Non-invasively differentiate non-alcoholic steatohepatitis by visualizing hepatic integrinαvβ3 expression with a targeted molecular imaging modality

BACKGROUND Non-invasive methods to diagnose non-alcoholic steatohepatitis(NASH),an inflammatory subtype of non-alcoholic fatty liver disease(NAFLD),are currently unavailable.AIM To develop an integrinαvβ3-targeted molecular imaging modality to differentiate NASH.METHODS Integrinαvβ3 expression was assessed in Human LO2 hepatocytes Scultured with palmitic and oleic acids(FFA).Hepatic integrinαvβ3 expression was analyzed in rabbits fed a high-fat diet(HFD)and in rats fed a high-fat,high-carbohydrate diet(HFCD).After synthesis,cyclic arginine-glycine-aspartic acid peptide(cRGD)was labeled with gadolinium(Gd)and used as a contrast agent in magnetic resonance imaging(MRI)performed on mice fed with HFCD.RESULTS Integrinαvβ3 was markedly expressed on FFA-cultured hepatocytes,unlike the control hepatocytes.Hepatic integrinαvβ3 expression significantly increased in both HFD-fed rabbits and HFCD-fed rats as simple fatty liver(FL)progressed to steatohepatitis.The distribution of integrinαvβ3 in the liver of NASH cases largely overlapped with albumin-positive staining areas.In comparison to mice with simple FL,the relative liver MRI-T1 signal value at 60 minutes post-injection of Gd-labeled cRGD was significantly increased in mice with steatohepatitis(P<0.05),showing a positive correlation with the NAFLD activity score(r=0.945;P<0.01).Hepatic integrinαvβ3 expression was significantly upregulated during NASH development,with hepatocytes being the primary cells expressing integrinαvβ3.CONCLUSION After using Gd-labeled cRGD as a tracer,NASH was successfully distinguished by visualizing hepatic integrinαvβ3 expression with MRI.

CXCR4通过integrinαVβ3调控头颈部鳞癌细胞迁移的机制研究

目的:研究C-X-C型趋化因子受体4(CXCR4)-整合素αVβ3(integrinαVβ3)介导头颈部鳞癌细胞迁移的分子生物学机制。方法:应用头颈部鳞癌细胞株HN-5,采用慢病毒转染方法构建CXCR4过表达的细胞株,用荧光显微镜观察基因表达情况;采用脂质体法将CXCR4-siRNA、integrinαVβ3-siRNA分别转染至HN-5细胞和CXCR4过表达的HN-5细胞中,Transwell小室实验检测HN-5细胞迁移能力,Western blot实验检测CXCR4、integrinαVβ3、基质金属蛋白酶-9(MMP-9)、基质金属蛋白酶-2(MMP-2)和黏附斑激酶(FAK)-非受体型酪氨酸蛋白激酶(SRC)-细胞外信号调节激酶(ERK)信号通路相关蛋白的表达情况。结果:CXCR4过表达可上调HN-5细胞中integrinαVβ3表达水平,而抑制CXCR4表达可下调integrinαVβ3表达;CXCR4过表达可调控MMP-2和MMP-9促进癌细胞迁移,并激活FAK-SRC-ERK信号通路;而抑制integrinαVβ3表达可明显逆转CXCR4过表达对HN-5细胞的上述作用。结论:CXCR4过表达可通过integrinαVβ3促进头颈部鳞癌细胞迁移,其作用机制可能与激活FAK-SRC-ERK通路有关。

Ischemic preconditioning partially suppresses and postpones integrin α_Vβ_3 mRNA expression following transient global cerebral ischemia in C57BL/6 mice

Previous studies of integrin αvβ3 have focused on ischemic brain damage, although the role of integrin αvβ3 in ischemic preconditioning （IP） has rarely been reported. The present study analyzed the effects of IP on integrin αvβ3 mRNA expression following cerebral ischemia through the use of hematoxylin-eosin staining and real-time quantitative polymerase chain reaction techniques. Integrin avid3 mRNA expression in the ischemia group peaked at 24 hours after ischemia-reperfusion. In the IP ＋ ischemia group, integrin αvβ3 mRNA expression increased after 24 hours, but remained significantly less than the ischemia group, and expression continued to increase until 7 days after ischemiaJreperfusion. These results demonstrate that IP effectively attenuated upregulation of integrin αvβ3 mRNA expression at 24 hours after ischemia.

