Runx3 might participate in regulating dendriti cell function in patients with irritable bowel syndrome

*

1Ningxia Medical University, Yinchuan 750004, Ningxia Province, China

2Department of Gastroenterology, Ningxia People's Hospital, Yinchuan 750004, Ningxia Province, China

3Department of Gastroenterology, Hainan Provincial People’s Hospital, Haikou 570311, China

Runx3 might participate in regulating dendriti cell function in patients with irritable bowel syndrome

Hua-Zhi Wu1, Man-Ni Cai1, Yu An2, Cheng Lan3, Jia-Li Wei3, Xiao-Ning Sun3*

1Ningxia Medical University, Yinchuan 750004, Ningxia Province, China

2Department of Gastroenterology, Ningxia People's Hospital, Yinchuan 750004, Ningxia Province, China

3Department of Gastroenterology, Hainan Provincial People’s Hospital, Haikou 570311, China

Objective:To evaluate the expression levels and correlations among the expressions of transforming growth factor-β1(TGF-β1) , Runx3 and CD83 in colonic mucosal specimens from IBS patients.Methods:A total of 40 patients were selected, who were confirmed as IBS by Rome III standard and 40 healthy volunteers served as control. Colonic mucosal specimens of each subject were collected from colon sigmoideum with biopsy forceps. Runx3, TGF-β1, and CD83 (the marker for immunecompetent mature dendritic cells (DCs) mRNA in the sigmoid colon tissue were measured by real-time fluorescence quantitative PCR.Results:Compared with the control group, CD83 mRNA expressions were higher in patients with IBS than in healthy controls (P<0.05) and were associated with runt-related transcription factor 3 (Runx3) mRNA levels (r=-0.361,P<0.05). Meanwhile, Runx3 mRNA levels were associated with TGF-β1mRNA expressions in irritable bowel syndrome (IBS) patients (r=0.402,P<0.05). However, there was no correlation between the mRNA expressions of TGF-β1and CD83 (P>0.05).Conclusions:The increase of abnormal dendritic cells might influence the occurrence and development of IBS. TGF-β1signal pathway might not be involved in Runx 3-regulated maturation of dendritic cells in IBS.

ARTICLE INFO

Article history:

Received 10 March 2014

Received in revised form 15 April 2014

党的十九大报告指出：“鼓励创业带动就业……促进高校毕业生等青年群体多渠道就业创业”[1]。近年来，“社会创业”逐渐成为一种世界性的教育潮流，在创造就业岗位、维护社会稳定和促进经济发展方面发挥着巨大的作用。我国高校也开始将社会创业教育作为高校创新创业教育改革的重要切入点，鼓励青年学生站在解决社会问题的出发点，接受社会创业教育，锤炼社会创业能力。这不仅是新时代落实创新型国家建设战略的必然要求，也是实现高等教育提质增效发展的应然选择。有鉴于此，厘清高校创业教育理念偏差，从由针对部分学生的精英式创客教育转向致力于面向全体学生，培育具有创新精神和素养的创新型人才为旨归的“社会创业”教育显得尤为必要。

Accepted 15 July 2014

Available online 20 September 2014

从表2的数据可以看出，速度测量不确定约为7.61×10-4m/s，而跟2.2节的分析结果，当信噪比48 dB时，理论上测速精度可达4.062 3×10-4m/s。这是因为我们只考虑了系统电学部分带来的测量不确定度，而测速系统中存在时钟抖动，光学器件误差等因素，均会对测速精度带来一定影响。

采用热盘炉预焚烧技术可使固体废弃物的整个燃烧过程发生在热盘炉内，最大程度上减少废弃物燃烧对分解炉产生的冲击，并增加固体废弃物炉内停留时间5～45min，使固体废弃物在1045℃的高温下彻底分解。

Irritable bowel syndrome

Runt-related transcription factor 3

Transforming growth factor-β1

Dendriti cell

CD83

西地那非是欧洲泌尿外科协会(European Association of Urology，EAU)指南推荐治疗勃起功能障碍(erectile dysfunction，ED)的一线治疗药物，能安全有效的改善各种ED患者的勃起功能[1]。早泄 (premature ejaculation，PE)是临床上最常见的男性性功能障碍，其患病率高达20%～30%[2]。有研究表明，30%～50%的ED患者同时存在PE[3]。为探究西地那非治疗合并PE的ED患者的临床疗效及安全性，我们对合并PE的ED患者应用按规律联合按需服用西地那非疗法进行了临床观察，现报告如下。

