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AbstractContext 5-Fluorouracil (5-FU)-injured stromal cells may cause chronic bone marrow suppression; however, the underlying mechanism remains unclear. Angelica sinensis polysaccharide (ASP), the main biologically active ingredient of the Chinese herb, Angelica sinensis (Oliv.) Diels (Apiaceae), may enrich the blood and promote antioxidation.Objective This study investigated the protective antioxidative effects of ASP on perivascular mesenchymal progenitors (PMPs) and their interactions with hematopoietic cells.Materials and methods PMPs were dissociated from C57BL/6 mouse femur and tibia and were subsequently divided into the control, ASP (0.1 g/L), 5-FU (0.025 g/L), and 5-FU + ASP (pre-treatment with 0.1 g/L ASP for 6 h, together with 0.025 g/L 5-FU) then cultured for 48 h. Hematopoietic cells were co-cultured on these feeder layers for 24 h. Cell proliferation, senescence, apoptosis, and oxidative indices were detected, along with stromal osteogenic and adipogenic differentiation potentials. Intercellular and intracellular signaling was analyzed by real-time quantitative reverse transcription polymerase chain reaction and Western blotting.Results ASP ameliorated the reactive oxygen species production/scavenge balance in PMPs; improved osteogenic differentiation; increased SCF, CXCL12, VLA-4/VCAM-1, ICAM-1/LFA1, and TPO/MPL, Ang-1/Tie-2 gene expression. Further, the ASP-treated feeder layer alleviated hematopoietic cells senescence (from 21.9 ± 1.47 to 12.1 ± 1.13); decreased P53, P21, p-GSK-3β, β-catenin and cyclin-D1 protein expression, and increased glycogen synthase kinase (GSK)-3β protein expression in co-cultured hematopoietic cells.Discussion and conclusions ASP delayed oxidative stress-induced premature senescence of 5-FU-treated feeder co-cultured hematopoietic cells via down-regulation of overactivated Wnt/β-catenin signaling. These findings provide a new strategy for alleviating myelosuppressive stress.

ABSTRACTObjectives: High reactive oxygen species (ROS) levels lead to cell death, and the testes are among the most vulnerable organs to oxidative damage. Rg1, an active ingredient extracted from the natural medicine ginseng, has potential anti-inflammatory, antioxidant and antiapoptotic properties. Our previous studies showed that Rg1 can effectively improve spermatogenic function in mice, but the specific mechanism remains unclear. The purpose of this study was to investigate the effect of Rg1 on oxidative stress and spermatogonium apoptosis in D-gal-induced testicular toxicity and elucidate the associated mechanism.Methods: Male C57BL/6 mice at 6–8 weeks of age were intraperitoneally injected with D-gal (200 mg/kg) for 42 days to establish a testicular injury model, and on day 16, 40 mg/kg Rg1-rich saline was injected intraperitoneally. Concurrently, we established an in vitro model of D-gal-damaged spermatogonia, which was treated with Rg1.Results: We found that treatment with the ginsenoside Rg1 reduced D-gal-induced oxidative stress and spermatogonium apoptosis in vivo and in vitro. Mechanistically, we found that Rg1 activated Akt/bad signaling and reduced D-gal-induced spermatogonium apoptosis.Discussion: We provide evidence showing that the antioxidant effect of Rg1 is mediated by the Akt/GSK-3β/NRF2 axis. Based on these findings, we consider Rg1 a potential treatment for testicular oxidative damage.

B-lymphocytopenia among myelosuppression is the most intractable side effect of chemotherapy. Here, we investigated ways to alleviate 5-fluorouracil-caused stress hematopoietic impairment. We found that intraperitoneally injected ASP (Angelica sinensis polysaccharides) (100 mg/kg per day), one main active ingredient of Angelica sinensis, for consecutive 7 days, significantly recovered mouse bone marrow pro-B and pre-B cells, reversed the capacity of CFU-PreB colony forming, thus alleviating B cell reduction in the spleen and peripheral blood, as well as ameliorating immunoglobin from spleen and serum. The mechanism is related to the protective effects of ASP on IL-7 producing cells, including perivascular Leptin+ and CXCL12+ mesenchymal stem and progenitor cells (MSPCs), thus promoting IL-7 production, and activating IL-7R-mediated STAT5, PI3K-AKT signaling, including survival signals and EBF1, PAX5 transcription factor expression. Additionally, ASP’s IL-7 promoting effect was demonstrated to be associated with maintaining osteogenesis/adipogenesis balance of MSPCs via the NRF2 antioxidant pathway. Collectively, our findings indicate that ASP reverse stress B-lymphocytopenia via improving Nrf2 signaling, promoting IL-7 production in MSPCs, and subsequently maintaining survival, proliferation, and differentiation of B cell progenitors, which may represent a promising therapeutic strategy.

Abstract Hematopoietic stem cells show biological manifestations of aging, diminished hematopoietic function and abnormal differentiation, which can lead to leukemia. It is therefore important to explore the mechanism underlying hematopoietic stem cell aging to develop strategies for delaying the process. Our evaluations revealed that the number of bone marrow hematopoietic cells (BMHCs) started to decrease significantly after 45 years of age, and the number of senescent BMHCs, as determined by senescence-associated beta-galactosidase staining, gradually increased with age. In addition, BMHCs from individuals over 45 years of age presented with notably reduced proliferative capacity, increased G1-phase cell cycle arrest, and significantly decreased generation of mixed colony forming units, which suggests that BMHCs enter senescence during middle age. Furthermore, we observed significantly lower antioxidant capacity and a significant increase in oxidative damage products, a gradual increase in the expression of senescence-associated proteins and genes, and a gradual decrease in the expression of cell cycle related proteins in BMHCs after middle age. Taken together, these findings offer both a theoretical and experimental basis for better understanding of the senescence progression of BMHCs and the optimal timing for anti-senescence drug interventions in clinical practice.

