内容翻译自2016年广州中医药大学第一附属医院妇儿中心科研团队发表在《国际血液学杂志（International Journal of Hematology）》上的《阿胶对改善地中海贫血孕妇贫血症和血红蛋白成分的治疗作用》。
Therapeutic effect of Colla corii asini on improving anemia and hemoglobin compositions in pregnant women with thalassemia
论文作者（英文名未翻译）：Yanfang Li1 · Hui He2 · Lilin Yang3 · Xiangyi Li1 · Daocheng Li1 · Songping Luo3
通讯作者（英文名未翻译）：Songping Luo firstname.lastname@example.org
1 Department of Obstetrics, The First Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, China
2 Department of Clinical Laboratory, The First Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, China
3 First School of Clinical Medicine, Guangzhou University of Chinese Medicine, No. 12 Jichang Road, Guangzhou 510405, Guangdong, China
Received: 15 April 2016 / Revised: 14 July 2016 / Accepted: 19 July 2016 . The Japanese Society of Hematology 2016
收稿日期：2016年4月15日/ 修订日期：2016年7月14日/ 接受日期：2016年7月19日。
Currently there is no consensus on treating ane-mia in pregnant thalassemia patients. In China, Colla corii asini (CCA) has been widely used for treating anemia for more than 2000 years. However, its clinical application in the thalassemia population is limited by a lack of quanti-tative evidence. The present study aims to investigate the therapeutic effect of CCA in increasing hemoglobin (Hb) concentration and improving abnormal hemoglobin com-positions in pregnant patients with β-thalassemia. Seventy-two pregnant patients who met inclusion criteria were ran-domly assigned to either the treatment group or control group. Patients in the treatment group were given 15 g of CCA, while the control group were observed and followed up without any treatment. Levels of Hb, serum iron (SI), serum ferritin (SF) and three types of Hb components [adult hemoglobin (HbA), fetal hemoglobin (HbF), minor adult hemoglobin (HbA2)] were measured before and after treatment. Treatment with CCA led to a significant increase of Hb. The major Hb component induced by CCA was HbA, while levels of both HbA2 and HbF dropped after treatment. CCA treatment significantly increased SI, while SF remained unaffected. Our data suggest that CCA can improve anemia and optimize Hb components in pregnant patients with thalassemia without affecting iron reserves.
目前关于妊娠地中海贫血患者贫血的治疗尚未达成共识。在中国，阿胶（Colla corii asini，CCA）已广泛被用于治疗贫血症超过两千多年了。然而，由于缺乏量化的证据，其在地中海贫血人群中的临床应用还很少。 本研究旨在探讨阿胶在增加妊娠β地中海贫血患者血红蛋白（Hb）浓度和改善异常血红蛋白组成方面的治疗效果。 符合纳入标准的72名孕妇随机分配到治疗组或对照组。 治疗组患者给予15g 阿胶，而对照组进行观察和随访，无需任何治疗。测定治疗前后血红蛋白（Hb）水平，血清铁（SI），血清铁蛋白（SF）和三种血红蛋白（Hb）成分[成人血红蛋白（HbA），胎儿血红蛋白（HbF），成人血红蛋白次要成分（HbA2）]。 用阿胶治疗导致血红蛋白（Hb）显着增加。由阿胶诱导产生的主要血红蛋白（Hb）组分是成人血红蛋白（HbA），而成人血红蛋白次要成分（HbA2）和胎儿血红蛋白（HbF）的水平在治疗后下降。 阿胶治疗显着增加血清铁（SI），而血清铁蛋白（SF）不受影响。我们的数据表明，阿胶可以改善贫血并优化怀孕地中海贫血患者的血红蛋白（Hb）成分，而不影响铁储备。
Keywords Thalassemia · Anemia · Pregnancy · Hemoglobin · Colla corii asini
Thalassemia is a type of hemolytic anemia disease caused by genetic defect of synthesis in one or more globin chains. Among all the single genetic disorders thalassemia has the highest incidence rate in the world and causes heavy bur-dens on public health system. In China, the southern prov-inces suffer from high incidence of thalassemia, which is particularly common in the population of Guangdong, Guangxi and Yunnan provinces. Epidemiological stud-ies showed that in Guangdong alone about 17.83 % of the 14,332 pregnant women across 21 regions examined were diagnosed as carriers of thalassemia .
