1.Impact of AMPKgamma silencing on AMPK activation and intracellular lipids regulation.
Acta Pharmaceutica Sinica 2013;48(6):842-7
The study is aimed to confirm the silencing efficiency of the vector in human hepatocellular liver carcinoma cell line (HepG2), and observe effects of AMPKgamma silencing on the AMPK stimulating activity and lipid synthesis of cordycepin (CCS), a natural product with known AMPK activating function. The downregulating efficacy of siRNAs on AMPKgamma expression was confirmed in our previous study. The double stranded shRNA Oligo was ligated to lentivirus vector and verified by sequencing. The lentiviral which can effectively inhibited protein expression levels of AMPKgamma was selected by Western blotting, and the regulation of CCS on protein expression of AMPKgamma and p-AMPK in AMPKgamma silence cells were detected by Western blotting analysis. The lipid accumulation in cells was observed by Oil-Red O stain and cells were collected for the estimation of cholesterol (TC), triglyceride (TG). The results showed that the lentiviral vector carrying a shRNA targeting the AMPKgamma gene was successfully constructed. Western blotting analysis confirmed that GR085 had the highest interfering efficiency. Treatment with CCS can significantly increase the levels of phospho-AMPK in normal cells, and the level of TC, TG was reduced, but in AMPKgamma silence cells the effects of CCS on AMPK activation and lipid synthesis were almost completely abolished without changing the expression levels of total AMPK or AMPKgamma protein. In conclusion, the AMPKgamma gene may be related to AMPK activation and intracellular lipids regulation by CCS.
2.Expression of Hsa_circ_0003928 in diabetic foot ulcer and its relationship with severity and prognosis
Shaona WANG ; Jiaohua WANG ; Yubin WU ; Yonghuan SU ; Wenli ZHAN
Chinese Journal of Diabetes 2024;32(2):108-112
Objective To investigate the expression of serum Hsa_circ_0003928 in patients with diabetic foot ulcer(DFU)and its relationship with the severity and prognosis of disease.Methods 113 DFU patients were selected as the study subjects.According to the severity of infection,19 cases were classified into level 1(no infection),40 cases at level 2(mild infection),20 cases at level 3(moderate infection),and 34 cases at level 4(severe infection).According to the prognosis of DFU patients,they were divided into good prognosis group(GP,n=63)and poor prognosis group(PP,n=50).The baseline data and levels of IL-6,C-RP and Hsa_circ_0003928 were compared among the four groups.Logistic regression was used to analyze the risk factors of poor prognosis in patients with DFU.The receiver operating characteristic(ROC)curve was used to analyze the value ofHsa_circ_0003928,C-RP and IL-6 in predicting the poor prognosis in DFU patients.Results The DFU duration,infection grade 3~4,serum creatinine,uric acid,BUN,C-RP,IL-6 and Hsa_circ_0003928 levels in PP group were significantly higher than those in GP group(P<0.05 or P<0.01).Grade 3~4 DFU patients had higher Hsa_circ_0003928 expression than grade 1~2(P<0.01).Logistic regression analysis showed that long duration of DFU,infection grade 3~4,higher levels of BUN,C-RP,IL-6 and Hsa_circ_0003928 were risk factors for poor prognosis in DFU patients.ROC curve showed that Hsa_circ_0003928 had the greatest AUC(0.882,95%CI 0.819~0.942)in predicting poor prognosis in DFU patients,with sensitivity 87.5%and specificity 85.6%,respectively.Conclusion Elevated Hsa_circ_0003928 is associated with DFU severity and poor prognosis,which has certain predictive value for the prognosis of DFU patients.