肿瘤整合素α_vβ_3受体显像的研究现状

整合素αvβ3受体在多种恶性肿瘤细胞表面有高水平的表达,尤其在恶性肿瘤组织新生血管内皮细胞膜,成熟血管内皮细胞和绝大多数正常器官系统则无表达或几乎不能被探及。含有RGD(精氨酸-甘氨酸-天冬氨酸)序列的小分子多肽是整合素αvβ3受体拮抗剂,对整合素αvβ3受体具有高度的选择性与亲和力,是一类具有潜在临床应用价值的肿瘤受体显像剂。

粘附分子CD44V6及Integrinαvβ_3在胆囊癌中的表达及意义

目的研究粘附分子CD44V6及Integrinαvβ_3在胆囊癌组织及胆囊炎中表达是否存在统计学意义的差异,其表达量与胆囊癌的分化程度之间的关系。方法应用免疫组织化学的方法检测20例胆囊癌及20例慢性胆囊炎组织中CD44V6及Integrinαvβ_3的表达,结合临床病理结果进行分析。结果在胆囊癌中CD44V6和Integrinαvβ_3表达阳性率分别为80.0%和70.0%。CD44V6及Integrinαvβ_3在胆囊癌中表达存在协同性,与胆囊癌分化程度呈正相关(均P<0.05)。结论 CD44V6及Integrinαvβ_3在胆囊癌中高表达(显著高于对照组),并且随着胆囊癌的分化程度降低其表达阳性分值增高;CD44V6及Integrinαvβ_3的表达与原发性胆囊癌的良恶性及分化程度相关;在相同标本中,CD44V6高表达往往伴有Integrinαvβ_3高表达,在胆囊癌的诊断上具有一致性,可联合作为临床监测胆囊癌发生、转移的参考指标。

Implication of Expression of Osteopontin and Its Receptor Integrin αvβ3 in the Placenta in the Development of Preeclampsia

To investigate the expression of osteopontin （OPN） and its receptor integrin αvβ3 in the placental tissue from pregnant women complicated with preeclampsia, the expression of OPN and αvβ3 in the placenta of the pregnant women with preeclampsia and healthy pregnant women was detected by immunohistochemistry, Western blotting and RT-PCR. Our results showed that OPN and αvβ3 protein were expressed in the placenta from normal pregnant woman and those with preeclampsia. OPN was located in the placental syncytiotrophoblasts and the cytoplasm of capillary endothelial cells and integrin αvβ3 was mainly expressed on the surface of trophoblast cells. Expression of OPN and integrin αvβ3 in the placental tissue from preeclampsia subjects was significantly lower than that from the control group （P〈0.05）. Compared with the control group, expression of OPN in the placental tissue from preeclampsia group was significantly lower （P〈0.05） but there was no significant difference in the expression of αv and β3 between the preeclampsia group and the controls. It is concluded that OPN and its receptor integrin αvβ3 may be involved in the pathogenesis of preeclampsia.

大肠癌中αvβ3 integrin表达与血管生成拟态的关系及分子机制的研究

目的:探讨大肠癌中是否存在血管生成拟态(VM),阐述VM存在的临床意义;分析VM与αvβ3 integrin表达的关系;研究大肠癌中αvβ3 integrin表达的临床意义及其与FAK、MMP-2、VEGF表达的相关性。方法:对227例大肠癌组织切片进行CD31免疫组织化学和PAS染色及αvβ3 integrin、FAK、VEGF、MMP-2免疫组织化学染色。结果:VM在大肠癌中阳性率为18.06%(41/227),αvβ3 integrin在大肠癌中阳性率为56.83%(129/227);VM阳性组、αvβ3 integrin阳性组生存时间均较阴性组短,差异均有统计学意义(P<0.05);VM阳性组的αvβ3 integrin阳性率较阴性组高,差异有统计学意义(P<0.05);αvβ3 integrin与FAK、VEGF、MMP-2表达呈正相关。结论:大肠癌中VM的存在、αvβ3 integrin的表达与肿瘤的不良预后有关;αvβ3 integrin可能通过与FAK、VEGF、MMP-2的相互作用,参与了大肠癌VM的形成。

