The expression of neurokinin-1 and preprotachykinin-1 in breast cancer cells depends on the relative degree of invasive and metastatic potential

Breast cancer has a predilection for metastasis to the bone marrow. The preprotachykinin-I (PPT-I) gene has a central role in the early migration of breast cancer cells into the bone marrow, making this organ a latent repository of the cancer cells. This study investigated whether the invasive and metastatic potential of breast cancer cells correlate with the expression of the PPT-I gene and the receptors for its peptides, neurokinin-1 (NK-1) and NK-2. The studies compared cells that are non-tumorigenic (MCF12A), low metastatic and invasive potential (MCF7), and sublines of MCF with increased invasive and metastatic potential (LCC1 and LCC2). LCC2, but not LCC1 is tamoxifen resistant. Quantitative RT-PCR showed increased expression of PPT-I, NK-1 and NK-2 mRNA LCC1 and LCC2. MCF7 required stimulation by phorbol ester for NK-1 induction. The levels of NK-2 mRNA were significantly increased in LCC2. Clonogenic assays with specific receptor antagonists showed a predominant role for NK-2 in the proliferation of both LCC1 and LCC2. While the growth rate of LCC1 and LCC2 were similar, the latter showed increased migration. Use of a nude mouse model confirmed higher metastatic potential of LCC2, including increased migration to regions of the endosteum. Overall, these studies show a correlation between three neuroendocrine-related genes: PPT-I, NK-1 and NK-2 and the metastatic potential of specific breast cancer cells. These cells provide a model for future studies on bone marrow metastasis.This work was done at UMDNJ-New Jersey Medical School, Departments of Pathology and Laboratory Medicine and Medicine-Hematology/Oncology.     

急性坏死性胰腺炎肠粘膜损害与NK-IR的过度表达相关

目的:研究神经激肽-1受体(NK-1R)和神经激肽-2受体(NK-2R)在急性坏死性胰腺炎(ANP)大鼠末端回肠组织中的表达,探讨该受体的表达与ANP肠粘膜损害的关系。方法:健康成年Sprague-Dawley大鼠随机分为假手术对照组(50只)和ANP组(80只)。假手术对照组开腹后只翻动胰腺,ANP组大鼠胰胆管恒速逆行注射5%牛磺胆酸钠,制成ANP大鼠模型。应用逆转录聚合酶链反应(RT-PCR)检测末端回肠组织NK-1R和NK-2R的mRNA水平,应用Westernblot检测NK-1R的蛋白表达水平。结果:与假手术对照组相比,ANP末端回肠组织中NK-1R和NK-2R的mRNA水平过度表达。NK-1R的表达水平分别与肠粘膜的病理学评分(r=0.77,P＜0.01)和肠粘膜通透性(r=0.68,P＜0.01)呈明显正相关。结论:ANP时,肠组织中NK-1R和NK-2R的表达水平明显上调,P物质的过度作用加剧ANP时肠粘膜的病理损害,损害肠粘膜屏障的功能。     

SMURF1对雌激素受体α阳性乳腺癌细胞增殖影响的分子机制及其对乳腺癌患者预后的影响

目的探讨SMURF1对雌激素受体α阳性乳腺癌细胞增殖的影响及其分子机制,进一步探讨其表达水平与乳腺癌患者预后的相关性.方法首先采用体外实验探索SMURF1同ERα表达的关系,与此同时收集湖北医药学院附属国药东风医院2012年2月至2018年3月肿瘤科收治的89例乳腺癌病例的临床及病理资料进行回顾性分析.长期对患者进行随访,确定SUMRF1在乳腺癌患者中的表达与预后的相关性.结果体外研究表明敲低SMURF1将减少ERα阳性细胞在体外和体内的增殖,SMURF1敲低显著降低乳腺癌细胞中ERα靶基因的表达.远期随访无复发或转移的乳腺癌患者组织中SMURF1蛋白质、SMURF1 mR-NA水平均低于有复发的患者;无复发或转移的乳腺癌患者组织中ERα蛋白质水平显著低于有复发或转移的乳腺癌患者,两组差异有统计学意义(t=18.361,P<0.01);乳腺癌复发患者组织中SMURF1 mRNA表达水平显著高于未复发患者组织,差异具有显著统计学意义(t=13.512,P<0.001)同时相关性分析表明,SMURF1蛋白质水平的表达同ERα蛋白质水平具有正相关性(r=1.356,P<0.001).结论SUMRF1的表达有可能影响ERα的信号途径,同时与乳腺癌患者的预后有一定的相关性,SUMRF1的表达水平低者的生存期限更长.SMURF1和ERα信号在支持乳腺癌生长方面可能具有调控作用.靶向SMURF1可能是ERα阳性乳腺癌治疗的一种有前途的策略.同时SMURF1未来有可能作为预测患者预后的有效临床指标.     