1. Introduction

Irritable bowel syndrome (IBS) is a common intestinal disorder characterized by persistent or intermittent abdominal pain or discomfort,distention,and changes in stool pattern. It has been became a serious health problem that affacts an estimated 2.9%-15.6% of people in Asian countries nowadays[1]. IBS is associated with abnormal intestinal motion and sensations,intestinal infection,hypothalamic-pituitary-gut axis dysregulation. Recently, a growing number of findings proved that immune activation play a majoy role in the pathophysiology of IBS[2]. The intestinal mucosa contains numerous DCs, which can mediate the innate and the adaptive immune system[3,4].

[3] Steinman RM. Dendritic cells: versatile controllers of the immune system. Nat Med 2007; 13: 1155-1159.

Transforming growth factor-β1(TGF-β1) belongs to a well-defined multipotent cytokine family known to regulate several pathophysiological events and exhibit the broadest spectrum of biological activities. Runt-related transcription factor 3 (Runx 3) is a novel tumor-suppressor gene,it involved in the differentiation of immune cells[5], especially the each step of T-cell differentiation[6]. As a part of the innate immune response, Runx3, TGF-β1and dendriti cells have been postulated to participate in mucosal and systemic immune responses in many studies,but the relationship of them have not been reported. The aim of this study was to evaluate the expression levels and correlations among the expressions of TGF-β1, Runx3 and CD83 in colonic mucosal specimens from IBS patients.

2. Materials and methods

2.1. Subjects and specimens

A total of 40 patients were selected,who were confirmed as IBS by Rome III standard and were admitted from March to December, 2013 at the clinic of digestive diseases, Hainan Provincial People’s Hospital (Haikou, China). All patients were evaluated by colonic-rectum endoscopy and showed normal colonic tissue. Meanwhile, 40 healthy volunteers were selected as control. Colonic mucosal specimens ofeach subject were collected from colon sigmoideum with biopsy forceps and specimens were preserved in stored at -80 ℃immediately for subsequent RNA extraction.

2.2. Real-time PCR

在生活自理能力方面，通过对793位入住老年人的调查，发现383位老年人完全正常（48.30%），279位老年人具有不同程度功能下降（35.18%），131位老年人存在功能明显障碍（16.52%）。380位男性老年人中，188位完全正常（49.47%），132位具有不同程度功能下降（34.74%），60位存在功能明显障碍（15.79%）。413位女性老年人中，195位完全正常（47.22%），147位具有不同程度功能下降（35.59%），71位存在功能明显障碍（17.19%），男性和女性在生活自理能力水平的分布位置相同（P＞0.05）（见表 6）。

Table 1 Primers in this study.

2.3. Statistical analysis

Statistical analysis was performed using SPSS software version 17.0. Results are reported as means±SD Student’st-test was used to compare Runx3, TGF-β1and CD83 mRNA levels between the IBS and control group. Correlations between two data sets were assessed using Persons correlation test. P-values of <0.05 were considered significant.

3. Results

3.1. Runx3, TGF-β1and CD83 mRNA expressions in sigmoid colon tissue from IBS patients

Compared with the control group, CD83 mRNA expressions were higher in patients with IBS than in healthy controls (P<0.05) (Figure 1). However , there was no significance in colonic mRNA expressions of Runx 3 (P>0.05) and TGF-β1(P>0.05) when compared with the control group.

3.2. Correlations among mRNA expressions of TGF-β1, Runx3 and CD83

CD83 mRNA expressions were associated with Runx3 mRNA levels (r=-0.361,P<0.05). Meanwhile, Runx3 mRNA levels were associated with TGF-β1mRNA expressions in IBS patients (r=0.402,P<0.05) (Figure 2). But there is no correlation between the mRNA expressions of TGF-β1and CD83 (P>0.05).

Figure 1. Runx3, TGF-β1and CD83 mRNA expressions in sigmoid colon tissue from IBS patients.Compared with the control group, CD83 mRNA expressions were higher in patients with IBS than in healthy controls (P<0.05). However, there was no significance in colonic mRNA expressions of Runx 3 (P>0.05) and TGF-β1(P>0.05) when compared with the control group.