Effect of lentivirus-mediated integrin αVβ3-sh RNA on tumor growth of mice with lung cancer xenografts

Objective: To study the effect of lentivirus-mediated integrin αVβ3-sh RNA on tumor growth of mice with lung cancer xenograft. Methods: Lung cancer tissue, paracancer tissue and normal tissue were collected and integrin αVβ3 expression was detected; BALB/c nude mice were selected, divided into integrin αVβ3 knockdown group(KD group) and negative control group(NC group), and inoculated with cells stably infected by integrin αVβ3-sh RNA lentivirus and cells stably infected by negative control-sh RNA lentivirus respectively, the growth of tumor tissue was continuously observed, and the number of apoptosis cells as well as the expression of angiogenesis, apoptosis and invasion genes in tumor tissue were detected. Results: m RNA content and protein content of integrin αVβ3 in lung cancer tissue were significantly higher than those in paracancer tissue and normal tissue; increasing trend of tumor tissue volume of KD group was weaker than that of NC group, and tumor volume at various points in time of KD group was lower than that of NC group; m RNA contents and protein contents of VEGF, FGF, EGF, Bcl-2, MMP-9, MMP-12 and MMP-13 in tumor tissue of KD group were lower than those of NC group, and apoptosis index as well as m RNA content and protein content of Bax were higher than those of NC group. Conclusions: The expression of integrin αVβ3 increases in lung cancer tissue, and lentivirus-mediated integrin αVβ3-sh RNA can inhibit tumor growth of mice with lung cancer xenografts.

Effects of Integrins and Integrin αvβ3 Inhibitor on Angiogenesis in Cerebral Ischemic Stroke

Summary： Integrins such as αvβ3, α5β31 play a key role in angiogenesis regulation, invasion and metastasis, inflammation, wound healing, etc. The up-regulation of integrin αvβ3 after cerebral ischemic stroke can promote angiogenesis, which in turn improves functional recovery. In addition, the integrin αvβ3 inhibitor can block the blood-brain barrier （BBB） leakage induced by vascular endothelial growth factor （VEGF） and also can reduce inflammatory reaction, decrease the deposition of fibrinogen. Other studies showed that integrin αvβ3 is not essential in revascularization. Therefore, the effect of integrin αvβ3 in the whole process of brain function recovery merits further study.

Milk fat globule epidermal growth factor 8 alleviates liver injury in severe acute pancreatitis by restoring autophagy flux and inhibiting ferroptosis in hepatocytes

BACKGROUND Liver injury is common in severe acute pancreatitis(SAP).Excessive autophagy often leads to an imbalance of homeostasis in hepatocytes,which induces lipid peroxidation and mitochondrial iron deposition and ultimately leads to ferroptosis.Our previous study found that milk fat globule epidermal growth factor 8(MFG-E8)alleviates acinar cell damage during SAP via binding toαvβ3/5 integrins.MFG-E8 also seems to mitigate pancreatic fibrosis via inhibiting chaperone-mediated autophagy.AIM To speculate whether MFG-E8 could also alleviate SAP induced liver injury by restoring the abnormal autophagy flux.METHODS SAP was induced in mice by 2 hly intraperitoneal injections of 4.0 g/kg L-arginine or 7 hly injections of 50μg/kg cerulein plus lipopolysaccharide.mfge8-knockout mice were used to study the effect of MFG-E8 deficiency on SAPinduced liver injury.Cilengitide,a specificαvβ3/5 integrin inhibitor,was used to investigate the possible mechanism of MFG-E8.RESULTS The results showed that MFG-E8 deficiency aggravated SAP-induced liver injury in mice,enhanced autophagy flux in hepatocyte,and worsened the degree of ferroptosis.Exogenous MFG-E8 reduced SAP-induced liver injury in a dose-dependent manner.Mechanistically,MFG-E8 mitigated excessive autophagy and inhibited ferroptosis in liver cells.Cilengitide abolished MFG-E8’s beneficial effects in SAP-induced liver injury.CONCLUSION MFG-E8 acts as an endogenous protective mediator in SAP-induced liver injury.MFG-E8 alleviates the excessive autophagy and inhibits ferroptosis in hepatocytes by binding to integrinαVβ3/5.

