Search

The major feature that distinguishes acute promyelocytic leukemia (APL) cells from other malignant hematopoietic cells is the expression of promyelocytic leukemia-retinoicacid receptor α (PML-RARα) fusion protein, which contributes to the inhibition of RARα-regulated hematopoietic cellular differentiation. The complete remission rate of APL exceeded 90% with the application of all-trans retinoic acid (ATRA), arsenic trioxide (ATO), and anthracycline-based chemotherapy. However, the 7-year cumulative incidence of relapses was reported as 28.6% in APL maintenance with ATRA and daunorubicin and even reached 33% in the ATRA maintenance treatment group. The relapse/refractory patients showed resistance to ATRA and/or ATO, which has been identified as a clinically significant problem. Currently, no effective drugs are available to reverse the ATRA resistance. Matrine (MAT) is the main active component of Sophora flavescens. It has been used to treat chronic hepatitis for several years in China. Recently, the molecule has been proven to exhibit an anti-leukemic effect. Previous studies have revealed that MAT can reverse the ATRA resistance of NB4-LR1 cells when coupled with ATRA. The treatment with 0.1mmol/L MAT and 1μmol/L ATRA can restore the ability of NB4-LR1 cells to differentiate, which might be related to the increased level of cyclic adenosine monophosphate and protein kinase A activity in NB4-LR1 cells, reduced telomerase activity and downregulated expression of topoisomerase II beta (TopoIIβ) (Wu, et al. Planta Med 2014). Subsequently, this study aimed to investigate the mechanism underlying the degradation of the PML/RARα fusion protein in the presence of MAT and ATRA in NB4 and NB4-LR1 cell lines. ATRA-sensitive (NB4) and ATRA-resistant (NB4-LR1) cell lines were used. Nitroblue tetrazolium reduction assay and flow cytometry were used to detect the differentiation ability. The activity of ubiquitin-proteasome and autophagy-mediated pathways in both cells treated with ATRA with or without MAT were compared in protein and mRNA level (Western blot analysis, qRT-PCR), the Fluorescent substrate Suc-LLVY-AMC detection was used to detect the activity of proteasome, and electron microscope for observing autophagosome. MG 132(proteasome inhibitor), rapamycin (autophagy activator, RAPA), hydroxychloroquine (lysosomal inhibitor, HCQ) and STI571 [retinoic acid receptor alpha (RARα) ubiquitin stabilizer] were used as positive controls. The effect of MAT was observed in vivo using xenografts. Results showed that MAT improved the sensitivity of NB4-LR1cells to ATRA treatment, which was consistent with the expression of PML-RARα fusion protein (Fig.A-C).The ubiquitin proteasome pathway plays a crucial role in protein degradation. MAT promoted the ubiquitylation level in NB4-LR1 by stabilized the 20S protein expression and enhanced the activity of the proteasome (Fig.D). ATRA inhibited the expression of RARα in NB4-LR1 cells, which was contradictory to that in NB4 cells. MAT can stabilize the expression of RARα in NB4-LR1 cells, whereas MG132 downregulated the expression of RARα in both cell lines, which hampered the differentiation of NB4 cells (Fig.E-G). In addition to UPP, the autophagy pathway also had a significant role in arsenious acid- or ATRA-mediated PML-RARα fusion protein degradation. MAT could promoted the autophagy in NB4-LR1 cells, with an increase in microtubule-associated protein 1 light chain3 (LC3)-II and LC3-II/LC3-I ratio and exhaustion of P62 (Fig.H-K). A similar phenomenon was observed in mouse xenografts (Fig.L-N). In summary, the present study revealed the difference in the chain reaction of sensitive and resistant APL cell lines (NB4 and NB4-LR1, respectively) to the treatment of ATRA, explaining the mechanism underlying the resistance to ATRA. ATRA decrease the level of 20S core subunit and the RARα in NB4-LR1 cells, but could not activate the autophagy process. These effects were reversed by the combination of MAT. The proteasome inhibitor might hamper the RARα stabilization and hence was not advantageous for the differentiation of cells. It also induced autophagy in NB4-LR1 cells. MAT induced the activation of UPP and mediated the autophagic degradation process, which synergistically induced the degradation of PML-RARα fusion protein and promote the differentiation of NB4-LR1 cells. Figure. Figure. Disclosures No relevant conflicts of interest to declare.

