Clinical Effect and Mechanism of Yisui Shengxue Granules in Thalassemia Patients with Mild, Moderate, or Severe Anemia

Yan-Ling Cheng, Xin-Hua Zhang, Yu-Wen Sun, Wen-Juan Wang, Su-Ping Fang, Zhi-Kui Wu, Yan-Ling Cheng, Xin-Hua Zhang, Yu-Wen Sun, Wen-Juan Wang, Su-Ping Fang, Zhi-Kui Wu

Abstract

Yisui Shengxue granules, which is a Chinese traditional medicine, can increase hemoglobin, red blood cells, and Ret of thalassemia patients with mild, moderate, and severe anemia and thus relieve clinical anemia symptoms. Studies on mechanism found that Yisui Shengxue granules can increase the proliferation ability of hematopoietic stem cells. Emodin promoted colony forming of hematopoietic stem cells. Yisui Shengxue granules can increase the activity of GSH-PX in bone marrow blood and decreased the severity of inclusion bodies on the cytomembrane of RBCs. YSSXG attenuated anemia symptoms in patients with thalassemia mostly by increasing the proliferation of hematopoietic stem cells and decreasing the hemolysis of RBCs.

Figures

Figure 1
Figure 1
Scatterplot of the change of hemoglobin details of each patient before and after one-, two-, and three-month treatment. (a, b, and c) Change of Hb content of each patient in mild group before and after one-, two-, and three-month treatment, respectively. (d, e, and f) Change of Hb content of each patient in moderate group before and after one-, two- and three-month treatment, respectively. (g, h, and i) Change of Hb content of each patient in severe anemia group before and after one-, two- and three-month treatment, respectively.
Figure 2
Figure 2
Change of clinical blood parameters of thalassemia patients in three groups (mild, moderate, and severe). (a) Change of Hb before and after one-, two-, and three-month treatment. (b) Change of RBC before and after one-, two-, and three-month treatment. (c) Change of Ret before and after one-, two-, and three-month treatment. ∗In mild group, compared with before treatment, ∗p < 0.05. +In moderate group, compared with before treatment, +p < 0.05; ++p < 0.01; +++p < 0.001. #In severe group, compared with before treatment, #p < 0.05; ##p < 0.01.
Figure 3
Figure 3
Proliferation ability of hematopoietic stem cell (CD34+) before and after three-month intervention with YSSXG.
Figure 4
Figure 4
Colonies formation of hematopoietic stem cells treated with emodin. CFU-E (colony forming unit-erythroid), BFU-E (burst forming unit-erythroid), CFU-GM (colony forming unit-granulocyte, macrophage), CFU-GEMM (colony forming unit-granulocyte, erythrocyte, macrophage, megakaryocyte). (a) CFU-E after 3-day incubation. (b) BFU-E and CFU-GM after 6-day incubation. (c) BFU-E and CFU-GM after 10-day incubation. (d) CFU-Mix after 13-day incubation.
Figure 5
Figure 5
Colony forming three days after treatment. Colonies were observed under an inverted phase contrast microscope; CFU-E was indicated with orange arrow. (a) CFU-E in control group (100x). (b) CFU-E in group with 4 μM emodin (100x). (c) CFU-E in group with 9 μM emodin group (100x).
Figure 6
Figure 6
Colony forming 6 days after treatment. Colonies were observed under an inverted phase contrast microscope; BFU-E was indicated with orange arrow and CFU-GM was indicated with green arrow. (a) BFU-E in control group (40x). (b) BFU-E in control group (100x). (c) BFU-E in group with 4 μM emodin (40x). (d) BFU-E in group with 4 μM emodin (100x). (e) BFU-E and CFU-GM in group with 9 μM emodin (40x). (f) BFU-E in group with 9 μM emodin group (100x).
Figure 7
Figure 7
Colony forming 10 days after treatment. Colonies were observed under an inverted phase contrast microscope; BFU-E was indicated with orange arrow and CFU-GM was indicated with green arrow. (a) BFU-E and CFU-GM in control group (40x). (b) BFU-E and CFU-GM in group with 4 μM emodin (40x). (c) BFU-E and CFU-GM in group with 9 μM emodin (40x). (d) CFU-GM in group with 4 μM emodin (100x). (e) CFU-GM in group with 9 μM emodin (100x). (f) BFU-E in group with 9 μM emodin, and it can be distinguished by the reddish or brownish color (200x).
Figure 8
Figure 8
Colony forming 13 days after treatment. (a) CFU-GEMM in control group (60x). (b) CFU-GEMM in group with 4 μM emodin (60x). (c) CFU-GEMM in group with 9 μM emodin (60x).
Figure 9
Figure 9
Benzidine staining to identification of erythroid differentiation. Benzidine staining positive cells were where the white arrows indicate, cells present with blue. Colonies were collected from a well using a blunt-ended needle, resuspended with PBS, and then stained with benzidine. Cells were observed under an inverted phase contrast microscope after 10 min. (a) Cells before benzidine staining. (b) Benzidine staining positive cells after staining (40x). (c) Benzidine staining positive cells after staining (100x). (d) Benzidine staining positive cells after staining (100x).
Figure 10
Figure 10
Comparison of changes in hemolysis-related cytokines before and after three months of treatment. (a) Comparison of changes in SOD activity. (b) Comparison of changes in MDA activity. (c) Comparison of changes in GSH-PX activity. Both bone marrow and peripheral blood of five patients were collected before and after treatment. A paired-sample t-test was used to analyze data before and after treatment, +p < 0.05.
Figure 11
Figure 11
Inclusion bodies deposition in erythrocyte membrane by transmission electron microscope (TEM) (in a 0.20 μm field of vision). Inclusion bodies were indicated with red arrow. (a)/(b)/(c) are inclusion bodied within red blood cells before treatment (magnification times, ×8000); (d)/(e)/(f) are inclusion bodies within red blood cells after three months of treatment.

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