Inhibitory Effect of Jinkui Shenqi Pills on Glucocorticoid-Enhanced Axial Length Elongation in Experimentally Myopic Guinea Pigs / 中国结合医学杂志

OBJECTIVE@#To explore the underlying mechanism of inhibition by Jinkui Shenqi Pills (JKSQP) on glucocorticoid-enhanced axial length elongation in experimental lens-induced myopia (LIM) guinea pigs.@*METHODS@#Sixty 2-week old male guinea pigs were randomly divided into 4 groups with 15 guinea pigs in each group, according to the random numbers generated by SPSS software: control, LIM, saline and JKSQP groups. The control group includes animals with no treatment, while the guinea pigs in the other 3 groups received lens-induced myopization on the right eyes throughout the experiment (for 8 weeks). The saline and JKSQP groups were given daily intraperitoneal injections of 10 mg/kg hydrocortisone for 2 consecutive weeks at the same time, and then orally administered either saline or JKSQP [13.5 g/(kg•d) for 6 consecutive weeks. Body weight, anal temperature and animal appearance were observed and recorded to evaluate the GC-associated symptoms. The ocular parameters, including refraction and axial length, were measured by streak retinoscopy and A-scan ultrasonography, respectively. The levels of plasma hormones associated with the hypothalamic-pituitary-adrenal axis (HPAA), including free triiodothyronine, free thyroxine, estradiol and testosterone, were measured by radioimmunoassay, and cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate were measured by enzyme-linked immunosorbent assay. In addition, the mRNA and protein expressions of retinal amphiregulin (AREG) was measured by quantitative real-time polymerase chain reaction and Western blotting, respectively.@*RESULTS@#JKSQP effectively increased body weight and anal temperature, improved animal appearance and suppressed axial length elongation in glucocorticoid-enhanced myopic guinea pigs with normalization of 4 HPAA-associated plasma hormones (all P<0.05). The plasma level of cAMP was significantly increased, whereas the plasma level of cGMP and the mRNA and protein expressions of retinal AREG were decreased after treatment with JKSQP (all P<0.05).@*CONCLUSION@#JKSQP exhibited a significant inhibitory effect on axial length elongation with decreased expression of AREG in the retina, and normalized 4 HPAA-associated plasma hormones and the expression of cAMP and cGMP in GC-enhanced myopic guinea pigs.

The repair effect and mechanism of amphiregulin on injured lung tissue in mice / 中华急诊医学杂志

Objective:To investigate the repair effect of amphiregulin (Areg) on injured lung tissue in mice with acute respiratory distress syndrome (ARDS) and its underlying mechanism.Methods:The ARDS mouse model was made by tracheal infusion of lipopolysaccharide (LPS), and bronchoalveolar lavage fluid (BALF) was extracted for 7 consecutive days. Adult male C57BL/6 mice were randomly (random number) divided into 5 groups ( n=4 per group): (1) Control group; (2) Areg group: mice were treated intraperitoneally (i.p.) with recombinant Areg; (3) LPS+PBS group; (4) LPS+Areg group; and (5) LPS+Anti-Areg group; mice were instilled with LPS, then were injected i.p. with PBS, Areg or Areg neutralization antibody (Anti-Areg) 30 min later. Lung tissue and BALF were extracted at day 1, 3, 5 and 7 after ARDS. HE staining was used to evaluate the pathological changes of lung tissues. The total protein content in BALF was detected by BCA method, and the concentrations of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), IL-1β and immunoglobulin M (IgM) were determined by ELISA method. The phosphorylated levels of epidermal growth factor receptor (EGFR) and expressions of proliferating cell nuclear antigen (PCNA) and surface proteins-C (SP-C) were tested by Western blot. The immunofluorescence was used to detect the co-expression of PCNA and SP-C in lung tissues. One-way analysis of variance was used to compare the mean values of normally distributed measurement data between groups. Comparisons between groups were performed using the least significant difference t-test. Results:Compared with that at before modeling [(51.05±2.47) pg/mL], Areg concentrations were increased significantly at day 1 [(71.97±6.51) pg/mL; P<0.01] and day 3 [(147.58±7.56) pg/mL, P<0.01] in the BALF after ARDS. At day 1 after ARDS, there were significant interstitial edema, neutrophil infiltration and alveolar collapse in the LPS+PBS group and LPS+Areg group. Compared with the LPS+PBS group at day 3, 5 and 7, the pathological changes of lung tissues were notably improved in the LPS+Areg group, while were more serious in the LPS+Anti-Areg group. Compared with the control group, the LPS+PBS group had higher levels of neutrophil number, total protein, IgM, TNF-α, IL-1β, and IL-6. However, Areg treatment significantly reduced the levels of these indicators. Moreover, the expressions of PCNA (1.34±0.10), SP-C (1.48±0.10) and p-EGFR (0.92±0.032) in the LPS+Areg group were significantly up-regulated compared with those in the LPS+PBS group (0.88±0.03, 1.06±0.15, and 0.68±0.03, all P<0.01). And compared with the LPS+PBS group, PCNA and SP-C double positive cells were significantly increased in the LPS+Areg group, but decreased in the LPS+Anti-Areg group. Conclusions:Areg enhances the proliferation of alveolar typeⅡ epithelial cells by activating EGFR pathway, therefore promotes the repair of lung tissues during ARDS development.