地中海贫血是由一种或多种珠蛋白链中的合成遗传缺陷引起的一种溶血性贫血疾病。 在所有单基因遗传性疾病中，地中海贫血在世界上发病率最高，并在公共卫生体系中造成沉重负担。在中国，南方地区是地中海贫血的高发地区，这在广东，广西和云南省的中尤其普遍。 流行病学研究显示，仅在广东，21个地区的14,332名孕妇中约有17.83％被诊断为地中海贫血基因携带者。
Recent studies showed that compared with healthy con-trols, women with thalassemia are associated with a wide range of abnormality and adverse pregnancy outcomes including cardiovascular disease, thrombotic disease, spon-taneous miscarriage, premature delivery, oligohydramnios, fetal growth restriction and low birth weight. In addition, the severity of adverse pregnancy outcomes is closely linked to low level of hemoglobin (Hb) and high level of fetal hemoglobin (HbF) as well as iron-overloading related complications during pregnancy [2, 3]. Therefore, it is essential to improve the anemic state and optimize the compositions of Hb in pregnant thalassemia patients.
最近的研究表明，与健康对照组相比，地中海贫血女性与各种异常和不良妊娠结果相关，包括心血管疾病，血栓性疾病，自发性流产，早产，羊水过少，胎儿生长受限和低出生体重。 此外，不良妊娠结果的严重程度与妊娠期间血红蛋白（Hb）水平低，胎儿血红蛋白（HbF）水平高以及铁过载相关并发症密切相关[2,3]。 因此，改善妊娠地中海贫血患者的贫血状态和优化血红蛋白的组成是至关重要的。
As the pregnancy progresses, women experience a series of physiological change, which can lead to physiological and nutritional anemia in thalassemia patients without prior or only mild anemia, further complicating the conditions of thalassemia patients during pregnancy. The risk of anemia becomes higher when the patients enter third trimester and many of them may need treatment for disease management . Currently there is no consensus on treating anemia in pregnant thalassemia patients. Due to the absence of an effec-tive treatment scheme with improved safety, many thalas-semia patients are prone to develop low level of Hb, which can severely impact the fetal growth and maternal health.
随着妊娠的进行，女性经历一系列生理变化，这可能导致地贫患者生理性和营养性贫血，而此前没有或仅有轻度贫血，这使妊娠期地中海贫血患者的病情进一步复杂化。 当患者进入妊娠晚期时，患有贫血的风险会增高，其中许多患者可能需要疾病管理。 目前对怀孕地中海贫血患者治疗贫血没有共识。 由于缺乏安全性有效治疗方案，许多地中海贫血患者容易发生血红蛋白降低，这可能会严重影响胎儿生长和孕妇健康。
In traditional Chinese medicine (TCM), Colla corii asini (CCA, E’jiao) is a gelatin-like preparation derived from donkey hide and has been widely used in clinical antanemic therapy for more than 2000 years. In the last decade, many studies had addressed the effect of CCA on the antane-mia process using modern pharmacological approaches. The results indicated that collagen protein and polysac-charides which are the main components of CCA can pro-mote hematopoiesis by a number of mechanisms which eventually increase the peripheral erythrocyte counts and Hb concentration . Therefore, we proposed that these hematopoietic effects of CCA might also contribute to the treatment of thalassemia which is caused by insufficient or abnormal Hb concentration. However, its clinical applica-tion in the thalassemia population is limited by the lack of quantitative evidence. This study aims at investigating the therapeutic effect of CCA in treating mild anemia during pregnancy among β-thalassemia patients.
在中药中，阿胶是一种来自驴皮的明胶样制剂，已被广泛用于临床贫血治疗两千多年了。在过去的十年中，许多研究已经用现代药理学方法研究阿胶对于贫血过程的影响。 结果表明，阿胶主要成分胶原蛋白和多糖结合物可通过多种机制促进造血功能，能够最终增加外周红细胞计数和血红蛋白浓度。因此，我们提出阿胶的这些造血作用也可能有助于治疗由Hb浓度不足或异常引起的地中海贫血。然而，由于缺乏定量证据，其在地中海贫血人群中的临床应用还有限。 本研究旨在探讨阿胶治疗β地中海贫血患者妊娠期轻度贫血的疗效。
Materials and methods
This is a randomized, open labelled, controlled prospec-tive clinical trial. Subjects were pregnant women diagnosed of minor or intermediate β-thalassemia at the First Affili-ated Hospital of Guangzhou University of Chinese Medi-cine between March 2015 and December 2015. The study was reviewed and approved by the institutional Medical Research Ethics Committee and patient recruitment was conducted in strict accordance to the inclusion and exclu-sion criteria. Informed consent was obtained from all sub-jects before enrollment.
这是一项随机，开放标签，控制的前瞻性临床试验。 受试者为2015年3月至2015年12月在广州中医药大学第一附属医院诊断为轻微或中度β-地中海贫血的孕妇。 该研究由医学研究伦理委员会审查并批准，患者招募严格按照纳入和排除标准进行。 在测试之前所有受试者都已签署知情同意书。
Diagnosis, inclusion and exclusion criteria
Patients were diagnosed as carriers of thalassemia based on genetic test. Classification of mild and intermedi-ate β-thalassemia was based on the guidelines published by the Thalassemia International Federation in 2008 .
