本文選題：紫紺型先心病　切入點(diǎn)：慢性缺氧　出處：《第三軍醫(yī)大學(xué)》2012年博士論文　論文類(lèi)型：學(xué)位論文

【摘要】：背景： 紫紺先天性心臟病是由心內(nèi)右向左分流引起的，可造成患者慢性缺氧及紫紺，臨床新生兒的發(fā)病率及死亡率很高�；颊邫C(jī)體可通過(guò)增加血紅蛋白、肌紅蛋白、細(xì)胞內(nèi)線(xiàn)粒體的數(shù)量及加強(qiáng)糖酵解等多種代償方式，以增加氧氣的釋放，并減低能量的消耗，重塑細(xì)胞的能量供求平衡，使心肌細(xì)胞能夠最大限度的適應(yīng)缺氧環(huán)境，維持細(xì)胞基本的生存和功能。 鐵是肌紅蛋白、細(xì)胞色素及線(xiàn)粒體電子傳遞鏈中輔基的重要組成成分，是機(jī)體不可缺少的重要元素，主要參與心肌細(xì)胞對(duì)氧氣的轉(zhuǎn)運(yùn)和利用。在低氧環(huán)境中，有很多參與鐵穩(wěn)態(tài)的基因表達(dá)發(fā)生變化，其中很多基因?yàn)榈脱跽T導(dǎo)因子HIFs(Hypoxia-inducible factors HIFs)的直接靶點(diǎn)。HIFs可直接作用于目的基因的低氧反應(yīng)原件(hypoxia-responsive elements HRE)，以增強(qiáng)其表達(dá)。轉(zhuǎn)鐵蛋白受體-1(Transferrin Receptor-1TfR-1)為心肌細(xì)胞攝取鐵的主要蛋白之一。有學(xué)者研究發(fā)現(xiàn)，在TfR-1基因的啟動(dòng)子上存在一個(gè)HRE。但TfR-1在心肌慢性缺氧與鐵代謝間存在著怎樣的作用仍不明確，我們提出假設(shè)：在慢性缺氧環(huán)境下，心肌細(xì)胞可能通過(guò)調(diào)節(jié)TfR-1的表達(dá)從而改變心肌鐵代謝，以實(shí)現(xiàn)心肌慢性缺氧適應(yīng)。 本研究將通過(guò)觀察紫紺先心病患兒血清鐵及總鐵結(jié)合力、心肌中非血紅素鐵的含量以及TfR-1的表達(dá)情況，明確是否在紫紺先心病患兒外周血及心肌組織中伴隨鐵代謝的改變，進(jìn)而探討心肌組織內(nèi)鐵含量及TfR-1對(duì)心肌組織慢性缺氧適應(yīng)的意義。 研究方法： 本研究經(jīng)新橋醫(yī)院倫理委員會(huì)批準(zhǔn)，共分三個(gè)分題： 分題一：選取紫紺先心病患兒23例，非紫紺先心病患兒24例。以靜脈血為標(biāo)本，檢測(cè)患兒血紅蛋白含量、紅細(xì)胞壓積，以亞鐵嗪法檢測(cè)血清鐵及總鐵結(jié)合力，以了解紫紺先心病患兒機(jī)體鐵儲(chǔ)備的改變。 分題二：選取紫紺先心病患兒20例，非紫紺先心病患兒15例。以術(shù)中右室流出道心肌組織為標(biāo)本，使用亞鐵嗪法檢測(cè)心肌組織中非血紅素鐵含量，并用Perls-DAB法檢測(cè)心肌組織中鐵的分布，以了解紫紺先心病患兒心肌組織攝取鐵的變化。 分題三：以分題二中入組病例右室流出道心肌為標(biāo)本，采用Western blot法檢測(cè)心肌組織中TfR-1蛋白的表達(dá)量，免疫組化法檢測(cè)心肌組織中TfR-1的分布和定位，以了解TfR-1蛋白在紫紺先心病患兒心肌組織中的表達(dá)變化。 研究結(jié)果： 主要結(jié)果如下： 1.兩組患兒血紅蛋白含量及紅細(xì)胞壓積都基本處于正常范圍內(nèi)，但組間差異較大。與非紫紺組相比，紫紺組先心病患兒血紅蛋白含量明顯升高(166.7g/L±22.1g/L比125.6g/L±11.9g/L, p0.05)，紫紺先心病患兒平均紅細(xì)胞壓積也明顯高于非紫紺組(52.4%±8.8%比37.7%±3.1%，p0.05)。 2.亞鐵嗪法檢測(cè)患兒血清鐵及總鐵結(jié)合力，與非紫紺先心病患兒相比，紫紺組患兒血清鐵明顯下降(23.25μmol/L±4.4μmol/L比31.34μmol/L±4.2μmol/L, p0.05)，總鐵結(jié)合力增加(69.59μmol/L±5.8μmol/L比52.67μmol/L±9.2μmol/L, p0.05),與非紫紺先心病患兒組差異顯著。 3.亞鐵嗪法檢測(cè)患兒心肌非血紅素鐵含量，與非紫紺組患兒相比，紫紺組患兒心肌非血紅素鐵明顯升高，相比有顯著差異(6.12μmol/g±0.67μmol/g比4.02μmol/g±0.68μmol/g，p0.05)。Perls-DAB法檢測(cè)心肌中鐵的分布，非紫紺組患兒心肌可見(jiàn)少量陽(yáng)性顆粒，紫紺組患兒心肌可見(jiàn)大小不等的陽(yáng)性顆粒明顯增多。 4.Western blot檢測(cè)結(jié)果顯示：與非紫紺組患兒相比，紫紺組患兒心肌中TfR-1蛋白表達(dá)明顯增強(qiáng)，其相對(duì)光密度比值(OD)明顯強(qiáng)于非紫紺組(0.62±0.02比0.39±0.02，p0.001)。