To determine the effect of combined treatment with Chinese medicine (CM) and allogeneic hematopoietic stem cell transplantation (allo-HSCT) on patients with severe aplastic anemia (SAA).Eleven patients were treated with CM plus allo-HSCT. Nine patients received a conditioning regimen consisting of fludarabine (Flu), anti-thymocyte globulin (pig ALG), or anti-lymphocyte globulin (Rabbit ATG) and cyclophosphamide (CY), and two patients received pig ALG and CY. All patients were treated with Kidney (Shen)-reinforcing, blood-activating, and stasis-removing (KBS) herbal preparation beginning at 1 week before transplantation and ending at 8 weeks after transplantation. Chimerism status was assessed by analyzing short tandem repeat (STR) polymorphisms.All patients recovered hematopoietic function and none had graft failure. The median number of days required for the absolute neutrophil count (ANC) increased to0.5×10(9)/L was 15 days (12-22 days) and for spontaneous platelet recovery to20×10(9)/L without post-transplantation transfusion was 17 days (15-27 days). Nine patients were long-term survivors and achieved full donor chimerism. The overall cumulative incidence of acute graft versus host disease (GVHD) grades I-II and III-IV was 18.2% (2/11) and 9.1% (1/11), respectively. The overall accumulated incidence of chronic GVHD was 27.3% and all patients had limited chronic GVHD. At a median follow-up time of 32 months (range: 12-97 months), 9 patients were still alive. The estimated 5-year overall survival (OS) rate was 81.8%. The incidence of treatment-related mortality, 2-year post-transplantation, was 18.2%. Two patients died from GVHD after transplantation.Treatment with the KBS formulation may reduce the rate of graft failure and treatment-related mortality and improve the rate of OS in SAA patients with allo-HSCT.

To explore differences in bone marrow angiogenesis seen in aplastic anemia (AA) patients presenting with differential Chinese medicine (CM) syndrome, and to correlate these differences with clinical pathology.Thirty-five patients were enrolled, including 18 with "yang deficiency syndrome" and 17 with "yin deficiency syndrome." Bone marrow biopsies and serum were collected. Microvessel density (MVD) and positive expression of vascular endothelial-derived growth factor (VEGF) were detected by immunohistochemisty. Hypoxia inducible factor -1α (HIF-1α), and VEGF expression were assayed by enzyme-linked immunoabsorbent assay (ELISA), serum lactate dehydrogenase (LDH) was tested by enzyme method and liquid chip technology was used to detected the expression of interleukin (IL)-2, IL-4, IL-6, IL-10, interferon (IFN)-γ and tumor necrosis factor (TNF)-α.Counts for leukocytes, absolute neutrophils and platelets in "yin deficiency syndrome" were lower than those found in "yang deficiency syndrome" (P0.05). MVD and VEGF expression, and the positive rate of CD34 and VEGF in bone marrow were lower in AA, especially in "yin deficiency syndrome" (P0.01 or P0.05). "Yin deficiency syndrome" displayed decreased VEGF and LDH expression, and enhanced expression of HIF-1α as compared to "yang deficiency syndrome" (P0.05). Levels of IL-4 and IL-6 were higher in AA (P0.01), but IL-10 was decreased (P0.05). High TNF-α expression was seen in "yang deficiency syndrome" and IFN-γ expression was decreased in "yin deficiency syndrome" as compared with normals (P0.01 and P0.05, respectively).AA patients have lower MVD than normals, especially in "yin deficiency syndrome." MVD might differentially correlate to disease severity, and could be dependent on bone marrow or serum VEGF expression and LDH. Additionally, IL-2, IL-10, IL-4 and IFN-γ were negatively associated while IL-6 and TNF-α were positively associated with MVD.