In accordance with the diagnostic criteria of the World Health Organization, patients were classified as mild ane-mia (60 g/L ≤ Hb < 110 g/L) and anemia during pregnancy (Hb < 110 g/L) .
根据世界卫生组织的诊断标准，患者被分为轻度贫血（60 g /L≤Hb<110 g / L）和妊娠期贫血（Hb <110 g / L）。
(1) Pregnant women diagnosed as thalassemia carri-ers by genetic test with clinical presentation of minor or intermediate β-thalassemia; (2) patients with mild ane-mia (80 g/L ≤ Hb < 110 g/L) prior to study enrollment; (3) singleton pregnancy; (4) patients having not received blood transfusion or any forms of anti-anemia treatment in conventional medicine or TCM in the last 12 weeks; (5) informed consent obtained.
（1）通过基因检测诊断为地中海贫血的孕妇，临床表现为轻度或中度β地中海贫血; （2）入选研究前轻度贫血（80 g /L≤Hb<110 g / L）的患者; （3）单胎妊娠; （4）过去12周内没有接受输血或任何形式的常规药物或中药的抗贫血治疗的患者; （5）获得知情同意书。
(1) Patients with major thalassemia; (2) patients with severe anemia (Hb < 80 g/L) prior to study enrollment; (3) twin or multiple pregnancies; (4) patients with any of the following abnormalities: immunodeficiency, primary diseases involving cardiovascular system, liver, kidney, gastrointestinal tract, endocrine system and hematological system; (5) allergic to two or more drugs; (6) patients with mental illness or poor compliance to medical treatment; (7) patients having received blood transfusion or any forms of anti-anemia treatment in conventional medicine or TCM in the last 12 weeks; (8) no informed consent obtained.
（1）重型地中海贫血患者; （2）入选研究前患有严重贫血（Hb <80 g / L）的患者; （3）双胞胎或多胎妊娠; （4）具有以下任何异常的患者：免疫缺陷，涉及心血管系统，肝，肾，胃肠道，内分泌系统和血液系统的原发性疾病; （5）对两种或多种药物过敏; （6）精神病患者或依从性差的患者; （7）过去12周内在常规药物或TCM中接受过输血或任何形式的抗贫血治疗的患者; （8）未获得知情同意。
Once diagnosed as thalassemia and enrolled in the study, patients would be required to cease taking medications that contain iron and folic acid. Patients in the treatment group received daily 15 g oral CCA (Shandong Dong-E E-Jiao Co., Ltd) in powder form for 4 consecutive weeks. The dos-age was adjusted to 10 g per day for 6 consecutive weeks if patients encounter any of the following side effects: swollen gums, dry or sore throat, ulcers in oral cavity, local eczema (perioral or alinasal). Patients in control groups do not receive any intervention. Subjects had their blood drawing at baseline, 4 or 6 weeks after initiation of the study.
一旦诊断为地中海贫血症并参加研究，患者将被要求停止服用含有铁和叶酸的药物。 治疗组患者连续4周每天口服阿胶（山东东阿阿胶有限公司）15克。 如果患者出现下列任何副作用：口腔肿胀，干燥或喉咙痛，口腔溃疡，局部湿疹（口周或鼻腔），剂量年龄调整为每天10克，连续6周。 对照组患者不接受任何干预。 受试者在研究开始后的基线数据和4或6周后抽血化验。
Evaluation of effectiveness
10 mL of blood were drawn after overnight fasting for at least 8 h next early morning before and after treatment for following analysis:
(1) level of Hb was measured by automated hematology analyzerXE5000 (Sysmex, Japan);
(2) levels of adult hemoglobin (HbA), minor adult hemo-globin (HbA2) and fetal hemoglobin (HbF) in periph-eral blood by automated capillary electrophoresis (Sebia, France);
(3) level of serum iron (SI) by full automatic bio-chemistry analyzer cobas8000 (Roche, Germany);
(4) level of serum ferritin (SF) by full automatic immune analyzer I 2000 SR (Abbott, USA).
（4）全自动免疫分析仪I 2000 SR（Abbott，美国）测定血清铁蛋白（SF）水平。
Evaluation of safety
Any symptoms that may be caused by adverse effect were recorded throughout the treatment. Peripheral blood was drawn from subjects before and after treatment to moni-tor changes of blood cell compositions and function of liver and kidney: total white blood count [WBC (×109/L)]; platelet count (×109/L); the percentage of neutrophil (%); serum alanine aminotransferase (U/L); serum aspartate aminotransferase (U/L); serum urea nitrogen (mmol/L); serum creatinine (μmol/L).