Figure 2. Correlations among mRNA expressions of TGF-β1, Runx3 and CD83.CD83 mRNA expressions were associated with Runx3 mRNA levels (r=-0.361, P<0.05). Meanwhile, Runx3 mRNA levels were associated with TGF-β1mRNA expressions in IBS patients (r=0.402,P<0.05). But there is no correlation between the mRNA expressions of TGF-β1and CD83 (P>0.05).

4. Discussion

IBS is a functional gastrointestinal disorder characterized by abdominal pain and altered bowel habits in the absence of specific and unique organic pathology. Its pathophysiology is still not entirely clear.Emerging evidence revealed that inflammation play a crucial role in the occurrence and development of IBS[7,8].

[6] Kohu K, Kubo M, Ichikawa H, Ohno S, Habu S, Sato T, et al. Pleiotropic roles of Runx transcription factors in the differentiation and function of T lymphocytes. Curr Immunol Rev 2008; 4: 101-115.

Some research shows that TGF-β1can inhibit the maturation of DC. TGF-β1plays an important role in mediating the balance of the inflammation responses within the intestinal mucosa. Notably, TGF-β1promotes early dendritic cells’ development in vitro and suppresses immature dendritic cells’ activation and maturation[13]. For exampled treated with TGF-β1in vitrobecome less responsive to maturation stimuli such as IL-1β and TNF-α[14], thereby reducing the efficacy of dendritic cells to stimulate T lymphocytes.

The mammalian RUNX3 gene resides on human chromosome 1p36.1 and mouse chromosome 4 respectively. It belongs to the RUNX family of transcription factors, which contains three genes. In mice, Sugaiet al[5] found that the loss of Runx3 in T cells resulted in suppression of Treg cell function which lead to the development of colitis in Runx3-/- animals.

Runx3 is highly expressed in DCs, where its functions as a component of TGF-β signaling cascade[15]. Runx3 KO DCs do not respond to TGF-β, their maturation is accelerated and accompanied by an increase defficacy to stimulate T cells,the abnormal DCs function constitute a primary immune system defect associated with spontaneous development of lung inflammation in the KO mice[15].

[5] Sugai M, Aoki K, Osato M, Nambu Y, Ito K, Taketo MM, et al. Runx3 is required for full activation of regulatory T cells to prevent colitis-associated tumor formation. J Immunol 2011; 186: 6515-6520.

4.依法保护民营企业合法权益。加强民营企业产权保护，依法保障民营企业和企业家合法财产不受侵犯、合法经营不受干扰。严厉打击针对民营企业的不正当竞争行为，对侵犯民营企业商标、专利、商业秘密等知识产权以及损害民营企业商业信誉、商品声誉的违法行为，及时予以纠正。对民营企业和人员的一般违法行为，慎用查封、扣押、冻结等措施，最大限度降低对涉案企业正常生产经营活动的不利影响。依法必须采取查封、扣押、冻结措施处置涉案财物的，必须严格区分个人财产和企业法人财产、区分违法所得和合法财产、区分涉案人员个人财产和家庭成员财产，结案后及时解封、解冻非涉案财物。

Interestingly, in IBS patients, there is a significant correlation between Runx3 mRNA and TGF-β1, suggesting that Runx3 - TGF-β1inflammation pathway might be involved in human IBS. Runx3 mRNA and CD83 mRNA also have certain correlation, but there is no correlation between TGF-β1mRNA and Runx3 mRNA,this may imply that Runx3 may regulate the maturation of dendritic cells through other signal transduction system in IBS patients.

Conflict of interest statement

We declare that we have no conflict of interest.

几十年来，右心前导联高而宽的R波被认为是后壁心肌梗死的征象。但MRI-MDE与心电图相互关系的新证据表明，在排除右心室肥大、完全性右束支阻滞或预激综合征等影响QRS波形态的因素后，在急性冠脉综合征发展过程中，V1导联出现高R波提示侧壁心肌梗死，并且梗死面积通常更大、透壁程度更深。

[1] Gwee KA, Bak YT, Ghoshal UC, Gonlachanvit S, Lee OY, Fock KM, et al. Asian consensus on irritable bowel syndrome. J Gastroenterol Hepatol 2010; 25: 1189-1205.

