中期因子和微血管密度在乳腺癌中的表达

目的　探讨中期因子 (MK)与微血管密度 (MVD)的表达与乳腺癌生物学行为的关系及对预后的意义。方法　采用免疫组织化学方法检测 67例乳腺浸润性导管癌组织MK、MVD表达水平 ,并对其与乳腺癌临床病理特征的关系进行统计学分析。结果　本组 67例中 ,MK阳性表达者 3 7例 ,占 5 5 .2 %。MK表达水平及MVD值均与肿块大小 ,腋淋巴结状态及复发和转移有关。MK表达阳性者的MVD均值 (90 .75± 3 3 .0 5 )明显高于MK表达阴性组 (70 .48± 3 1.3 3 ,P <0 .0 1)。结论　MK在乳腺癌血管生成中发挥重要作用 ,MK的阳性表达与乳腺癌的发生、发展和预后密切相关 ,可作为一种有价值的肿瘤标志和预后指标。     

全结肠系膜切除术联合术后乌司他丁治疗右半结肠癌的效果及对血清中期因子、肿瘤特异性生长因子水平的影响

目的:探讨全结肠系膜切除术联合术后乌司他丁治疗右半结肠癌的效果及对血清中期因子(MK)、肿瘤特异性生长因子(TSGF)水平的影响。方法:选取2015年2月至2018年2月上海交通大学医学院附属新华医院崇明分院收治的右半结肠癌根治术患者120例作为研究对象,按照匹配原则分为对照组和研究组,每组60例。两组患者均进行全结肠系膜切除术,在此基础上,研究组患者术后给予乌司他丁静脉滴注治疗。观察两组患者的治疗效果,治疗前后主要肝肾功能指标[丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转移酶(AST)、总胆红素(TBIL)、肌酐及尿素氮(BUN)]、MK、TSGF、炎症因子[肿瘤坏死因子α(TNF-α)、白细胞介素6(IL-6)及白细胞介素8(IL-8)]及免疫功能指标水平和主要并发症发生情况。结果:研究组患者术后进食流质时间、恢复正常饮食时间和住院时间明显短于对照组,差异均有统计学意义(P<0.05)。术后1、3 d,两组患者ALT、AST、TBIL、肌酐和BUN水平均较治疗前明显升高,但研究组患者明显低于对照组,差异均有统计学意义(P<0.05)。治疗后,两组患者血清MK、TSGF、TNF-α、IL-6、IL-8、CD4+及CD8+水平较治疗前明显降低,且研究组患者明显低于对照组,差异均有统计学意义(P<0.05)。研究组、对照组患者的并发症发生率分别为26.67%(16/60)、21.67%(13/60),差异无统计学意义(P>0.05)。结论:全结肠系膜切除术联合术后乌司他丁治疗右半结肠癌的疗效显著,能明显缩短患者的住院时间,改善患者预后,且对术后患者肝肾功能具有一定的保护作用,可显著降低肿瘤相关标志物水平,同时安全性较高。     

Inhibitory effect of midkine-binding peptide on tumor proliferation and migration

Background: To investigate the inhibitory effect of midkine-binding peptides on human umbilical vein endothelial cells (HUVECs) proliferation and angiogenesis of xenograft tumor. Methods: The midkine-binding peptides were panned by Ph.D.-7^™ Phage Display Peptide Library Kit, and the specific binding activities of positive clones to target protein were examined by phage ELISA. The effect of midkine-binding peptides on proliferation of HUVECs was confirmed by MTT test. The xenograft tumor model was formed in BALB/c mice with the murine hepatocarcinoma cells H22 (H22). Microvessel density (MVD) was analyzed by immunohistochemistry of factor VIII staining. Results: Midkine-binding peptides have the inhibitory effects on tumor angiogenesis, a proliferation assay using human umbilical vein endothelial cells (HUVECs) indicated that particular midkine-binding peptides significantly inhibited the proliferation of the HUVECs. Midkine-binding peptides were also observed to efficiently suppress angiogenesis induced by murine hepatocarcinoma H22 cells in BALB/c nude mice. Conclusion: The midkine-binding peptides can inhibit solid tumor growth by retarding the formation of new blood vessels. The results indicate that midkine-binding peptides may represent potent anti-angiogenesis agents in vivo.     