在整个治疗过程中记录可能由不良作用引起的任何症状。 从治疗前后的受试者抽取外周血，监测血细胞组成和肝肾功能的变化：总白细胞计数[WBC（×109 / L）];血小板计数（×109 / L）;中性粒细胞百分比（％）;血清丙氨酸转氨酶（U / L）;血清天冬氨酸转氨酶（U / L）;血清尿素氮（mmol / L）;血清肌酐（μmol/ L）。
Sample size calculation and patient randomization
Calculation of sample size to detect statistical signifi-cance was based on the study by Xu and colleagues where 3 month treatment of CCA led to increase of Hb level by
66.9 % . The estimated change of Hb level after 4 weeks of CCA is 20 % and null hypothesis is rejected if α value
<0.05 (power, 0.90; loss of follow-up rate, 0.2). In total 72 patients were included in the final study and patients were randomized to either treatment or the control group in a 2:1 ratio by statistical package for social sciences (SPSS) 21.0 random number generator.
样本大小的计算以检测统计显着性是基于徐和其同事的研究，其中服用阿胶3个月导致Hb水平升高66.9％。 因此预计服用阿胶4周血红蛋白水平的变化为20％，如果α<0.05，则零假设被拒绝（统计功效，0.90;失访率，0.2）。 总共72名被试者被纳入最终研究，通过社会科学统计软件包（SPSS 21.0）随机数生成器以2：1的比例随机分配被试者到治疗组或对照组。
Statistical analysis was performed by SPSS 21.0. Continu-ous variables are expressed as mean ± standard deviation. Categorical variables are expressed as n (%). Homogene-ity of variance analysis was performed. Group comparison was conducted with independent t test. Enumeration data was present as rates or proportions and analyzed with Chi-square test. P < 0.05 was considered statistically significant.
统计分析由SPSS 21.0进行。 连续变量表示为平均值±标准差。 分类变量表示为n（％）。 进行方差齐性分析。 组间比较采用独立t检验。枚举数据以比率或比例存在，并用卡方检验进行分析。 P <0.05被认为有统计学显著意义。
Fig. 1 Depiction of study procedure
Eighty-one patients were recruited and 76 were included in the final study (Fig. 1). Demographic characteristics were shown in Table 1. No significant differences were detected between the control and treatment groups in terms of preg-nancy history, body mass index (BMI) and gestational week at the time of enrollment. Age of both groups ranged from 23 to 32 years, with an average age of 28.11 years. Gestational age ranged between 25 and 33 weeks with the average of 28.69 weeks. All subjects are married Chinese Han female without history of stillbirth.
最终研究招募了81名患者，其中76名被纳入研究（图1）。 表1显示了人口特征。 对照组和治疗组之间在妊娠史，体重指数（BMI）和入选时的孕周方面没有显着差异。 两组年龄介于23至32岁之间，平均年龄为28.11岁。 胎龄介于25至33周之间，平均为28.69周。 所有科目都是已婚中国汉族女性，没有死胎史。
Table 1 The demographic and clinical characteristics at baseline
Change of level and proportion in Hb before and after treatment
We examined the level of Hb and the proportion of each Hb component in subjects’ peripheral blood before and after treatment. As shown in Table 2, before treatment, subjects from both groups had similar levels of Hb and Hb compo-nent. Therefore, the subjects did not differ in their severity of anemia and thalassemia. After receiving oral CCA for 4 weeks, patients experienced a significant increase of Hb concentration by an average 9.96 ± 2.75 g/L. On the other hand, the Hb concentrations in the control group dropped by 4.54 ± 2.14 g/L from baseline when no intervention was given. Level of HbA increased significantly after treat-ment while a simultaneous decrease of HbA2 and HbF was observed in subjects treated with CCA. The changes of different Hb components showed an opposite trend in control group where HbF and HbA2 increased while HbA decreased from baseline. Between treatment and control groups, the differences of three Hb components differed substantially after treatment (P < 0.001).
我们在治疗前后检查了受试者外周血中Hb水平和每种Hb组分的比例。 如表2所示，治疗前，两组受试者的Hb和Hb成分水平相似。 因此，受试者的贫血和地中海贫血的严重程度没有差异。 口服阿胶4周后，患者Hb浓度显着增加平均值9.96±2.75 g / L。另一方面，当没有干预时，对照组的Hb浓度比基线下降了4.54±2.14 g / L。在治疗后HbA水平显着增加，同时在用阿胶治疗的受试者中观察到HbA2和HbF同时降低。不同Hb组分的变化在对照组中呈现相反的趋势，其中HbF和HbA2增加，而HbA从基线降低。 在治疗组和对照组之间，治疗后三种Hb组分的差异显着不同（P <0.001）。
Table 2 The difference of concentration and component of Hb and iron metabolism indexes before and after treatment (standard deviation)
Effect on iron metabolism
As shown in Table 2, treatment of CCA increased SI while decreasing the level of SF in the subjects. In the control group, both SI and SF dropped slightly at the end of the study. Between treatment and control group, the change of SI was significant different after treatment of CCA while SF remained unaffected regardless of the treatment.