[2] Lee HF, Hsieh JC, Lu CL, Yeh TC, Tu CH, Cheng CM, et al. Enhanced affect/cognition-related brain responses during visceral placebo analgesia in irritable bowel syndrome patients. Pain 2012; 153: 1301-1310.

另一方面，作为老牌学科，《组织学与胚胎学》的教学仍然基本采用传统式的课堂讲授。教学普遍存在“一言堂”的现象，以教师为中心的观念非常浓厚，这样往往会造成教学过程单调乏味，没有良好的师生互动。课上、课下缺乏获得资源和开展协作式学习的平台，致使学生的作业与问题等得不到及时的反馈和解决。更为糟糕的其实是学生的学习过程不能得到应有的评价，往往是以期末考试试卷的成绩为主。这种“一锤子买卖”使得学生在平时的学习过程中无法客观地、及时地看到自己的表现，进而便不能做到及时的自我调整和提高，久而久之便抹杀了学生日常学习的积极性。

[4] Steinman RM, Hemmi H. Dendritic cells: translating innate to adaptive immunity. Curr Top Microbiol Immunol 2006; 311: 17-58.

Similar to the result of our previous study on rat model with IBS, the colonic level of Runx3 mRNA and TGF-β1mRNA did not show any remarkable changes in IBS group compared with their control group. This negative data may be result from the clinic type of IBS, the lasting time of the inflammation and the depth of the samples. But the expression of CD83 mRNA was higher than the control group, which indicates the maturation of dendritic cells maybe a important role in IBS patients.

中小型工业企业的创始人普遍都不是受过高等教育的高精尖人才，大多是凭着自身努力和钻研起家的草根，虽然有很强的毅力和拼搏精神，但是由于缺乏理论教育，企业往往做每一件事都是摸着石头过河。企业内部工作中推诿扯皮现象屡见不鲜，每个人都觉得自己很忙，但是工作的结果又都往往很粗糙、漏洞百出。企业管理层面对这些现象，总是头疼医头，脚疼医脚，不知道主要问题出在哪里，也不清楚该采取哪种预案应对这些不足，无法形成系统的管理体系。

Dendritic cells (DCs) are key regulators in the immune system. DCs are found in two functionally distinct states: immature and mature dendritic cells. Immature DCs are present in peripheral tissues and are mainly phagocytic cells. Mature DCs are specialized antigen presentation cells that orchestrate innate and adaptive immune responses, they can act by priming abnormal T cell responses to the enteric flora in organized lymphoid tissues. Sustain DCs reactivity within the inflamed mucosa via the release of proinflammatory cytokines, for example, mature DCs expressing higher levels of costimulatory molecules (CD40,CD80, and CD86) and increased amounts of IL-12p40 and IL-23p19 upon CD40 ligation[9]. While IL-12p40 and IL-23p19 can form IL-23, which is important for the growth and stabilization of Th17 cells in the mouse and their differentiation in humans[10], while Th17 cell differentiation can adrive inflammation. DCs play an important role in the occurrence and development of intestinal disease. Several observations suggest that DCs may play a pathogenic role in humans and in mouse models of inflammatory bowel diseases (IBDs), including Crohn disease and ulcerative colitis, are chronic relapsing inflammatory diseases of the gastrointestinal tract. Murakamiet al[11] showed an increase of mucosal CD83+and CD86+cells producing macrophage inhibitory factor, which is thought to contribute to neutrophil recruitment and activation in ulcerative colitis. Cremonet al[12] demonstrated that the magnitude of the immune infiltrate detected in patients with IBS was markedly lower than that of active or quiescent ulcerative colitis in some comparative studies, this may imply that DCs also plays a role in intestinal inflammation in IBS patients.

Total RNA was isolated from colonic tissue using the RNeasy Micro Kit (Qiagen, Santa Clarita, CA), reverse transcription was performed using the Power RT Kit cDNA (Bio TeKe, Beijing, China) according to the manufacturer’s protocols. Real-time PCR was performed to measure expression levels of target mRNAs using a 2×SYBR realtime RT-PCR premixture (Bio TeKe, Beijing, China). Briefly, the reactions were incubated at 95 ℃ for 2 min, followed by 45 cycles of denaturation at 95 ℃ for 15 s, annealing at 60 ℃ for 60 s in ABI PRISM 7300 Sequence Detection System Thermal Cycler(Applied Biosystems). Oligonucleotides (Table 1) were designed by Primer 5 software and synthesised at the Sangon Biotech (Shanghai, China).