Ouabain inhibits anchorage-independent growth in human lung cancer cells via integrinαvβ3 reduction

Physiological effects of ouabain (Fig. 1A), a human endogenous hormone, have been intensively investigated nowadays.Recent studies demonstrate anticancer activity of ouabain in sensitization of apoptosis and suppression of migration in lung cancer cells (1,2)Focusing on metastatic process, the ability of cancer cells to grow in an anchorage-independent condition determines the success of cancer metastasis [3].

ANALYSIS OF T CELL CLONALITY BY CDR3 SIZE OF T-CELL ANTIGEN RECEPTOR Vβ REPERTOIRE IN HCL AND c-ALL

Objective: To analyze the distribution and clonality of TCR Vβ subfamily T cells in hairy cell leukemia (HCL) and common-acute lymphoblastic leukemia (c-ALL). Methods: Peripheral blood mononuclear cell samples from 3 cases of HCL and 1 case of c-ALL were investigated for analysis of complementarity determining region 3 (CDR3) size of T cell receptor Vβ repertoire using reverse transcriptase-polymerase chain reaction (RT-PCR). The products were further analyzed by genescan to identify T cell clonality. Results: Some Vβ subfamily PCR products from 4 patients contained monopeak (monoclone) or a dominant peak (oligoclone). In contrast, multipeak (polyclone) distributions were found in all Vβ subfamily PCR products from normal control cases. Conclusion: T cell clonal expansion may be found in HCL and c-ALL cases that may indicate a host response directed against leukemia related antigen. In addition, it may be useful to detect the minimal residual disease.

Effects of controlled superovulation on the expression of HOXA10,integrin αvβ3 and LIF in mice during peri-implantation

Objective:To observe the expression of endometrial homologous box gene A10(HOXA10),Integrinαvβ3 and leukemia suppressor factor(LIF)in peri-implantation mice,and to investigate the effect of controlled superovulation(COH)on endometrial receptivity in mice.Methods:After COH model preparation and blastocyst transplantation,kunming pure-bred mature mice were randomly divided into natural blastocyst+natural pseudocyst group,COH blastocyst+natural pseudocyst group,natural blastocyst+COH pseudocyst group,and COH blastocyst+COH pseudocyst group.The pregnancy rate and average number of beds in each group were observed on day 6 of conception.Western blot and RT-PCR were used to detect the expression of HOXA10,Integrinααvβ3,LIF protein and mRNA in the endometria of mice on day 5 of conception.Results:The pregnancy rate and average number of beds in the COH group were lower than those in the natural receptor group.The expression of HOXA10,Integrinααvβ3 and LIF in endometrium was significantly lower than that of the natural receptor mice(P<0.01).Conclusion:Superovulation drugs down-regulate the expression of HOXA10,Integrinαvβ3 and LIF in endometrium during the implantation window,reduce endometrial receptivity,and lead to low clinical pregnancy rate.