Antagonist activity of the antipsychotic drug lithium chloride and the antileukemic drug imatinib mesylate during glioblastoma treatment in vitro

Objectives: Glioblastoma (GBM), the most common primary tumour of the central nervous system, is characterised by a high malignancy and poor prognosis. The aims of this study were to investigate whether the combination of imatinib mesylate (IM) and lithium chloride (LiCl) exhibited a synergistic effect in treatment and to determine whether midkine (MK) affected the fate of this treatment in vitro.

Methods: Monolayer and spheroid cultures of the T98G human GBM cell line were treated with an IM and LiCl combination for 72 h. The cell proliferation index, apoptotic index, cell cycle distribution, apoptotic and anti-apoptotic protein levels, and cAMP level as well as the cellular morphology and ultrastructure were evaluated.

Results: All applications inhibited cell proliferation and induced apoptosis. The most substantial decreases in cell proliferation and the caspase-3, epidermal growth factor receptor (EGFR), platelet derived growth factor receptor-alpha (PDGFR-α), multidrug resistance protein-1 (MRP-1), aquaporin-4 (AQP-4) and cAMP levels were induced by the LiCl treatment, which exhibited more pronounced effects compared with the combination treatment. LiCl was less effective in decreasing the MK and B cell lymphoma-2 (Bcl-2) levels compared with the combination treatment. The most substantial decrease in the p170 levels was identified following the combination treatment, whereas IM induced the second greatest decrease. LiCl alone had no effect on the p170 levels. IM induced the most substantial decrease in the phospho-glycogen synthase kinase 3-beta (p-GSK-3β)/glycogen synthase kinase 3-beta (GSK-3β) ratio, and LiCl induced the second most substantial decrease. Both LiCl and the combination treatment induced G2 + M arrest, whereas IM induced G0 + G1 arrest after 72 h of exposure. An apoptotic appearance and autophagic vacuoles were commonly identified in the LiCl, combination and IM groups, respectively.

Conclusions: The combination of IM and LiCl exhibited an antagonist effect, and MK had a role at this antagonism.     

健胃愈疡颗粒对胃溃疡大鼠胃组织MK、和表皮生长因子受体表达的影响

目的：探讨健胃愈疡颗粒的疗效及抗复发机制。方法：以Okabe改良法复制大鼠实验性胃溃疡,采用免疫组织化学和原位杂交技术,观察该药对胃溃疡组织Midkine(MK)和表皮生长因子受体（EGFR）表达的影响。结果：模型组MK和EGFR的表达较正常组增高,而健胃愈疡治疗组MK和EGFR的表达增高较模型组更为明显。结论：健胃愈疡颗粒可增加MK和EGFR的上增性表达,这可能是其促进溃疡愈合和提高溃疡愈合质量从而减少溃疡复发的物质基础。     

中期因子对大鼠心肌梗死后心室重塑的保护作用

目的观察中期因子（MK）对大鼠急性心肌梗死（AMI）后心室重塑的影响。方法成年雄性Wistar大鼠48只,随机分为4组：空白对照组（Control组）、伪手术组（sham组）、梗死模型组（AMI组）和MK干预组（MK组）。结扎大鼠左冠状动脉前降支（LAD）,建立AMI动物模型。模型制备成功后,在环绕LAD周围注射MK为MK干预组。4周后,对大鼠行血流动力学指标检测,称重,计算全心体重指数;取组织制备标本,Masson染色鉴定胶原生成情况;免疫组化检测心肌微血管;TUNEL检测细胞凋亡数;Westernblot分析左心室内Bel-2蛋白、P—ERK1蛋白含量的表达。结果MK组大鼠心功能较AMI组得到明显的改善（P〈0．05）;全心体重指数较AMI组明显降低（3．788±0．630比4．725±0．610,P〈0．05）。sham组与Control组几乎无胶原形成,而AMI组与MK组有明显的胶原组织。在梗死及梗死周边区微血管数目MK组多于AMI组（30．662±1．794比15．275±0．389,P〈0．05）,心肌细胞凋亡率明显低于AMI组（27．2±3．2比47．8±4．5,P〈0．05）。MK组Bcl-2蛋白表达较AMI组增多（1．8748±1．0406比1．3637±0．2528,P〈0．05）,P—ERK1蛋白表达较AMI组增多（1．2751±0．6353比0．7862±0．5470,P〈0．05）。结论MK对大鼠心梗后心室重塑产生保护作用,其机制可能与MK上调了P—ERK1蛋白表达有关。     