Expression of amphiregulin in enchondromas and central chondrosarcomas

OBJECTIVES: The aim of this study was to evaluate the role of amphiregulin protein, an epidermal growth factor receptor ligand, in cartilaginous tumors. METHODS: Amphiregulin expression was examined in 31 enchondromas and 67 chondrosarcomas using immunohistochemistry analysis. RESULTS: Overall, 15 enchondromas (48.40%) and 24 chondrosarcomas (35.82%) were positive for amphiregulin. According to the receiver operating characteristic curve test, no difference in amphiregulin expression was observed between enchondromas and low-grade chondrosarcomas (p=0.0880). Additionally, 39 lesions (16 in short bones, 13 in long bones, and 10 in flat bones) were positive for amphiregulin, exhibiting a higher percentage of positive cells (p=0.0030) and intensity of immunohistochemical expression (p=0.0055) in short bone lesions than in others. Among 25 enchondromas localized in short bones, 15 expressed amphiregulin; however, all 6 cases localized in long bones were negative for this marker (p=0.0177). CONCLUSIONS: Amphiregulin did not help in distinguishing enchondromas from low-grade chondrosarcomas. The present study is the first to document the expression of this immunohistochemical marker in enchondromas. Furthermore, amphiregulin expression in enchondromas was localized in short bones, indicating a phenotypic distinction from that in long bones. This distinction may contribute to an improved understanding of the pathogenesis of these lesions.

Isorhapontigenin protects against doxorubicin-induced cardiotoxicity

As an effective anticancer drug, the clinical limitation of doxorubicin (Dox) is the time- and dose-dependent cardiotoxicity. Yes-associated protein 1 (YAP1) interacts with transcription factor TEA domain 1 (TEAD1) and plays an important role in cell proliferation and survival. However, the role of YAP1 in Dox-induced cardiomyopathy has not been reported. In this study, the expression of YAP1 was reduced in clinical human failing hearts with dilated cardiomyopathy and Dox-induced

IL-22 regulates positively amphiregulin expression of keratinocytes involved in psoriasis / 西安交通大学学报(医学版)

Objective: To investigate the effect of IL-22 on the expression of Amphiregulin (AREG) in HaCaT and further understand its role in psoriasis vulgaris (PsV). Methods: The mRNA expressions of IL-22, IL-22R1, IL-22BP and AREG were detected by RT-qPCR in PsV lesions (n=26) and healthy control (HC) skin specimens (n=15). RT-qPCR and Western blot were applied to detect the expression of AREG in HaCaT cells stimulated with IL-22 (20 ng/mL) and its soluble receptor IL-22BP (20 ng/mL) for 24 h. Results: The mRNA expressions of IL-22 (P<0.001), IL-22R1 (P<0.001) and AREG (P<0.01) were significantly increased respectively by 36 times, 24 times and 15 times in PsV compared with those in HC. In addition, there were positive correlations between the mRNA levels of AREG and IL-22 (r=0.49, P<0.05). IL-22 could upregulate the mRNA level of AREG by 22 times and protein expression by 6 times in HaCaT cells (P<0.01). IL-22BP could inhibit the effect of IL-22 (P<0.05). Conclusion: IL-22 may regulate positively amphiregulin expression of keratinocytes involved in psoriasis, and IL-22BP may inhibit this role as a blocker in treating psoriasis.