神经降压素及其受体1与乳腺癌

神经降压素是一种分布于脑和胃肠道的多肽,具有广泛的中枢和外周效应.近来研究表明神经降压素及其1型受体的信号与乳腺癌的增殖、抗凋亡、侵袭转移方面具有密切的联系;而神经降压素受体1仅在乳腺癌中表达将为开发神经降压素类似物为基础的新型肿瘤显像剂及靶向治疗药物提供广泛的应用前景,有助于乳腺癌的早期诊断和治疗.     

5-羟色胺受体和P物质受体在培养的人胎盘滋养层细胞的定位研究

目的：探讨培养的人胎盘绒毛滋养层细胞是否存在5－羟色胺受体（5－HTR）和P物质受体（SPR）,方法：采用细胞培养法和免疫细胞化学技术。结果：培养的滋养层细胞为5－HTR与SPR免疫阳性反应,免疫阳性物质位于胞浆内,胞核为阴性。结论：人胎盘滋养层细胞自身含有5－HTR和SPR,为在离体培养条件下研究5－HT和SP对胎盘激素的合成与分泌的调控作用提供了形态学依据。     

侧脑室注射P物质对大鼠胃电-机械活动的影响

观察大鼠侧脑室注射P物质 (SP)及药物对胃电 机械活动和体表胃电活动 (EGG)的影响 ,应用SP受体拮抗剂和阿托品 (atropine)探讨SP的受体机制 ;比较了冷应激后胃对SP和SP受体拮抗剂的动力学反应性的变化。雄性Wistar大鼠 4 4只 ,随机分为 7组 :正常对照 ;SP ;SP受体拮抗剂 ;atropine ;冷应激对照 ;冷应激 +SP ;冷应激 +SP拮抗剂。结果表明 :(1)侧脑室SP给药 10 μL ,(10 -8mol/L)能引起胃窦运动明显增强 (P <0 0 5 ) ,(2 )应激使胃运动幅度极显著升高、频率加快 (P <0 0 1) ,应激后大鼠对侧脑室注入P物质的胃动力学反应性较前明显增强 (P <0 0 5 )。(3)侧脑室注入SP受体拮抗剂 ([D Pro2 ,D Phe7,D Trp9] SubstanceP) 10 μL ,(10 -8mol/L)在确定SP的神经递质作用中没有明显作用。侧脑室注射M受体阻断剂atropine10 μL ,(1× 10 -7mol/L)不能阻断脑内P物质对胃的收缩效应 ,结果提示 ,中枢注入的P物质可以促进胃电 机械活动的调节因子参与中枢增强胃肠的调控 ;SP在中枢可能是通过第二信使而发挥兴奋性递质作用的     

Substance P and the regulation of inflammation in infections and inflammatory bowel disease

Substance P (SP) and its natural analogue hemokinin‐1 (HK1) are produced by lymphocytes and macrophages, and at times B cells. These peptides are an important component of the immune response during several infections and in inflammatory bowel disease (IBD). The synthesis of SP and HK1 in leucocytes is subject to immune regulation. IL12 and IL23 stimulate T cells and macrophages to make SP respectively. The cytokines driving HK1 production are not presently defined. These peptides act through a shared receptor called neurokinin‐1. T cells, macrophages and probably other immune cell types can express this receptor. Several cytokines IL12, IL18 and TNFα as well as T‐cell antigen receptor activation induce neurokinin‐1 receptor expression on T cells, while IL10 blocks receptor display. TGFβ delays internalization of the SP/neurokine‐1R complex on T cells resulting in stronger receptor signalling. One of the functions of SP and neurokinin‐1 receptor is to enhance T cell IFNγ and IL17 production, amplifying the proinflammatory response. Thus, SP and HK1 have overlapping functions and are part of a sophisticated immune regulatory circuit aimed at amplifying inflammation at mucosal surfaces and in other regions of the body as shown in animal models of infection and IBD.     