免疫組化結(jié)果顯示：在紫紺組患兒心肌胞漿中及胞膜上TfR-1蛋白廣泛表達(dá)，較非紫紺組明顯增加。 結(jié)論： 1.在紫紺先心病患兒體內(nèi)，外周血血紅蛋白及紅細(xì)胞壓積的升高，伴隨血清鐵下降以及總鐵結(jié)合力的增加。提示在慢性缺氧的情況下，患兒機(jī)體處于一種潛在的鐵缺乏狀態(tài)，大量的鐵可能被機(jī)體組織攝取，用于適應(yīng)這重慢性低氧環(huán)境，如合成血紅蛋白及紅細(xì)胞數(shù)量以增加血液的攜氧能力，造成機(jī)體鐵儲(chǔ)備下降。 2.紫紺先心病患兒心肌組織中鐵顆粒明顯增多，，并且伴隨非血紅素鐵濃度也明顯升高，表明除血紅蛋白的合成需要更多的鐵，心肌組織內(nèi)相應(yīng)含鐵蛋白合成的增強(qiáng)，也需要心肌組織攝取大量的鐵，這一改變可能為心肌慢性缺氧適應(yīng)中的關(guān)鍵環(huán)節(jié)。 3.紫紺先心病患兒心肌組織中TfR-1的表達(dá)明顯高于非紫紺組。提示在慢性缺氧環(huán)境中，TfR-1作為心肌組織中攝取鐵的主要分子，其表達(dá)增強(qiáng)有助于心肌細(xì)胞攝取更多的鐵，協(xié)助心肌細(xì)胞完成低氧適應(yīng)機(jī)制。
[Abstract]:Background:
Cyanotic congenital heart disease is caused by intracardiac right to left shunt, can cause patients with chronic hypoxia and cyanosis, morbidity and mortality in neonates is high. Patients can increase hemoglobin, myoglobin, the number of mitochondria in cells and enhance glycolysis and other compensatory way to increase oxygen release, and to reduce energy consumption, energy supply and demand balance remodeling cells, myocardial cells can adapt to hypoxic environment, to maintain the survival and function of cells.
Iron is myoglobin, cytochrome and an important component of mitochondrial electron transfer cofactor in the chain, is an important element of the indispensable, mainly involved in the myocardial cells of oxygen transport and utilization. In hypoxia environment, there are many expression of genes involved in iron homeostasis changes, many of them because of hypoxia inducible factor HIFs (Hypoxia-inducible factors HIFs) hypoxia response of the original direct targets of.HIFs can be directly applied to the target gene (hypoxia-responsive elements HRE), to improve the expression of transferrin receptor. -1 (Transferrin Receptor-1TfR-1) is one of the main protein in myocardial uptake of iron. Some scholars have found that in the promoter of TfR-1 gene, but the existence of a HRE. TfR-1 in the myocardium of chronic hypoxia and iron metabolism exist between what role is not clear, we hypothesized that in chronic hypoxia environment, myocardial cell The myocardial iron metabolism may be altered by regulating the expression of TfR-1 in order to realize the chronic hypoxia adaptation of the myocardium.