Expression of amphiregulin in different types of nasal polyps and its correlation with tissue remodeling / 中华耳鼻咽喉头颈外科杂志

Objective@#To explore the expression of amphiregulin (AREG) in nasal polyps patients with different degrees of eosinophil infiltration, and to analyze the correlation between AREG and tissue remodeling.@*Methods@#Forty-eight patients underwent endoscopic sinus surgery in the Department of Otorhinolaryngology Head and Neck Surgery, Remin Hospital, Wuhan University from July 2017 to August 2018 were recruited, including 40 males and 8 females, aged from 16 to 60 years old. The subjects were divided into three groups: control group (n=14), eosinophilic chronic sinusitis with nasal polyps (ECRSwNP) group (n=19) and noneosinophilic chronic rhinosinusitis with nasal polyps (non-ECRSwNP) group (n=15). The relative expression of AREG in nasal mucosa was detected by Western blot assay and immunohistochemical staining. Tissue remodeling was detected by HE staining, AB-PAS staining and Masson staining. Kruskal-Wallis test was used for comparison among multiple groups, and Spearman correlation analysis was conducted between the expression level of AREG and the related indexes of tissue remodeling.@*Results@#The expression of AREG in ECRSwNP group was significantly higher than that in non-ECRSwNP group and control group (median protein expression of Western blot was 1.592 vs 0.617 vs0.582, all P<0.05). The degree of epithelial injury and goblet cell metaplasia in ECRSwNP group was significantly higher than that in control group (all P<0.05), the percentage of collagen fibrosis area in ECRSwNP group was significantly lower than that in control group (P=0.01). In chronic rhinosinusitis with nasal polyps (CRSwNP) patients, the area of mucous glands was negatively correlated with the expression of AREG (r=-0.616, P<0.05), and the percentage of collagen fibrosis area was negatively correlated with the expression of AREG (r=-0.738, P<0.05).@*Conclusion@#The expression of AREG is higher in ECRSwNP patients, which is related to the process of tissue remodeling.

Effects of amphiregulin antibody on the eyes of guinea pigs with binocular lens-induced myopic / 眼科新进展

Objective To investigate the effects of amphiregulin antibody on the axial length,diopter and posterior sclera thickness in eyes of bilateral lens-induced myopic guinea pigs.Methods A total of 60 healthy three-colored short-hair guinea pigs were randomly divided into 5 groups:myopic model group,low dose group,medium dose group,high dose group and normal control group,12 rats in each group,and both eyes of the guinea pigs in the former four groups were induced by-10 D lens,while the normal group did not make any treatment.After wearing of goggles for 2 weeks,the right eyes of guinea pigs were intravitreally administrated with 5 μg,10 μg,20 μg anphiregulin antibody for the low,medium and high dose group,respectively.At the same time,the left eyes of these groups were given the same dose of Ringer' s buffer (buffer group).Continuous wearing of the lenses before and after injection of antibody was allowed.Diopter and axial length of guinea pigs were measured before wearing of the lenses,2 weeks after wearing lenses and 5 weeks after the experiment (after 3-times injections of antibody).Moreover,the thicknesses of retina,choroid and sclera in the posterior pole were detected by PAS staining.Finally,the expression of amphiregulin and EGFR mRNA and protein was detected by real time quantitative PCR and immunofluorescence.Results After 2 weeks of lens induction,the axial length of the myopic model group increased,but the diopter decreased when compared with the normal control group,and the differences were statistically significant (both P ＜ 0.05).Compared with the buffer group,after intravitreal injection of amphiregulin antibody,the axial length in the low,medium,and high dose groups decreased,while the diopter increased,and the scleral thickness at the posterior pole increased significantly in a dose-dependent manner,with statistically significant differences (all P ＜ 0.05).However,there was no significant change in retinal thickness at the posterior pole of all groups.Real-time quantitative PCR and immunofluorescence showed the expression of amphiregulin mRNA and protein in the retina was upregulated the myopic model group and buffer group,but downregulated in the high,medium,and low dose groups.Furthermore,when compared with the buffer group,the expression level of epidermal growth factor receptor in the retina of the low dose group was decreased (t =2.606,P =0.022),but there was no significant difference in the other groups.Conclusion After injection of amphiregulin antibody into the eyes of bilateral lens-induced myopic guinea pigs,the diopter increases,but the axial length is significantly shortened,and the posterior sclera is thickened,which may involve a decrease in the expression of amphiregulin.