恩度联合放疗对 Lewis 肺癌小鼠 MVD 及αvβ3 mRNA 表达的影响

目的：观察恩度联合放疗对Lewis肺癌的生长抑制作用，检测肿瘤中微血管密度（ MVD）及αvβ3 mRNA的表达，探讨其在肿瘤血管“正常化”及协同抑瘤作用中的可能机制。方法荷瘤小鼠随机分成4组：空白对照组（NC），恩度组（ES），放射治疗组（RT），恩度联合放疗组（ES＋RT）。从开始治疗后隔日测量肿瘤体积，绘制生长曲线。免疫组化法及RT-PCR分别检测各组肿瘤组织中MVD及αvβ3 mR-NA的表达情况。结果（1）ES＋RT组的抑瘤作用最为明显；（2）与空白对照组相比，RT组随时间延长微血管计数逐渐增加，ES组及ES＋RT组下降，其中ES＋RT组比ES组下降更为明显，差异有统计学意义（P＜0．05）；（3）与空白对照组相比，RT组αvβ3 mRNA表达水平升高，而ES及ES＋RT组则降低，ES＋RT组降低更为明显，ES组在第6天扩增倍数最低，各治疗组在d8～d12均有统计学差异（ P＜0．05）。结论恩度联合放疗显著抑制Lewis肿瘤的生长；恩度能使肿瘤MVD明显减少，形态趋于正常；恩度可下调αvβ3 mRNA的表达，改善肿瘤组织紊乱的血管网，降低乏氧，可能是放疗增敏的机制之一。

放射性标记RGD肽行肿瘤αvβ3受体显像研究的进展

恶性肿瘤的持续生长、侵袭转移与肿瘤血管生成密切相关,在这个过程中整合素αvβ3起着重要的作用。αvβ3主要通过和细胞外基质(ECM)中的一些含RGD的三肽序列配体结合发挥作用,文献报道其在新生血管内皮细胞有强烈表达,而在正常细胞表达极少或缺乏,因此,放射性标记RGD肽作为高特异性的标记物对恶性肿瘤进行核医学显像,能够达到早期诊断和治疗目的。本文就近年来国内外RGD肽的标记方法和αvβ3受体显像研究进行了综述。

EGFR、αVβ3、ADAM23在大肠癌中的表达及临床意义

目的探讨表皮生长因子受体(epidermal growth factor receptor,EGFR)、去整合素-αVβ3(integrin alpha-v-beta3,αVβ3)、去整合素-蛋白酶23(adisintegrin and metalloprotease 23,ADAM23)在结直肠癌(colorectal cancer,CRC)及癌旁正常组织中的表达、相关性及临床意义。方法选取CRC患者癌组织61例作为CRC组,依据肝转移情况分为转移组和无转移组,选取其中CRC患者癌旁正常组织20例作为对照组,采用免疫组化法检测所有组织中EGFR、αVβ3、ADAM23表达情况。结果 CRC组EGFR、αVβ3表达阳性率明显高于对照组,CRC组ADAM23表达阳性率明显低于对照组,差异有统计学意义(P<0.001);肝转移组EGFR、αVβ3表达阳性率明显高于无肝转移组,ADAM23表达阳性率明显低于无肝转移组,差异有统计学意义(P<0.05)。肠癌原发灶中EGFR表达与αVβ3表达无明显相关(P=0.152),ADAM23表达与αVβ3或EGFR表达呈显著负相关,差异有统计学意义(P<0.001)。结论 ADAM23在CRC组织中呈低表达,EGFR、αVβ3呈高表达,ADAM23表达与αVβ3或EGFR表达呈显著负相关。以此为靶点三者与CRC的发生、发展有关,可作为评估CRC病情的重要参考指标,指导药物开发和临床治疗重要方向,值得进一步推广。