如表2所示，阿胶治疗增加了SI，同时降低了受试者的SF水平。 在对照组中，SI和SF在研究结束时略有下降。 在治疗组和对照组之间，在治疗阿胶后SI的变化有显着差异，而SF无论治疗如何都不受影响。
Among the treatment group, 78.0 % (39/50) of subjects completed the full course of 4-week treatment while the rest changed to 6-week treatment scheme due to inflamma-tory side effects including sore throat, swollen gums, ulcers in oral cavity and local eczema (perioral or alinasal). Two patients suffered from unbearable inflammatory side effects and discontinued the intervention. No subjects experienced any of the following severe adverse events: systemic rash and throat edema. No significant change was detected in WBC, platelet count, and liver function between control and treatment group (data not shown).
在治疗组中，78.0％（39/50）的受试者完成了4周的全部疗程，而其余的改为6周的治疗方案，因为炎症副作用包括喉咙痛，牙龈肿胀，口腔溃疡腔和局部湿疹（口周或鼻翼）。 两名患者出现难以忍受的炎症副作用，并停止治疗。 没有受试者出现任何以下严重不良事件：系统性皮疹和咽喉水肿。 WBC，血小板计数和肝功能在对照组和治疗组之间没有显着变化（数据未显示）。
Thalassemia is a genetic abnormality that manifests hemo-globin, the major oxygen binding protein in red blood cells. The severity of the disease depends on the type and loca-tion of the mutation. Normal tetrameric Hb is consisted of two non-α chain (β, δ, γ) and two α chain. The composi-tions of the four globin chains determine the type of Hb. There are three major types of Hb: (1) adult hemoglobin (HbA), consisted of two α chains and two β chains (α2β2), which account for more than 95 % of total Hb; (2) minor adult hemoglobin (HbA2), consisted of two α chains and two δ chains (α2δ2), accounting for 2–3 % of total Hb; (3) fetal hemoglobin (HbF), consisted of two α chains and two γ chains (α2γ2), accounting for 1–2 % of total Hb .
β-Thalassemia is characterized by mutations in β-globin which result in reduced synthesis or complete loss of β chain. As a result, the relative amount of α chain as well as the compensatory synthesis of γ and δ chain increases, leading to elevated level of HbF and HbA2. The excess globin chain polymer further deposit on the membrane of red blood cell (RBC) and impair the mechanical stabil-ity and transformation ability by inducing immunological damage and reactive oxygen species. Eventually patients suffer from hemolysis and ineffective hematopoiesis [8, 9]. Studies in the last few decades have demonstrated that the imbalanced synthesis of Hb globin chain and abnormal Hb compositions are the major causes of thalassemia and therefore it is essential to target these mechanisms for bet-ter clinical management [8, 9].