3.Vinpocetine, a phosphodiesterase 1 inhibitor, mitigates atopic dermatitis-like skin inflammation
Yeon Jin LEE ; Jin Yong SONG ; Su Hyun LEE ; Yubin LEE ; Kyu Teak HWANG ; Ji-Yun LEE
The Korean Journal of Physiology and Pharmacology 2024;28(4):303-312
Atopic dermatitis (AD) is the most common inflammatory pruritic skin disease worldwide, characterized by the infiltration of multiple pathogenic T lymphocytes and histological symptoms such as epidermal and dermal thickening. This study aims to investigate the effect of vinpocetine (Vinp; a phosphodiesterase 1 inhibitor) on a 1-chloro-2,4-dinitrobenzene (DNCB)-induced AD-like model. DNCB (1%) was administered on day 1 in the AD model. Subsequently, from day 14 onward, mice in each group (Vinp-treated groups: 1 mg/kg and 2 mg/kg and dexamethasone- treated group: 2 mg/kg) were administered 100 µl of a specific drug daily, whereas 0.2% DNCB was administered every other day for 30 min over 14 days. The Vinp-treated groups showed improved Eczema Area and Severity Index scores and trans-epidermal water loss, indicating the efficacy of Vinp in improving AD and enhancing skin barrier function. Histological analysis further confirmed the reduction in hyperplasia of the epidermis and the infiltration of inflammatory cells, including macrophages, eosinophils, and mast cells, with Vinp treatment. Moreover, Vinp reduced serum concentrations of IgE, interleukin (IL)-6, IL-13, and monocyte chemotactic protein-1. The mRNA levels of IL-1β, IL-6, Thymic stromal lymphopoietin, and transforming growth factor-beta (TGF-β) were reduced by Vinp treatment. Reduction of TGF-β protein by Vinp in skin tissue was also observed. Collectively, our results underscore the effectiveness of Vinp in mitigating DNCB-induced AD by modulating the expression of various biomarkers. Consequently, Vinp is a promising therapeutic candidate for treating AD.
4.Analysis of technical difficulties of single-port and reduced port laparoscopic radical gastrectomy for gastric cancer
Su YAN ; Xinfu MA ; Kang ZHAO ; Xiaoqian CHEN ; Can GUO ; Qingqing WANG ; Liang WANG ; Chun WANG ; Chenghao LIU ; Yubin MA
Chinese Journal of Digestive Surgery 2019;18(3):222-228
Single-port and reduced-port laparoscopic radical gastrectomy as the innovative surgery for gastric cancer are gradually accepted nowadays,and more attentions are also paid to single-port and reduced-port laparoscopic radical gastrectomy due to its better cosmetic effect,less pain,lower incidence rate of surgical site infection and more advantages in enhanced recovery after surgery.However,in the early stage,the development of single-port and reduced-port laparoscopic radical gastrectomy were facing challenges and obstacles on account of limited surgical skills,the lack of special laparoscopic instruments,laparoscope and multi-port Trocar.In recent years,the dilemma and difficult situations were gradually resolved following by surgical techniques innovation,laparoscopic instruments and facilities improvement.It is believed that single-port and reduced-port laparoscopic radical gastrectomy will have a good prospect and breakthrough in the field of gastric cancer treatment in the future.
5.MicroRNA-145 Gene Modification Enhances the Retention of Bone Marrow-Derived Mesenchymal Stem Cells within Corpus Cavernosum by Targeting Krüppel-Like Factor 4
Daoyuan HU ; Yunlong GE ; Yuhang XI ; Jialiang CHEN ; Hua WANG ; Chi ZHANG ; Yubin CUI ; Lizhao HE ; Ying SU ; Jun CHEN ; Cheng HU ; Hengjun XIAO
The World Journal of Men's Health 2024;42(3):638-649
Purpose:
The poor retention and ambiguous differentiation of stem cells (SCs) within corpus cavernosum (CC) limit the cell application in erectile dysfunction (ED). Herein, the effects and mechanism of microRNA-145 (miR-145) gene modification on modulating the traits and fate of bone marrow-derived mesenchymal stem cells (BMSCs) were investigated.
Materials and Methods:
The effects of miR-145 on cell apoptosis, proliferation, migration, and differentiation were determined by flow cytometry, cell counting kit-8, transwell assays and myogenic induction. Then, the age-related ED rats were recruited to four groups including phosphate buffer saline, BMSC, vector-BMSC, overexpressed-miR-145-BMSC groups. After cell transplantation, the CC were harvested and prepared to demonstrate the retention and differentiation of BMSCs by immunofluorescent staining. Then, the target of miR-145 was verified by quantitative real-time polymerase chain reaction and immunohistochemical. After that, APTO-253, as an inducer of Krüppel-like factor 4 (KLF4), was introduced for rescue experiments in corpus cavernosum smooth muscle cells (CCSMCs) under the co-culture system.