Angelica Sinensis polysaccharide (ASP), the main active component of Angelica sinensis, possesses antioxidative and anti-apoptotic properties. In this study, we have investigated the antagonistic effect of ASP on 5-FU-induced injury of mouse spleen in vivo and splenocytes in vitro, and its possible mechanism. Our results showed that ASP inhibited 5-FU-induced decreases in spleen weight and organ index in mice, restored the number of peripheral blood leukocytes and lymphocytes, repaired spleen structure disorder and functional impairment, rescued serum IL-2, IL-6, and IFN-γ levels, and relieved 5-FU-induced mitochondrial swelling， reduced the oxidant accumulation including MDA and ROS, whereas increasing the activities of GSH, SOD and CAT. The mechanism may be related to ASP downregulation of Keap1 protein expression thus motivating the nuclear translocation of Nrf2. Furthermore, ASP alleviated the apoptosis of spleens in vivo and splenocytes in vitro, and reactivated PI3K / AKT signalling. In conclusion, the protective effect of ASP on spleens and splenocytes may be related to the reduction of oxidative stress and apoptosis via reactivation of Nrf2 and PI3K/AKT pathways. This study has provided a new protective agent for minimizing the spleen injury caused by 5-FU and a new idea for improving the prognosis of chemotherapy patients.

Oxidative stress induced by chemotherapeutic agents causes hepatotoxicity. 5-Fluorouracil (5-FU) has been found to have a variety of side effects, but its toxic effect on the liver and the mechanism are still unclear. Angelica polysaccharide (ASP), the main active ingredient of Dang Gui, has antioxidative stress effects. In this study, we investigated the antagonistic effects of ASP on 5-FU-induced injury in the mouse liver and human normal liver cell line MIHA and the possible mechanism. Our results show that ASP inhibited 5-FU-induced the decrease in Bcl-2 protein and the increase in Bax protein. ASP alleviated 5-FU-induced the increase in alanine aminotransferase (ALT), triglyceride (TG), and aspartate aminotransferase (AST) content; hepatic steatosis; and liver fibrosis. ASP restored 5-FU-induced swelling of mitochondria and the endoplasmic reticulum. 5-FU promoted the expression of Keap1 and increased the binding to NF-E2-related factor 2 (Nrf2) to reduce the nuclear translocation of Nrf2, thereby weakening the transcriptional activity of Nrf2 to inhibit the expression of HO-1; reducing the activity of GSH, SOD, and CAT to increase ROS content; and aggravating DNA damage (indicated by the increase in 8-OHdG). However, ASP reversed these reactions. In conclusion, ASP attenuated the 5-FU-induced Nrf2 pathway barrier to reduce oxidative stress injury and thereby inhibit the disorder of lipid anabolism and apoptosis. The study provides a new protectant for reducing the hepatic toxicity caused by 5-FU and a novel target for treating the liver injury.

Myelosuppression is the most common complication of chemotherapy. Decline of self-renewal capacity and stress-induced premature senescence (SIPS) of hematopoietic stem cells (HSCs) induced by chemotherapeutic agents may be the cause of long-term myelosuppression after chemotherapy. Whether the mechanism of SIPS of hematopoietic cells relates to chemotherapeutic injury occurred in hematopoietic microenvironment (HM) is still not well elucidated. This study explored the protective effect of Angelica sinensis polysaccharide (ASP), an acetone extract polysaccharide found as the major effective ingredients of a traditional Chinese medicinal herb named Chinese Angelica (Dong Quai), on oxidative damage of homo sapiens bone marrow/stroma cell line (HS-5) caused by 5-fluorouracil (5-FU), and the effect of ASP relieving oxidative stress in HM on SIPS of hematopoietic cells. Tumor-suppressive doses of 5-FU inhibited the growth of HS-5 in a dose-dependent and time-dependent manner. 5-FU induced HS-5 apoptosis and also accumulated cellular hallmarks of senescence including cell cycle arrest and typical senescence-associated β-galactosidase positive staining. The intracellular reactive oxygen species (ROS) was increased in 5-FU treated HS-5 cells and coinstantaneous with attenuated antioxidant capacity marked by superoxide dismutase and glutathione peroxidase. Oxidative stress initiated DNA damage indicated by increased γH2AX and 8-OHdG. Oxidative damage of HS-5 cells resulted in declined hematopoietic stimulating factors including stem cell factor (SCF), stromal cell-derived factor (SDF), and granulocyte-macrophage colony-stimulating factor (GM-CSF), however, elevated inflammatory chemokines such as RANTES. In addition, gap junction channel protein expression and mediated intercellular communications were attenuated after 5-FU treatment. Significantly, co-culture on 5-FU treated HS-5 feeder layer resulted in less quantity of human umbilical cord blood-derived hematopoietic cells and CD34+ hematopoietic stem/progenitor cells (HSPCs), and SIPS of hematopoietic cells. However, it is noteworthy that ASP ameliorated SIPS of hematopoietic cells by the mechanism of protecting bone marrow stromal cells from chemotherapeutic injury via mitigating oxidative damage of stromal cells and improving their hematopoietic function. This study provides a new strategy to alleviate the complication of conventional cancer therapy using chemotherapeutic agents.