阿胶通过调控珠蛋白水平治疗β-地中海贫血患者妊娠期贫血的通路研究

Guangdong province is a high incidence area of thalassemia. After pregnancy, anemia in β-thalassemia patients tends to turn more serious, the risk of adverse pregnancy outcomes accordingly increase. Currently, no consensus has been reached in treating pregnant thalassemia patients due to lack of safe and effective treatment. Regulation of globin gene expression is the key link of β-thalassemia treatment, but the accessible western medicines have many limitations, including no definitive efficacy, bone marrow suppression, carcinogenicity and teratogenicity, and are not suitable for pregnant patients. Some traditional Chinese medicines have been shown to treat anemia in patients with β-thalassemia by inducing γ globin gene expression and promoting fetal hemoglobin (HbF,α2γ2) synthesis, but lack of clinical evidences of pregnant patients. Our previous clinical study showed that Colla corii asini (CCA) can significantly increase the level of hemoglobin and adult hemoglobin (HbA,α2β2) in the pregnant women with β-thalassemia, it was speculated that CCA might induce β globin gene expression, which would be more beneficial to pregnant women than the γ globin gene inducer. But its regulatory pathway is not clear. This study is designed to explore the pathways and targets of regulating globin expression by using the transcriptomics method, which might be related to CCA treating pregnant anemia in β-thalassemia patients. And then the results of transcriptomics study will be further verified by expanding the clinical samples and implementing cell experiments, aiming to explore the mechanisms of CCA in treating β-thalassemia with pregnant anemia by regulating globin expression. It would be of important strategic significances to develop new strategies for the treatment of anemia in thalassemia patients and improve pregnancy outcomes of patients with β-thalassemia.

广东省是地中海贫血高发地区，β-地贫患者妊娠后贫血易进行性加重，不良妊娠结局风险显著增高，目前缺乏安全有效的治疗方法。调控珠蛋白基因表达是β-地贫治疗的关键环节，但已有的西药存在疗效不确切、骨髓抑制和致癌致畸等缺陷，不适合孕妇使用。一些中药被证明有诱导γ珠蛋白基因表达，促进胎儿血红蛋白（HbF，α2γ2）合成，改善地贫患者贫血的作用，但缺乏孕妇的研究数据。本课题前期研究已证实阿胶能明显提高β-地贫孕妇血红蛋白（Hb）和主要成人血红蛋白（HbA，α2β2)水平，推测其可能有诱导β珠蛋白基因表达作用，较γ珠蛋白基因诱导剂更有益于孕妇，但其调控通路尚不清楚。本研究拟用转录组学方法分析阿胶治疗β-地贫孕妇调节珠蛋白表达的作用通路及靶标，进而通过扩大临床样本及细胞实验进行验证，探讨阿胶通过调节珠蛋白表达治疗β-地贫孕妇贫血的作用机制，对开拓地贫贫血治疗的新策略，改善地贫孕妇妊娠结局具有重

广东省是地中海贫血（简称地贫）的高发区，地贫患者妊娠后贫血易进行性加重，不良妊娠结局风险增高，缺乏安全有效的治疗方法。课题组前期研究证实口服阿胶治疗明显提高β-地贫孕妇血红蛋白（Hb）和主要成人血红蛋白（HbA，α2β2)水平，推测其可能有调节珠蛋白链失衡的作用。本研究采用转录组测序方法，研究阿胶治疗β-地贫合并妊娠期贫血患者前后外周血细胞转录组水平的变化，分析阿胶的抗贫血作用通路及潜在作用靶标；基于转录组学结果，通过实施一个随机、双盲、安慰剂平行对照临床试验，共纳入68例轻型或中间型地贫合并轻度妊娠期贫血的孕妇，从mRNA和蛋白两个水平验证治疗前后靶标和4种珠蛋白水平的变化；从6例不同基因型的地贫孕妇和1例健康孕妇外周血中分离单核细胞，通过二步细胞培养法诱导的红系祖细胞为模型，研究阿胶对红系祖细胞分化增殖、靶标基因和珠蛋白含量的影响。研究结果显示：(1)βCD41-42(-TTCT)/βN是受试者中最主要的基因类型，治疗后红细胞计数和Hb上升最明显，对基因表达的调控作用也最为明显。(2)差异表达基因的主要分子功能、分子组成及生物学过程涉及激活尿嘧啶合成、信使RNA氨基酸转运、激活核糖体结合、调控细胞周期、调节血影蛋白相关细胞骨架表达。(3)基因THOC5和ZNF471是潜在的作用靶标，分别参与了THOC5通路和含KRAB结构域锌指蛋白通路。(4)阿胶治疗可能有上调β-地贫贫血孕妇的β和δ-珠蛋白链表达的作用。(5)阿胶治疗对基因型为βCD41-42（-TTCT）/βN的地贫贫血孕妇的α、β和γ-珠蛋白的上调作用更明显，且不同个体间的疗效差异更小。(6)阿胶体外消化液浓度为100ug/ml，培养时间为72小时时促进细胞增殖的效果最佳；阿胶消化液对地贫细胞的珠蛋白表达的调节作用与地贫基因型有关。研究初步阐述阿胶治疗地贫患者妊娠期贫血的疗效与基因类型之间的关系，为更精准选择适宜的治疗人群、提高临床疗效提供实践依据；从调控靶基因表达、平衡珠蛋白比例和影响红细胞骨架形成等角度，阐述阿胶可能的抗贫血作用机制，为其临床应用提供理论依据。