大鼠中央杏仁核内微量注射P物质抑制对大鼠胃电-机械活动

目的观察大鼠中央杏仁核内微量注射P物质（SP）对胃肌电和胃运动幅度的影响。方法用铂金丝双电极和应变片传感器引导胃肌电及胃运动信号,用MacLab系统记录并处理信号。结果中央杏仁核内注射4g／L的SP 1μL,可明显抑制胃电．机械活动;注射1．5g／L的SP受体拮抗剂（DPDPDT）1μL,可阻断内源性SP的作用,胃电-机械活动增强;并且DPDPDT可使外源性SP对胃电-机械活动的抑制效应减弱;用0．5g／L阿托品1μL阻断M受体后,SP对胃电-机械活动的抑制效应也减弱。结论SP在中央杏仁核可能通过与SP受体结合,抑制胃电-机械活动,而胆碱能神经元可能参与了SP抑制胃电-机械活动这一过程。     

大鼠脊髓白质后索内P物质受体阳性神经元的形态特征及其联系

本研究应用免疫组织化学方法和免疫荧光组织化学双标技术,观察了大鼠脊髓白质后索内的P物质(SP)受体(SPR)阳性神经元的形态特征及其联系。结果表明,脊髓白质后索内存在SPR阳性神经元,它们的胞体较小,常集中在两侧后索的中线上,呈三角形、圆形和多极形;它们的短树突在胞体周围呈放射状,但向后索表面行走的树突较直,且可达脊髓表面;在激光共聚焦显微镜下可见这些SPR阳性神经元呈NeuN阳性,但GFAP呈阴性;它们的胞体及其突起与SP、谷氨酸脱羧酶(GAD)、脑啡肽(ENK)和5-HT阳性纤维及终末形成紧密接触。上述结果说明脊髓白质后索内存在神经元,且呈SPR阳性;这些SPR阳性神经元的活动可能受到多种来源神经信号的调控。     

Fundamental Role of microRNAs in Androgen‐Dependent Male Reproductive Biology and Prostate Cancerogenesis

Male reproductive failure has been linked to successive development of various urologic diseases including prostate cancer. There is strong epidemiologic data in support of this association, it is important therefore to identify the fundamental grounds that lay beneath such a connection. Male reproductive biology, as sex determined, is significantly dependent upon the hormonal regulation of androgens. With the advancement of knowledge on androgen receptivity and epigenetic regulation, the role of new regulatory factors such as microRNAs becomes essential. This review focuses on unraveling the role of microRNA tight incorporation in androgen‐dependent male reproductive biology in the context of recent prostate cancer data.     

Neurokinin-1 Receptor Immunoreactive Neuronal Elements in the Superficial Dorsal Horn of the Chicken Spinal Cord: With Special Reference to Their Relationship with the Tachykinin-containing Central Axon Terminals in Synaptic Glomeruli

Synaptic glomeruli that involve tachykinin-containing primary afferent central terminals are numerous in lamina II of the chicken spinal cord. Therefore, a certain amount of noxious information is likely to be modulated in these structures in chickens. In this study, we used immunohistochemistry with confocal and electron microscopy to investigate whether neurokinin-1 receptor (NK-1R)-expressing neuronal elements are in contact with the central primary afferent terminals in synaptic glomeruli of the chicken spinal cord. We also investigated which neuronal elements (axon terminals, dendrites, cell bodies) and which neurons in the spinal cord possess NK-1R, and are possibly influenced by tachykinin in the glomeruli. By confocal microscopy, NK-1R immunoreactivities were seen in a variety of neuronal cell bodies, their dendrites and smaller fibers of unknown origin. Some of the NK-1R immunoreactive profiles also expressed GABA immunoreactivities. A close association was observed between the NK-1R-immunoreactive neurons and tachykinin-immunoreactive axonal varicosities. By electron microscopy, NK-1R immunoreactivity was seen in cell bodies, conventional dendrites and vesicle-containing dendrites in laminae I and II. Among these elements, dendrites and vesicle-containing dendrites made contact with tachykinin-containing central terminals in the synaptic glomeruli. These results indicate that tachykinin-containing central terminals in the chicken spinal cord can modulate second-order neuronal elements in the synaptic glomeruli.     