地中海贫血是一种基因异常，表现在血红蛋白，即红细胞中的主要氧结合蛋白。 疾病的严重程度取决于突变的类型和位置。 正常四聚体Hb由两条非α链（β，δ，γ）和两条α链组成。 四种珠蛋白链的组成决定了Hb的类型。 Hb主要有三种类型：（1）成人血红蛋白（HbA），由两条α链和两条β链组成（α2β2），占总Hb的95％以上; （2）轻微成人血红蛋白（HbA2）由两条α链和两条δ链（α2δ2）组成，占总血红蛋白的2-3％; （3）胎儿血红蛋白（HbF），由两条α链和两条γ链（α2γ2）组成，占Hb总数的1-2％。
β-地中海贫血的特征在于β-珠蛋白的突变，导致β链的合成减少或完全丧失。 结果，α链的相对量以及γ和δ链的代偿合成增加，导致HbF和HbA2的水平升高。 过量的珠蛋白链聚合物进一步沉积在红细胞（RBC）膜上，并通过诱导免疫损伤和活性氧损害机械稳定性和转化能力。 最终患者会出现溶血和无效的造血作用[8,9]。 过去几十年的研究表明，Hb珠蛋白链的不平衡合成和异常Hb组成是导致地中海贫血的主要原因，因此将这些机制作为更好的临床管理的基础是至关重要的[8,9]。
Despite of the progress in thalassemia management, no consensus has been reached in treating pregnant thalas-semia patients due to lack of safe and effective treatment, which present a major challenge to gynecologists and obstetricians. Therapies targeted at genetic mutations have captured much attention in the field of thalassemia. Early studies indicated that gene-targeting drugs can significant improve hemolytic anemia in patients with thalassemia. Currently the two major categories of drugs that regulate gene expression of globin chains are activators of γ chain and inhibitor of α chain. The former includes 5-azacy-tidine, hydroxyurea and other antineoplastic agents while the latter is consisted of anti-tuberculosis drugs like isonia-zid. Both categories are potentially teratogenic and carci-nogenic, rendering them unsuitable for pregnant patients. Besides, severe side effect like bone marrow suppression is another concern for clinicians to apply these drugs because fetus and newborns are at high risk of drug exposure through placenta and breast milk . Transfusion remains the only effective means for treating severe anemia during pregnancy. However about 60–80 % of patients with thalas-semia requires blood transfusion during pregnancy, with half of the populations never receiving any blood transfu-sion before. Therefore, an immune response may be elicited and cause massive production of anti-RBC antibodies in thalassemia pregnant women, exposing them to higher risk of autoimmune hemolytic anemia. In severe cases, patients may suffer from a vicious cycle of hemolytic anemia and repeated transfusion in addition to the danger of contract-ing infections disease and iron overloading [2, 11].
尽管在地中海贫血管理方面取得了进展，但由于缺乏安全有效的治疗，尚未就治疗怀孕的地中海贫血患者达成共识，这对妇产科医生和产科医生构成了重大挑战。 针对基因突变的疗法在地中海贫血领域引起了很多关注。 早期研究表明，基因靶向药物可显着改善地中海贫血患者的溶血性贫血。 目前调节珠蛋白链基因表达的两大类药物是γ链激活剂和α链抑制剂。 前者包括5-氮胞苷，羟基脲和其他抗肿瘤药物，后者则由抗结核药物如异烟肼组成。 这两个类别都有可能致畸和致癌，使其不适合孕妇。 此外，骨髓抑制等严重副作用是临床医生应用这些药物的另一个关注点，因为胎儿和新生儿通过胎盘和母乳暴露于高风险的药物暴露。 输血仍然是治疗妊娠期间严重贫血的唯一有效手段。 然而，大约60-80％的地中海患者在怀孕期间需要输血，其中有一半的人口从未接受任何输血。 因此，地中海贫血孕妇可能会引发免疫应答并引起抗RBC抗体的大量产生，从而使他们面临更高的自身免疫性溶血性贫血的风险。 在严重的病例中，患者可能遭受溶血性贫血和反复输血的恶性循环，此外还有感染疾病和铁过载的危险[2,11]。
Compared to conventional medicine where synthetic chemicals play an important role, TCM has the advantage of low toxicity and cost. Therefore, TCM is a lucrative resource for future exploration in treating thalassemia with broad prospects of clinical application. TCM practitioners classify thalassemia as a type of “blood deficiency” that is caused by both genetic and environmental factors like malnutrition and underdevelopment, the pathogenesis of which involves to the deficiency of qi, blood, yin, yang. The so-called “blood deficiency” in TCM is best defined as insufficient hematopoiesis in conventional medicine . Accord-ing to the theory of TCM, pregnant women are prone to ‘blood deficiency’ due to the physiological changes during pregnancy, similar to the aggravation of symptoms in preg-nant thalassemia patients .
与合成化学药品发挥重要作用的传统药物相比，中药具有毒性低，成本低的优点。 因此，中医治疗地中海贫血具有广阔的临床应用前景，是今后探索治疗地贫的有利资源。 中医从业人员将地中海贫血归类为由于营养不良和发育不良等遗传因素和环境因素共同造成的“血虚症”，其发病机制涉及气血，阴，阳不足。 这个中医学中所谓的“血虚症”最好定义为常规医学中的造血不足。 根据中医理论，孕妇因怀孕期间的生理变化而容易出现“血虚症状”，类似于地贫患者症状加重。
CCA, as one of the well-known TCM, has been used for treating insufficient hematopoiesis for more than 2000 years. Its therapeutic efficacy has been demonstrated in various hematological diseases including iron-deficiency anemia, aplastic anemia and thrombocytopenia. Clinical studies also showed that it can increase blood cell count and reduce bleeding from threatened abortion . With the development of modern pharmacology, several key com-ponents have been purified from CCA including collagen protein, amino acids, polysaccharides, volatile substances and inorganic substances . The hematopoietic effect and mechanism of CCA were investigated in anemic mice sepa-rately induced by 5-fluorouracil or cyclophosphamide [14, 15]. The results indicated that oral administration of CCA could activate the erythrocyte progenitor cells in bone mar-row and subsequently increase the percentage of periph-eral reticulocytes, eventually leading to the recovery of red blood cell counts and Hb concentration. By examining the expression level of granulocyte macrophage colony stimu-lating factor (GM-CSF) and erythropoietin (EPO) in the kidney and liver of the anemic mice, the investigators fur-ther confirmed that CCA treatment significantly increased serum GM-CSF and EPO level, which suggested that the hematopoietic effect of major components in CCA was partly due to promotion of the proliferation and differentia-tion of hematopoietic stem cells through stimulating GM-CSF and EPO secretion [4, 15]. These potential effect may explain the underlying mechanisms of CCA in the allevia-tion of thalassemia.