Effect of amphiregulin on focal cerebral ischemia/reperfusion injury in rats / 中国脑血管病杂志

Objectives To observe the effect of amphiregulin (Areg) via lateral ventricle injection on focal cerebral ischemia/reperfusion (I/R) injury in rats and to investigate its possible mechanism. Methods A total of 96 3-month old health specified pathogen free SD rats were randomly divided into 6 groups (n=16 in each group):sham operation group (sham group),only exposure of common carotid artery and bifurcation;I/R group,making I/R model;solvent control group,lateral ventricle injection of standard protein solution(5 μl);Areg group,lateral ventricle injection of Areg(2 μg/5 μl);AG1478 group [AG1478,a blocker of Areg receptor epidermal growth factor receptor(EGFR),lateral ventricle injection of AG1478 (2.5 μg/5 μl);Areg combined AG1478 (AAG) group,lateral ventricle giving AG1478 (2.5 μg/5 μl),and then giving Areg (2 μg/5 μl) after 30 mm.The model of focal cerebral I/R injury was induced after 30 min administration of the above last 4 groups.After 24 h of reperfusion,the volume of cerebral infarction, the neurobehavioral score and the number of apoptotic cells in the brain tissue were compared among the groups. After 6 h of reperfusion,the phosphorylation levels of EGFR and protein kinase B(Akt)in ischemic brain tissue were detected. Results Compared with the sham group,the cerebral infarction volume and the number of apoptotic cells in brain tissue were increased significantly,while the neurobehavioral score was decreased(all P<0.05).Compared with the I/R group,the volume of cerebral infarction,the number of apoptotic cells in the brain tissue were decreased significantly,and the neurobehavioral score was increased in the Areg group,the levels of EGFR and Akt phosphorylation were significantly higher (all P <0. 05). Compared with the I/R group,the volume of cerebral infarction and the number of apoptotic cells of the AG1478 group were increased,the levels of EGFR and Akt phosphorylation were decreased(all P<0.05);Compared with the Areg group,the volume of cerebral infarction and the number of apoptotic cells of the AAG group and AG1478 group were increased significantly,and the levels of EGFR and Akt phosphorylation were decreased significantly(all P<0.05). Conclusions Areg reduces the infarct volume in ischemic brain tissue,improves nerve function,and inhibits apoptosis by activating EGFR-Akt signaling pathway. Therefore,it has some protective effect for cerebral I/R injury.

Overexpression of amphiregulin promotes the growth of mouse colon carcinoma CT26 cells in vivo / 肿瘤

Objective: To investigate the effect of epidermal growth factoramphiregulin (AREG) on the growth of mouse colon carcinoma CT26cells and its related mechanisms.Methods: The protein expression level of AREG in different mouse cancer cells was detected by ELISA. Mouse colon carcinoma CT26 cells, melanoma B16 cellsand hepatocellular carcinoma LPC-Akt cells were transfected with the recombinant lentiviralplasmid carrying AREG gene, while the ones transfected with empty plasmid were used asthe negative controls. After AREG overexpression, the cell proliferation, colony-formingabilities and cell cycle progression in vitro were detected by MTT, colony-forming assay andFCM, respectively. After the homograft mouse model of CT26 cells was constructed, thegrowth of homograft tumor was observed, the distribution of immune cells in tumor tissueswas detected by FCM, furthermore the expression of chemokine was detected by real-timefluorescent quantitative PCR.Results: The levels of AREG expression were relatively low in mouse colon carcinoma CT26cells, melanoma B16 cells and hepatoma LPC-Akt cells. AREG overexpression did notmarkedly affect the proliferation, colony-forming abilities and cell cycle progression of thesethree types of tumor cells in vitro (all P > 0.05). However, in the homograft mouse model ofCT26 cells, AREG overexpression significantly promoted the growth of tumor cells in vivo (P <0.01), decreased the percentage of CD8+ T cells (P < 0.05), and reduced the mRNA level ofCC chemokine 5 ligand (CCL5) (P < 0.05) which was related to CD8+ T cell recruitment.Conclusion: AREG promotes the growth of mouse colon carcinoma CT26 cells in vivo . AREGmay affect the tumor microenvironment by regulating the production of chemokine which isrelated to CD8+ T cell recruitment.