Common genetic variants in pre-microRNAs were associated with increased risk of breast cancer in Chinese women

Small, noncoding RNA molecules, called microRNAs (miRNAs), are thought to function as either tumor suppressors or oncogenes. Common single-nucleotide polymorphisms (SNPs) in miRNAs may change their property through altering miRNA expression and/or maturation, and thus they may have an effect on thousands of target mRNAs, resulting in diverse functional consequences. However, it remains largely unknown whether miRNA SNPs may alter cancer susceptibility. We evaluated the associations of selected four SNPs (rs2910164, rs2292832, rs11614913, and rs3746444) in pre-miRNAs (hsa-mir-146a, hsa-mir-149, hsa-mir-196a2, and hsa-mir-499) with breast cancer risk in a case-control study of 1,009 breast cancer cases and 1,093 cancer-free controls in a population of Chinese women and we found that hsa-mir-196a2 rs11614913:T>C and hsa-mir-499 rs3746444:A>G variant genotypes were associated with significantly increased risks of breast cancer (odds ratio [OR], 1.23; 95% confidence interval [CI], 1.02-1.48 for rs11614913:T>C; and OR, 1.25; 95% CI, 1.02-1.51 for rs3746444:A>G in a dominant genetic model) in a dose-effect manner (P for trend was 0.010 and 0.037, respectively). These findings suggest, for the first time, that common SNPs in miRNAs may contribute to breast cancer susceptibility. Further functional characterization of miRNA SNPs and their influences on target mRNAs may provide underlying mechanisms for the observed associations and disease etiology.     

GABA(A) receptor facilitation of neurokinin release from primary afferent terminals in the rat spinal cord

Our goal was to test the following hypotheses: 1) GABA_A receptors facilitate neurokinin release from primary afferent terminals; 2) they do this by suppressing an inhibitory effect of GABA_B receptors; 3) the activation of these two receptors is controlled by the firing frequency of primary afferents. We evoked neurokinin release by stimulating the dorsal root attached to spinal cord slices, and measured it using neurokinin 1 receptor (NK1R) internalization. Internalization evoked by root stimulation at 1 Hz (but not at 100 Hz) was increased by the GABA_A receptor agonists muscimol (effective concentration of drug for 50% of the increase [EC₅₀] 3 μM) and isoguvacine (EC₅₀ 4.5 μM). Internalization evoked by root stimulation at 100 Hz was inhibited by the GABA_A receptor antagonists bicuculline (effective concentration of drug for 50% of the inhibition [IC₅₀] 2 μM) and picrotoxin (IC₅₀ 243 nM). Internalization evoked by incubating the root with capsaicin (to selectively recruit nociceptive fibers) was increased by isoguvacine and abolished by picrotoxin. Therefore, GABA_A receptors facilitate neurokinin release. Isoguvacine-facilitated neurokinin release was inhibited by picrotoxin, low Cl⁻, low Ca²⁺, Ca²⁺ channel blockers and N-methyl-d-aspartate receptor antagonists. Bumetanide, an inhibitor of the Na⁺-K⁺-2Cl⁻ cotransporter, inhibited isoguvacine-facilitated neurokinin release, but this could be attributed to a direct inhibition of GABA_A receptors. The GABA_B agonist baclofen inhibited NK1R internalization evoked by 100 Hz root stimulation (IC₅₀ 1.5 μM), whereas the GABA_B receptor antagonist (2S)-3-[[(1S)-1-(3,4-dichlorophenyl)ethyl]amino-2-hydroxypropyl](phenylmethyl) phosphinic acid (CGP-55845) increased NK1R internalization evoked by 1 Hz root stimulation (EC₅₀ 21 nM). Importantly, baclofen inhibited isoguvacine-facilitated neurokinin release, and CGP-55845 reversed the inhibition of neurokinin release by bicuculline. In conclusion, 1) GABA_B receptors located presynaptically in primary afferent terminals inhibit neurokinin release; 2) GABA_A receptors located in GABAergic interneurons facilitate neurokinin release by suppressing GABA release onto these GABA_B receptors; 3) high frequency firing of C-fibers stimulates neurokinin release by activating GABA_A receptors and inhibiting GABA_B receptors, whereas low frequency firing inhibits neurokinin release by the converse mechanisms.     