作为着名的中医药之一，阿胶已被用于治疗超过2000年的造血功能不足。 其疗效已被证明在包括缺铁性贫血，再生障碍性贫血和血小板减少症在内的各种血液疾病中。 临床研究还表明，它可以增加血细胞计数，减少先兆流产的出血。 随着现代药理学的发展，已经从阿胶中提纯了几种主要成分，包括胶原蛋白，氨基酸，多糖，挥发性物质和无机物。 在5-氟尿嘧啶或环磷酰胺分别诱导的贫血小鼠中研究阿胶的造血作用和机制[14,15]。 结果表明，口服阿胶可以激活骨髓中的红细胞祖细胞，并随后增加外周网织红细胞的百分比，最终导致红细胞计数和Hb浓度的恢复。 通过检测贫血小鼠的肾脏和肝脏中粒细胞巨噬细胞集落刺激因子（GM-CSF）和促红细胞生成素（EPO）的表达水平，研究人员进一步证实阿胶治疗显着增加了血清GM-CSF和EPO这提示阿胶中主要成分的造血作用部分是由于通过刺激GM-CSF和EPO分泌来促进造血干细胞的增殖和分化[4,15]。 这些潜在的影响可能解释阿胶在地中海贫血的缓解机制中的作用。
In the past, majority of the studies on treating thalas-semia by TCM was restricted to mainly case report and observational studies. Quantitative studies in pregnant patients with systematic intervention are lacking, imped-ing greater usage of CCA in this particular population . Our results suggest that powdered CCA alone can improve Hb levels and optimize Hb components without affecting iron reserves. SI is a common indicator of functional iron but can be easily influenced by diet and acute inflammation. Therefore, it is considered as an unreliable parameter for evaluating iron status. On the other hand, SF is an indica-tor of iron reserve and the most commonly used biomarker of iron status in pregnancy . We observed an increase of SI in the treatment group which may be due to the rich iron contained in CCA . It is of clinical importance to maintain a balanced iron metabolism because treatment-induced iron overloading is a severe issue in patients with thalassemia. A small fraction of patients experienced minor adverse events including sore throat and oral ulcers. All the side effects can be managed and alleviated via reducing the dosage and prolonging the treatment duration.
过去，大多数中医药治疗地中海贫血的研究主要限于病例报告和观察性研究。缺乏有系统干预的孕妇的定量研究，阻碍了在这一特定人群中更多地使用阿胶 。 我们的研究结果表明，粉末阿胶单独可以提高血红蛋白水平，优化血红蛋白组分，而不影响铁储备。 SI是功能性铁的常见指标，但易受饮食和急性炎症的影响。 因此，它被认为是评估铁质状态的不可靠参数。 另一方面，SF是铁储备指标，也是妊娠期铁最常用的生物标志物。 我们观察到治疗组SI增加，这可能是由于阿胶中含有丰富的铁。 维持铁代谢平衡具有临床重要性，因为治疗性铁超载是地中海贫血患者的严重问题。 一小部分患者出现轻微不良事件，包括喉咙痛和口腔溃疡。 所有的副作用都可以通过减少剂量和延长治疗时间来控制和缓解。
Recent studies have reported that imbalance of glo-bin chain synthesis and abnormal compositions of Hb can largely affect the severity of thalassemia. Moreover the levels of HbF and HbA are closely linked to adverse preg-nancy outcome in women. It has been demonstrated that elevated HbF level is associated with high rate of spontane-ous abortion in thalassemia patients. One plausible expla-nation is that compared to the other two adult hemoglobins (HbA and HbA2), HbF has a higher affinity for oxygen, causing reduced dissociation of oxygen at local tissue and cells and subsequent local hypoxia. Thus it is speculated that by reducing the level of HbF, one can prevent adverse pregnancy outcome, particularly miscarriage by improving oxygen supply to the embryo . Our results suggest that oral administration of CCA not only significantly increase the level of HbA, but also marginally decrease the level of HbF and HbA2. The total improvement in HbA is com-parable to the total decline in HbA2 and HbF. However, the changes in HbA2 percentages and HbF percentages are only modest and likely within the range of laboratory variance, possibly caused by the insufficient sample size. The impact on HbF and HbA2 should be further verified in future study with larger sample size.
