 ones show the significant differences. Conclusions The results of questionnaire show satisfied reliability, correlation coefficient, discrimination and coverage. This indicates this mode could be employed in evaluation of teaching. The design, difficulty and schedule are the major factors of IT experiments.
Effect evaluation and feedback of ultrasound medicine online teaching during the COVID-19 pandemic
2021, 41(3):  463-466. 
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Objective To explore the application of in ultrasonic theory teaching online during the COVID-19 pandemic. Methods The standardized training of students, postgraduates and refresher doctors who have participated in the online theoretical teaching of ultrasound in the Peking University Third Hospital. Compared with the teaching assessment results in previous years as the objective index, the questionnaire was used as the subjective evaluation index. Results There was no significant difference between the experimental group and the control group. 78.3% of the questionnaire results showed that the online teaching method got better than the conventional offline teaching method. Conclusions The teaching quality of online teaching is similar to that of conventional teaching mode, and teachers and students have better acceptance and satisfaction of online teaching mode.
Basic & Clinical Medicine
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Add: 5 Dong Dan San Tiao, Beijing 100005
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E-mail: basic_clinic@vip.163.com
Website: http://jcyxylc.pumc.edu.cn