Results:
In vitro, miR-145 inhibited the migration and apoptosis of BMSCs and promoted the differentiation of BMSCs into smooth muscle-like cells with stronger contractility. In vivo, the amount of 5-ethynyl-2′-deoxyuridine (EdU)+cells within CC was significantly enhanced and maintained in the miR-145 gene modified BMSC group. The EdU/CD31 co-staning was detected, however, no co-staining of EdU/α-actin was observed. Furthermore, miR-145, which secreted from the gene modified BMSCs, dampened the expression of KLF4. However, the effects of miR-145 on CCSMCs could be rescued by APTO-253.
Conclusions
Overall, miR-145 modification prolongs the retention of the transplanted BMSCs within the CC, and this effect might be attributed to the modulation of the miR-145/KLF4 axis. Consequently, our findings offer a promising and innovative strategy to enhance the local stem cell-based treatments.
6.Protective effect of ulinastatin on acute bone loss in sepsis
Peng YANG ; Yubin TANG ; Jing YANG ; Jian LIU ; Runjie YAO ; Lin CHEN ; Nan SU
Chinese Journal of Tissue Engineering Research 2024;28(35):5649-5655
BACKGROUND:Sepsis-induced systemic inflammation leads to rapid bone mass loss;however,there is a lack of effective treatments.Ulinastatin is an anti-inflammatory drug,but its protective effect and mechanism on bone under sepsis-induced systemic inflammation are still unclear. OBJECTIVE:To explore whether ulinastatin can relieve acute bone loss caused by lipopolysaccharide. METHODS:(1)Animal experiment.Thirty male C57BL/6 mice were randomly divided into three groups(n=10 per group):control group,model group and experimental group.The control group was injected intraperitoneally with normal saline,the model group was injected intraperitoneally with lipopolysaccharide,and the experimental group was injected intraperitoneally with lipopolysaccharide and ulinastatin.In the experimental group,ulinastatin was injected continuously for 3 days.After intraperitoneal injection of ulinastatin for 14 days,femoral tissues were taken for CT scanning and pathological observation.(2)Cell experiment.C57BL/6 mouse primary osteoblasts were isolated and divided into three groups:the control group was routinely cultured,lipopolysaccharide was added to the model group,and lipopolysaccharide with ulinastatin was added to the experimental group.Cell proliferation and osteogenic differentiation were detected.C57BL/6 mouse bone marrow mononuclear cells were isolated and divided into three groups:the control group was routinely cultured,lipopolysaccharide was added to the model group,and lipopolysaccharide and ulinastatin were added to the experimental group.Osteoclast differentiation was detected. RESULTS AND CONCLUSION:(1)Animal experiment.CT scanning and hematoxylin-eosin staining showed that bone mass in lipopolysaccharide-treated mice was reduced but increased after treatment with ulinastatin.Tartrate resistant acid phosphatase staining showed that the number of osteoclasts in bone tissue increased in the model group,but significantly decreased in the experimental group compared with the model group.(2)Cell experiment.Cell counting kit-8 assay showed that lipopolysaccharide treatment inhibited the proliferation of osteoblasts,and ulinastatin elevated the proliferation of osteoblasts after lipopolysaccharide treatment.Alkaline phosphatase staining,alizarin red staining and osteogenesis-related gene(alkaline phosphatase,Runx2,osteocalcin,osteoblastin,nuclear factor κB receptor-activating factor ligand,osteoprotegerin)detection showed that lipopolysaccharide treatment inhibited osteogenic differentiation of osteoblasts and elevated the nuclear factor κB receptor-activating factor ligand/osteoprotegerin ratio;ulinastatin did not have any significant effect on the reduction of osteoblast function induced by lipopolysaccharide but decreased the nuclear factor κB receptor-activating factor ligand/osteoprotegerin ratio.Tartrate resistant acid phosphatase staining and osteoclast-related gene(tartrate resistant acid phosphatase and matrix metalloproteinase 9)detection showed that lipopolysaccharide treatment could promote osteoclast differentiation of bone marrow monocytes,while ulinastatin could inhibit lipopolysaccharide-induced osteoclast differentiation of bone marrow monocytes.(3)Overall,ulinastatin can significantly inhibit lipopolysaccharide-induced bone loss,mainly through promoting osteoblast proliferation and directly or indirectly inhibiting osteoclast differentiation to alleviate bone loss and achieve osteoprotective effects.