最近的研究报道，血红蛋白链合成的不平衡和血红蛋白的异常组成可以很大程度上影响地中海贫血的严重程度。 此外，HbF和HbA水平与女性不良妊娠结局密切相关。 已经证明，HbF水平升高与地中海贫血患者的自发性流产率高有关。 一种合理的解释是，与其他两种成人血红蛋白（HbA和HbA2）相比，HbF对氧有更高的亲和力，导致局部组织和细胞中氧的分解减少以及随后的局部缺氧。 因此据推测，通过降低HbF水平，可以预防不良的妊娠结局，特别是通过改善胚胎供氧的流产。 我们的研究结果表明，口服阿胶不仅会显着提高HbA水平，还会轻微降低HbF和HbA2的水平。 HbA的总体改善与HbA2和HbF的总体下降相当。然而，HbA2百分比和HbF百分比的变化只是适度的，可能在实验室变异的范围内，这可能是由样本量不足造成的。 对于HbF和HbA2的影响应在未来研究中进一步验证，并且样本量较大。
More importantly, our data demonstrated that CCA influences the level of HbA and HbF via mechanism that differs significantly from the conventional γ chain gene activator. The major role of γ chain gene activator is to pro-mote synthesis of γ chain in order to replace the defective β chain so that the excess α chain can form HbF with the γ chain because the newly form HbF can partially com-pensate for the loss of HbA . While this can heighten the total Hb, the increment in HbF also expose the patients to higher change of miscarriage. Other therapies includ-ing erythropoietin increased HbF substantially . On the contrary, CCA improves anemia in β-thalassemia by enhancing HbA but not HbF, although the exact molecular signaling pathway remains unclear. We suspect that CCA enhances total Hb mainly through induction of β-globin chain. As a result the compensatory synthesis of γ and δ chain is reduced, leading to higher level of HbA (α2β2) but lower levels of HbF (α2γ2) and HbA2 (α2δ2). This specu-lation should be confirmed in future study when more in-depth genetic study is coupled with biomarker analysis in order to provide a solid scientific basis for the treatment of anemia in pregnant women with thalassemia.
更重要的是，我们的数据表明，阿胶通过与常规γ链基因激活剂显着不同的机制影响HbA和HbF的水平。 γ链基因激活剂的主要作用是促进γ链的合成，以取代有缺陷的β链，使得过量的α链可以与γ链形成HbF，因为新形成的HbF可以部分补偿失去HbA 。 虽然这可以提高总Hb，但HbF的增加也使患者暴露于更高的流产变化。 其他疗法包括促红细胞生成素可显着增加HbF 。 相反，阿胶通过增强HbA而不是HbF改善了β地中海贫血的贫血，尽管确切的分子信号传导途径仍不清楚。 我们怀疑阿胶主要通过诱导β-珠蛋白链增强总Hb。 结果γ和δ链的代偿合成减少，导致HbA（α2β2）水平升高，但HbF（α2γ2）和HbA2（α2δ2）水平降低。 这种推测应该在未来的研究中得到证实，当更深入的遗传学研究与生物标志物分析相结合时，为地中海贫血孕妇贫血的治疗提供坚实的科学依据。
In summary, CCA significantly improves anemia symp-toms without alternating iron reserves in β-thalassemia patients or inducing severe side effect. The therapeutic effect is mainly mediated through simultaneous increase of HbA in β-thalassemia patients respectively. It may also exert its effect via reducing HbF and HbA2 which requires further study to confirm. This study demonstrates the potential of a single ingredient from TCM for treat-ing genetic disorder in pregnant patients. It brings insight to obstetricians and gynecologists to the potential of CCA in treating anemia in pregnant patients with β-thalassemia where optimization of Hb components holds the key for disease control.
总之，阿胶显着改善贫血症状，而不改变β-地中海贫血患者的铁储备或诱发严重的副作用。 治疗效果主要是通过同时增加β地中海贫血患者HbA介导的。 它也可能通过减少HbF和HbA2发挥作用，这需要进一步的研究来证实。 这项研究证明了来自TCM的单一成分用于治疗怀孕患者遗传性疾病的潜力。 它使产科医生和妇科医生了解阿胶治疗妊娠期β地中海贫血患者贫血的潜力，其中Hb组分优化是疾病控制的关键。
Compliance with ethical standards
Conflict of interest The authors have no conflicts of interest.
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