Amplified in breast cancer 1 expression in breast cancer

Aims: The amplified in breast cancer 1 (AIB1), steroid receptor co‐activator family member, acts as an oestrogen receptor (ER) co‐activator. Acting with HER‐2, it is thought to play a role in endocrine resistance by facilitating ER–growth factor crosstalk. The aim was to analyse AIB1 expression by immunohistochemistry and study its correlations with other prognostic variables in breast cancer and its effect on survival. Methods: A tissue microarray comprising tumours from 438 patients with 15.4 years’ median follow‐up was used. Interpretable AIB1 expression obtained in 395 patients was analysed along with other prognostic factors in breast cancer. Results: AIB1 expression scores ranged from 0 to 30; positive AIB1 expression (score > 14) was seen in 146/395 breast cancers; it correlated negatively with ER (P = 0.003) and progesterone receptor (PR) (P = 0.007), and positively with HER‐2 (P = 0.005) and tumour grade (P = 0.014). It did not correlate with nodal status (P = 0.437). Among ER+ patients, AIB1 expression showed a trend towards loss of PR expression (29% versus 20%; P = 0.14). AIB1 did not predict survival on univariate or multivariate analysis. Conclusions: AIB1 expression correlates with HER‐2 expression in breast cancer and shows a trend of association with loss of PR expression in ER+ tumours. Our study supports the postulated role of AIB1 in ER–growth factor interactions.     

The Journal of Pathology 2008 Jeremy Jass Prize for Research Excellence in Pathology

The first Jass Prize for Research Excellence has been awarded to a group from Hannover in Germany. These authors discovered the epigenetic inactivation of microRNA gene hsa‐mir‐9‐1 in human breast cancer and characterized its biological and clinical relevance. This frequent epigenetic silencing was found to occur early in the development of breast cancer, and illustrates another mechanism by which tumour development is influenced by genes that operate without expression as proteins. Copyright © 2009 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Epigenetic inactivation of microRNA gene hsa-mir-9-1 in human breast cancer

MicroRNAs (miRNAs) represent a new class of small non-coding RNAs regulating gene expression by inducing RNA degradation or interfering with translation. Aberrant miRNA expression has been described for several human malignancies and tumour suppressor functions have been ascribed to this new class of small regulatory RNAs. Accordingly, inactivation due to deletion or mutation has been found in human malignancies. Here, we describe the role of aberrant hypermethylation as an additional mechanism for miRNA gene inactivation in human breast cancer. Aberrant hypermethylation was shown for mir-9-1, mir-124a3, mir-148, mir-152, and mir-663 in 34-86% of cases in a series of 71 primary human breast cancer specimens. For comprehensive methylation analysis, combined bisulphite restriction analysis, bisulphite sequencing, and Pyrosequencing were employed. miRNA gene hypermethylation correlated strongly with methylation of known tumour suppressor genes (p = 0.003). After treatment of various breast cancer cell lines with the demethylating agent 5-aza-2'-deoxycytidine, reduction of mir-9-1 gene methylation and concomitant reactivation of expression could be observed. For the mir-9-1 gene, which is already hypermethylated in pre-invasive intraductal lesions, a good correlation between quantitative methylation level and reduction of expression could be demonstrated in a subset of primary human breast cancer specimen (r = 0.8). In conclusion, this study demonstrates that various microRNA genes are also affected by epigenetic inactivation due to aberrant hypermethylation and that this is an early and frequent event in breast cancer development.     

Alterations of microRNAs are associated with impaired growth of MCF-7 breast cancer cells induced by inhibition of casein kinase 2

Background and aim: Protein Kinase (casein kinase 2, CK2) is a pleiotropic serine-threonine kinase that is frequently dysregulated in many human tumors; microRNAs (miRNAs) are a class of small noncoding RNAs which play important roles in human cancers. This study aimed to investigate the role of CK2 and miRNA expression in breast cancer. Methods: Casein kinase 2 in MCF-7 breast cancer cell line was inhibited by the CK2 inhibitor TBB (4,5,6,7-tetrabromobenzotriazole), then cell proliferation was studied using MTT assay, and cell cycle distribution and apoptosis were detected by flow cytometry. The alteration of microRNAs expression profile was determined by microarray technology, followed by RT-PCR confirmation. Results: Here, we for the first time showed that inhibition of CK2 in MCF-7 breast cancer cells causes suppressed cell growth, which was related with dysregulation of the miRNA profile and altered expression. CK2 inhibition induced the up-regulated expression of 17 miRNAs and 10 down-regulated microRNAs which contributed to the impaired growth, inhibited cell cycle progress and increased apoptosis of MCF-7 cells by a CK2 inhibitor. Conclusions: These findings highlight the potential role of dysregulated miRNA expression regulated by CK2 in breast cancer.