7.Promotion effect of TGF-β-Zfp423-ApoD pathway on lip sensory recovery after nerve sacrifice caused by nerve collateral compensation.
Pingchuan MA ; Gaowei ZHANG ; Su CHEN ; Cheng MIAO ; Yubin CAO ; Meng WANG ; Wenwen LIU ; Jiefei SHEN ; Patrick Ming-Kuen TANG ; Yi MEN ; Li YE ; Chunjie LI
International Journal of Oral Science 2023;15(1):23-23
Resection of oral and maxillofacial tumors is often accompanied by the inferior alveolar nerve neurectomy, resulting in abnormal sensation in lower lip. It is generally believed that spontaneous sensory recovery in this nerve injury is difficult. However, during our follow-up, patients with inferior alveolar nerve sacrifice showed different degrees of lower lip sensory recovery. In this study, a prospective cohort study was conducted to demonstrate this phenomenon and analyze the factors influencing sensory recovery. A mental nerve transection model of Thy1-YFP mice and tissue clearing technique were used to explore possible mechanisms in this process. Gene silencing and overexpression experiments were then conducted to detect the changes in cell morphology and molecular markers. In our follow-up, 75% of patients with unilateral inferior alveolar nerve neurectomy had complete sensory recovery of the lower lip 12 months postoperatively. Patients with younger age, malignant tumors, and preservation of ipsilateral buccal and lingual nerves had a shorter recovery time. The buccal nerve collateral sprouting compensation was observed in the lower lip tissue of Thy1-YFP mice. ApoD was demonstrated to be involved in axon growth and peripheral nerve sensory recovery in the animal model. TGF-β inhibited the expression of STAT3 and the transcription of ApoD in Schwann cells through Zfp423. Overall, after sacrificing the inferior alveolar nerve, the collateral compensation of the ipsilateral buccal nerve could innervate the sensation. And this process was regulated by TGF-β-Zfp423-ApoD pathway.
Mice
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Animals
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Lip/innervation*
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Prospective Studies
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Mandibular Nerve/pathology*
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Sensation/physiology*
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Trigeminal Nerve Injuries/pathology*
8.GLUL stabilizes N-Cadherin by antagonizing β-Catenin to inhibit the progresses of gastric cancer.
Qiwei JIANG ; Yong LI ; Songwang CAI ; Xingyuan SHI ; Yang YANG ; Zihao XING ; Zhenjie HE ; Shengte WANG ; Yubin SU ; Meiwan CHEN ; Zhesheng CHEN ; Zhi SHI
Acta Pharmaceutica Sinica B 2024;14(2):698-711
Glutamate-ammonia ligase (GLUL, also known as glutamine synthetase) is a crucial enzyme that catalyzes ammonium and glutamate into glutamine in the ATP-dependent condensation. Although GLUL plays a critical role in multiple cancers, the expression and function of GLUL in gastric cancer remain unclear. In the present study, we have found that the expression level of GLUL was significantly lower in gastric cancer tissues compared with adjacent normal tissues, and correlated with N stage and TNM stage, and low GLUL expression predicted poor survival for gastric cancer patients. Knockdown of GLUL promoted the growth, migration, invasion and metastasis of gastric cancer cells in vitro and in vivo, and vice versa, which was independent of its enzyme activity. Mechanistically, GLUL competed with β-Catenin to bind to N-Cadherin, increased the stability of N-Cadherin and decreased the stability of β-Catenin by alerting their ubiquitination. Furthermore, there were lower N-Cadherin and higher β-Catenin expression levels in gastric cancer tissues compared with adjacent normal tissues. GLUL protein expression was correlated with that of N-Cadherin, and could be the independent prognostic factor in gastric cancer. Our findings reveal that GLUL stabilizes N-Cadherin by antagonizing β-Catenin to inhibit the progress of gastric cancer.