肝素结合生长因子Midkine基因在大肠杆菌中的克隆、表达和纯化

目的：用基因工程的方法构建、表达和纯化人肝素结合生长因子human Midkine（hMK），并进行活性测定。方法：利用RT-PCR技术从胎儿肾组织中扩增MK，将去除信号肽序列的hMK插入pET30a，构建成表达载体pET-hMK，经表达和亲和层析、纯化获得目的蛋白，3^H-TdR法测定活性。结果：hMK序列与Genebank发表的人MK基因编码序列一致，SDS-PAGE电泳显示表达出目的蛋白，3^H-TdR法测定有良好的活性。结论：人MK已被转人表达载体中，并在大肠杆菌中诱导表达，获得了hMK的表达菌株。     

Alteration of midkine expression in the ischemic brain of humans

Midkine (MK) is a heparin-binding growth factor that occurs as a product of the retinoic acid-inducible gene. Alteration of MK expression in ischemic brain lesions was examined in humans immunohistochemically in nine patients and in two control subjects without neurological disorders. Some neurons were MK-immunopositive, but no evident MK-immunoreactivity was observed in astrocytes in brains of control subjects. In the ischemic lesions, significant elevation of MK-immunoreactivity in the astrocytes and depletion of the reactivity in neurons were seen, especially in the early period, where edema and eosinophilic neurons were prominent. On the other hand, MK-immunoreactivity was not observed in hypertrophic and fibrillary astrocytes in the later period. These findings suggest that the MK in astrocytes play some role in the repair process in the early period of the ischemic brain lesions in humans.     

Intraventricular administration of the neurotrophic factor midkine ameliorates hippocampal delayed neuronal death following transient forebrain ischemia in gerbils

Midkine (MK) is a growth factor with neurotrophic activities, and is expressed during the early stages of experimental cerebral infarction in rats in the zone surrounding the infarct. To evaluate in vivo activity of MK in preventing neuronal death, MK produced in yeast (Pichia pastoris) was administered into the brain ventricle immediately before occlusion of the bilateral common carotid artery of Mongolian gerbils. MK administration at the dose of 0.5-2 microg immediately before occlusion was found to ameliorate delayed neuronal death in the hippocampal CA1 region caused by transient ischemia 7 days after the insult. The hippocampal neurons of the MK-administered gerbils tended to degenerate 14 and 21 days after the insult, but their numbers remained higher than those in saline-administered controls; however, the hippocampal neurons were degenerated 28 days after the insult. MK administration at 2 h after occlusion did not ameliorate the neuronal death. These findings suggested that the therapeutic time window was narrow. The two to four times repeated administration of 2 microg MK immediately before and at 1, 2, or 3 weeks after the occlusion were not significantly different for the hippocampal neuronal death at 28 days after the insult compared with a single injection, but were significantly effective compared with vehicle administration alone. These findings suggested that the therapeutic time window was relatively narrow. The potent neuroprotective activity of MK observed in vivo suggested that MK might be useful as a therapeutic reagent for prevention of neuronal death in neurodegenerative diseases.     

Absence of mouse pleiotrophin does not affect bone formation in vivo

Pleiotrophin (Ptn) is an extracellular matrix protein that regulates hippocampal synaptic plasticity and learning behavior in vivo. Since the overexpression of Ptn in transgenic mice leads to increased bone formation, we analyzed whether a deficiency in Ptn expression would have a negative effect on bone remodeling. Bones from Ptn-deficient mice and wild-type littermates were analyzed using radiography, μCT imaging and undecalcified histology. Biomechanical stability was determined in a three-point-bending assay. Cellular activities were assessed using dynamic histomorphometry and the determination of urinary collagen degradation products. Skeletons of Ptn-deficient mice have no gross abnormalities, displayed a normal size, and showed no differences in growth plate organization compared to wild-type littermates. There were no obvious differences in bone mass as determined by radiographic and μCT imaging. The absence of a bone remodeling phenotype in Ptn-deficient mice was further confirmed using static histomorphometry and biomechanical testing. Finally, the number, morphology, and function of osteoclasts, osteoblasts, and osteocytes were not altered in Ptn-deficient mice compared to wild-type littermates. The complete skeletal analysis of Ptn-deficient mice presented here demonstrates that the lack of Ptn in mice does not affect bone formation in vivo. Therefore, Ptn does not play a significant role in normal bone physiology.