This study through the observation of cyanosis congenital heart disease in children with serum iron and TIBC, non heme iron content and the expression of TfR-1 in myocardium, whether in cyanotic congenital heart disease in children with peripheral blood and myocardial tissue with iron metabolism, and then discuss the Fe and TfR-1 in myocardial tissue of myocardial adaptation to chronic the significance of hypoxia.
Research methods:
The study was approved by the Xinqiao Hospital Ethics Committee and divided into three points:
Topic: the cyanosis congenital heart disease in children with 23 cases of non cyanotic congenital heart disease in children. In 24 cases of venous blood were detected with hemoglobin, hematocrit, the FerroZine assay of serum iron and total iron binding capacity, in order to understand the cyanotic congenital heart disease in children body iron reserves change.
Section two: selection of cyanotic children with congenital heart disease in 20 cases, 15 cases of non cyanotic congenital heart disease in children. As of right ventricular outflow tract myocardium were non heme iron content of myocardial tissue were detected using FerroZine method, and the distribution of Perls-DAB was used to detect the myocardial tissue in congenital heart disease, in order to understand the changes of myocardial tissue in purple purple children with iron uptake.
Section three: the topic 2 cases of right ventricular outflow tract myocardium specimens, TfR-1 protein expression in myocardial tissue was detected by Western blot method, the distribution and localization of TfR-1 were detected by immunohistochemistry in myocardial tissue, in order to understand the TfR-1 protein in cyanotic children with congenital heart disease myocardial tissues.
The results of the study:
The main results are as follows:
1. two groups of children with hemoglobin and hematocrit were basically in the normal range, but the differences between the groups. Compared with non cyanotic group, cyanosis group of children with congenital heart disease hemoglobin content increased obviously (166.7g/L + 22.1g/L 125.6g/L + 11.9g/L, P0.05), cyanosis congenital heart disease in children with average hematocrit were significantly higher than those in non cyanosis group (52.4% + 8.8% to 37.7% + 3.1%, P0.05).
2. children with FerroZine method to detect serum iron and TIBC, compared with non cyanotic congenital heart disease in children, serum iron group cyanosis decreased significantly (23.25 mol/L + 4.4 mol/L than 31.34 mol/L + 4.2 mol/L, P0.05), total iron binding capacity increased (69.59 mol/L + 5.8 mol/L 52.67 mol/L + 9.2 mol/L, P0.05), and non cyanotic children with congenital heart disease group were significantly different.
3. children with FerroZine method detection of myocardial non heme iron content, compared with non cyanosis group, cyanosis group with myocardial non heme iron was significantly increased, there was significant difference (6.12 mol/g + 0.67 mol/g than 4.02 mol/g + 0.68 mol/g, P0.05).Perls-DAB method for detection of myocardial iron distribution, non cyanosis group a small amount of myocardial positive granules, positive granules were visible cyanosis myocardial sizes increased significantly.
4.Western blot test results show that: compared with non cyanosis group, significantly enhanced the expression of TfR-1 protein in cardiac muscle in cyanotic group, the relative optical density ratio (OD) was significantly stronger than that of non cyanosis group (0.62 + 0.02 to 0.39 + 0.02, p0.001). Immunohistochemistry results showed that: in the group with myocardial cytoplasm cyanosis and cell membrane TfR-1 protein is widely expressed, compared with non cyanotic group increased significantly.
Conclusion:
In 1. children with congenital heart disease in cyanosis, increased peripheral blood hemoglobin and hematocrit, serum iron decreased with the increase of TIBC. In chronic hypoxia condition, with the body in a potential iron deficiency, large amounts of iron may be the body tissue uptake, to adapt to this heavy chronic hypoxia environment, such as the number of synthesis of hemoglobin and red blood cells to increase blood oxygen carrying capacity, resulting in a decline in body iron reserves.
2. cyanosis congenital heart disease myocardial tissues of children with iron particles increase obviously, and the non heme iron concentration also increased significantly, except for that of hemoglobin synthesis needs more iron, myocardium containing enhanced ferritin synthesis, myocardial tissue uptake also needs a large amount of iron, this change may be the key link of myocardial adaptation to chronic hypoxia.
The expression of TfR-1 3. in cyanotic children with congenital heart disease myocardial tissue were significantly higher than those in non cyanotic group. In chronic hypoxia environment, TfR-1 as the main molecular iron uptake in myocardial tissue, its expression contribute to enhancement of myocardial uptake more iron, myocardial cells adapt to hypoxia assist complete mechanism.

【學(xué)位授予單位】：第三軍醫(yī)大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2012
【分類(lèi)號(hào)】：R725.4

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