[7] Corinaldesi R, Stanghellini V, Cremon C, Gargano L, Cogliandro RF, De Giorgio R, et al. Effect of mesalazine on mucosal immune biomarkers in irritable bowel syndrome:a randomized controlled proofof-concept study. Aliment Pharmacol Ther 2009; 30: 245-252.

[8] Feng B, La JH, Schwartz ES, Gebhart GF. Neural and neuroimmune mechanisms of visceral hypersensitivity in irritable bowel syndrome. Am J Physiol Gastrointest Liver Physiol 2012; 302: G1085-G1098.

“诗佛”王维擅作山水田园诗，但也写过“莫嫌旧日云中守，犹堪一战取功勋”这样激昂慷慨的颂歌。以你对王维的了解，你能猜出下面哪句诗不是出自王维之手吗？

[9] Krajina T, Leithauser F, Moller P, Trobonjaca Z, Reimann J. Colonic lamina propria dendritic cells in mice with CD4+T cellinduced colitis. Eur J Immunol 2003; 33: 1073- 1083.

[10] Korn T, Bettelli E, Oukka M, Kuchroo VK. IL-17 and Th17 cells. Annu Rev Immunol 2009; 27: 485-517.

3) 对于气动执行机构，若从经济角度考虑不便安装防火保护罩时，可以选择在气缸上加装易熔塞。当发生火灾时由于温度急剧升高，气缸温度达到或超过易熔塞的熔点时，易熔塞熔化将气缸内的压力泄放掉，使另一侧气缸内的弹簧或储气罐内的压缩空气推动活塞将阀门自动关闭。

[11] Murakami H, Akbar SM, Matsui H, Horiike N, Onji M. Macrophage migration inhibitory factor activates antigenpresenting dendritic cells and induces inflammatory cytokines in ulcerative colitis. Clin Exp Immunol 2002; 128: 504-510.

如果我们明确了弗雷格所说的“语句”与自然语句形同而实不同，那么自然不会对判断杠“|”的使用产生误解，也不会认为他把自然语言中的语句和单独词项混为一谈。为了使弗雷格的“语句”与自然语句的区别一目了然，我们可以为弗雷格逻辑系统中的“语句”加上标注:▲f，而具有判断力的表达式则为：|—▲f。那么，与自然语句“2+2=4”相当的并不是“（2+2=4）f”，也不是“—（2+2=4）f”，而是“|—（2+2=4）f”。

[12] Cremon C, Gargano L, Morselli-Labate AM, Santini D, Cogliandro RF, De Giorgio R. et al. Mucosal immune activation in irritable bowel syndrome: gender-dependence and association with digestive symptoms. Am J Gastroenterol 2009; 104: 392-340.

[13] Fogel-Petrovic M, Long JA, Misso NL, Foster PS, Bhoola KD, Thompson PJ. Physiological concentrations of transforming growth factor beta1selectively inhibit human dendritic cell function. Int Immunopharm 2007; 7: 1924-1933.

[14] Ohtani T, Mizuashi M, Nakagawa S, Sasaki Y, Fujimura T, Okuyama R, Aiba S. TGF-beta1 dampens the susceptibility of dendritic cells to environmental stimulation, leading to there quirement for danger signals for activation. Immunol 2009; 126: 485-499.

[15] Fainaru O, Woolf E, Lotem J, Yarmus M, Brenner O, Goldenberg D, et al. Runx3 regulated mouse TGF-β-mediated dendritic cell function and its absence results in airway inflammation. EMBO J 2004; 23: 969-979.

ment heading

10.1016/S1995-7645(14)60130-9

*Corresponding author: Xiao-Ning Sun, Department of Gastroenterology, Hainan Provincial People’s Hospital, Haikou 570311, China.

E-mail: xnsun_0108@163.com.

Foundation project: It is supported by Key Science and Technology Project of Hainan Province (Grant No.ZDXM20100042).