Modifiers of TGF-beta1 effector function as novel therapeutic targets of pulmonary fibrosis

Pulmonary fibrosis is a fatal progressive disease with no effective therapy. Transforming growth factor (TGF)-beta1 has long been regarded as a central mediator of tissue fibrosis that involves multiple organs including skin, liver, kidney, and lung. Thus, TGF-beta1 and its signaling pathways have been attractive therapeutic targets for the development of antifibrotic drugs. However, the essential biological functions of TGF-beta1 in maintaining normal immune and cellular homeostasis significantly limit the effectiveness of TGF-beta1-directed therapeutic approaches. Thus, targeting downstream mediators or signaling molecules of TGF-beta1 could be an alternative approach that selectively inhibits TGF-beta1-stimulated fibrotic tissue response while preserving major physiological function of TGF-beta1. Recent studies from our laboratory revealed that TGF-beta1 crosstalk with epidermal growth factor receptor (EGFR) signaling by induction of amphiregulin, a ligand of EGFR, plays a critical role in the development or progression of pulmonary fibrosis. In addition, chitotriosidase, a true chitinase in humans, has been identified to have modulating capacity of TGF-beta1 signaling as a new biomarker and therapeutic target of scleroderma-associated pulmonary fibrosis. These newly identified modifiers of TGF-beta1 effector function significantly enhance the effectiveness and flexibility in targeting pulmonary fibrosis in which TGF-beta1 plays a significant role.

The Effect of Interleukin-4 and Amphiregulin on the Proliferation of Human Airway Smooth Muscle Cells and Cytokine Release

Airway smooth muscle (ASM) hyperplasia and angiogenesis are important features associated with airway remodeling. We investigated the effect of IL-4 and amphiregulin, an epidermal growth factor family member, on the proliferation of human ASM cells and on the release of vascular endothelial growth factor (VEGF) and monocyte chemotactic protein (MCP)-1 from human ASM cells. Human ASM cells were growth-arrested for 48 hr and incubated with platelet-derived growth factor (PDGF)- BB, interleukin (IL)-4, amphiregulin, and VEGF to evaluate cell proliferation. The cells were treated with PDGF, IL-4 and amphiregulin to evaluate the release of VEGF, MCP-1. IL-4 suppressed unstimulated and PDGF-stimulated ASM cell proliferation. Amphiregulin stimulated ASM cell proliferation in a dose-dependent manner. VEGF did not have any influence on ASM cell proliferation. IL-4 stimulated VEGF secretion by the ASM cells in a dose-dependent manner and showed added stimulatory effects when co-incubated with PDGF. Amphiregulin did not promote VEGF secretion. IL-4 and amphiregulin showed no stimulatory effects on MCP-1 secretion. The results of this study showed that IL-4 had bifunctional effects on airway remodeling, one was the suppression of the proliferation of the ASM cells and the other was the promotion of VEGF release by the ASM cells, and amphiregulin can promote human ASM cell proliferation.

Expression of amphiregulin in human endometrium during the menstrual cycle / 北京大学学报(医学版)

Objective:To investigate the expression of amphiregulin in human endometrium during the menstrual cycle.Methods: Endometrial tissues were collected from the patients undergoing hysterectomy or endometrial biopsy.Real-time RT-PCR,in situ hybridization and immunohistochemistry were used to detect the expression characteristics of amphiregulin in human endometrium in proliferative and secretory phases.Results: Real-time RT-PCR showed the expression of amphiregulin mRNA in secretory phase was 32 times that in proliferative phase.The results from in situ hybridization and immunohistochemistry showed that amphiregulin was located in the cytoplasm and mainly expressed in the gland of endometrium.The expressions of amphiregulin mRNA in proliferative and secretory phases were 0.54?0.22 and 2.96?0.47(P