靶向整合素αvβ3受体isoDGR-2CY的^(131)I标记与生物活性评价

目的 131I标记αvβ3受体isoDGR-2CY模序,研究其在荷VX2肿瘤裸鼠体内的生物分布及显像。方法 (1)合成isoDGR-2CY模序并用131I标记。(2)20只荷VX2肿瘤裸鼠随机分成4组,每组5只,经尾静脉注射7.4 MBq(0.2 ml)131IisoDGR-2CY,分别于注射后2、5、12、24 h按组将裸鼠处死;另取5只荷瘤裸鼠作为竞争性抑制组,注射131I-isoDGR-2CY前1 h,先注射未标记的isoDGR-2CY,并于注射131I-isoDGR-2CY后12 h处死。各组取血及主要脏器,称取脏器质量并测量放射性计数,经衰减校正后计算每克组织百分注射剂量率(%ID/g)。(3)取6只荷VX2肿瘤裸鼠,实验前7 d在饮用水中加入0.1%碘化钾封闭甲状腺后,经尾静脉注射7.4 MBq的131I-isoDGR-2CY。4只作为显像组,另2只作为竞争性抑制组[注射131I-isoDGR-2CY前1 h,先注射未标记的isoDGR-2CY(isoDGR-2CY与131I-isoDGR-2CY摩尔比为10∶1)],分别于注射后2、4、8、12、24 h进行SPECT静态显像。结果合成的isoDGR-2CY模序纯度为97.06%,可以满足实验要求;131I-isoDGR-2CY标记率为(89.03±0.18)%,放化纯度(95.89±0.12)%,其在血清中具有良好的稳定性;131I-isoDGR-2CY主要通过肾脏排泄,肿瘤的靶向性明显,12 h靶/肌肉比值(T/M)为2.39;SPECT显示注射标记物1 h肿瘤即可见显影,12 h显影清晰,该显像可被未标记的isoDGR-2CY抑制。结论 isoDGR-2CY可被131I成功标记,肿瘤组织能特异性摄取131I-isoDGR-2CY,并通过SPECT清晰显像。

黄体功能不全患者子宫内膜上皮细胞整合素β_3亚单位与孕激素受体关系

目的 探讨黄体功能不全患者子宫接受性与上皮细胞雌、孕激素受体的关系。方法 应用免疫组化方法测定黄体功能不全患者子宫内膜上皮细胞整合素β3亚单位和雌、孕激素受体的表达。结果 黄体功能不全患者子宫内膜上皮细胞整合素 β3亚单位含量显著降低 ,而雌、孕激素受体含量明显升高。结论 黄体功能不全患者子宫接受性下降 ,且与黄体中期雌、孕激素受体失去下调密切相关。

整合素β3与VEGFR-3在胃癌组织中的表达及意义

目的探讨整合素β3、血管内皮细胞生长因子受体-3(VEGFR-3)在胃癌发生、发展中的作用。方法用免疫组织化学SP法检测65份胃癌术后癌组织标本(观察组)中整合素β3、VEGFR-3表达[定量以淋巴管密度(LMVD)表示],并与20份距胃癌组织10 cm的正常胃黏膜组织标本(对照组)作对比。结果观察组整合素β3阳性表达率及LMVD均明显高于对照组(P均<0.01)。观察组整合素β3阳性表达者LMVD明显高于阴性表达者(t=4.30,P<0.01),整合素β3阳性表达与LMVD呈正相关(r=0.473,P<0.01)。结论整合素β3和VEGFR-3与胃癌的淋巴管形成及浸润转移密切相关,可作为胃癌预后综合性评估的有效指标。

整合素αV与β3在鼻咽癌组织中的表达及其意义

目的探讨整合素αV、β3在鼻咽癌组织中的表达及其与鼻咽癌发生发展的关系。方法采用免疫组化Supervision二步法检测106例鼻咽癌组织、26例鼻咽黏膜慢性炎症组织中整合素αV、β3的表达,分析其与鼻咽癌患者各临床病理特征之间的关系。结果 106例鼻咽癌组织中,整合素αV、β3表达阳性率分别为81.1%(86/106)、94.3%(100/106),26例鼻咽黏膜慢性炎症组织均无整合素αV、β3阳性表达(P<0.01)。整合素αV、β3表达与鼻咽癌临床分期、淋巴结转移和远处转移有关(P<0.05),与患者性别、年龄无关(P>0.05)。整合素αV与β3表达无相关性(P>0.05)。结论整合素αV、β3与鼻咽癌的发生、发展有密切关系,两者互相作用共同促进鼻咽癌的侵袭转移。