Because the mechanism underlying the development of acute pancreatitis (AP) has not yet been fully clarified, it has been a hot but difficult topic in basic and clinical research for a long time. Currently, the dominant hypothesis for the pathogenesis of AP is that it is a disease of self-digestive acute chemical inflammation induced by trypsin activation. As proteins to trigger the inflammatory response cascade, Toll-like receptors (TLRs), especially TLR4, provide a new clue for studying the pathogenesis of AP from the source. Some studies have found that when TLR4 is activated by certain factors, it can amplify an inflammatory effect and aggravate the body's inflammatory response through a series of signal transduction. Toll-like receptor 4 may play an important role in the synthesis and release of proinflammatory cytokines, and the up-regulation of the TLR4 gene may be related with the development and progression of multiple organ injury during AP. As the "gate" of inflammatory response, TLR4 may be closely associated with the development and progression of multiple organ injury during AP. Understanding the roles of TLR4 in AP will help to further clarify the pathogenesis of AP and to search a new target for the treatment of AP.

One of the most common complications of acute pancreatitis is acute lung injury, during which intercellular adhesion molecule-1 (ICAM-1) plays an important role by participating in leukocyte adhesion and activation as well as by inducing the "cascade effect" of inflammatory mediators, pulmonary microcirculation dysfunction and even acute respiratory distress syndrome, multiple organ failure or death. Although it is generally believed that the modulatory mechanism of ICAM-1 during this process is associated with the activation of nuclear transcription factor kappa B which is mediated by IL-1, IL-6, IL-18 and oxygen free radical, etc., further studies are still required to clarify it. Since the upregulation of ICAM-1 expression in the lung during acute lung injury is one of main pathogeneses, the early detection of the ICAM-1 expression level may contribute to the prevention and treatment of acute lung injury. Moreover, reducing pulmonary ICAM-1 expression levels through treatment with anti-ICAM-1 monoclonal antibody (aICAM-1) and antagonists of the neurokinin 1 receptor, etc., should have a positive effect on protecting the lungs during acute pancreatitis. This review aims to further clarify the relationship between ICAM-1 and acute pancreatitis complicated by acute lung injury, and therefore provides a theoretical basis for the formulation of corresponding therapeutic measures in clinical practice for acute pancreatitis.

Aplastic anemia (AA) is a common hematologic disease characterized by hematopoietic failure of the bone marrow and pancytopenia of the peripheral blood. Some patients with AA suffer from transfusion-induced iron overload secondary to long-term blood transfusions due to moderate and severe anemia. Topics related to iron overload and chelation therapy have recently become important in hematologic diseases, especially in hematopoietic failure. However, no animal model of AA complicated by iron overload has been developed, which affects drug research and development. The purpose of our study is to establish a mouse model of aplastic anemia complicated by iron overload. We firstly controlled the total dose of iron supplements (2.0 *103 mg/kg) and compared how serum iron and ferritin levels varied among different dosage regimens to determine the best dosage and duration for the development of an iron overload model in mice (we finally chose the 200 mg/w/kg * 10 weeks). A composite model of AA was successfully established on the principle of immune-mediated bone marrow failure: DBA/2 mice were euthanized by cervical dislocation and the thymuses were taken out using aseptic techniques for the preparation of cell suspensions of 5 * 106/mL; Balb/c mouse were given whole-body irradiation with 60Co 6.0 Gy at 1 Gy/min, after which 0.2 mL of the prepared cell suspensions was injected into each mouse via the caudal vein within 4 h. We then compared the differences in liver volume, peripheral hemogram, bone marrow pathology, serum iron, serum ferritin, pathological iron deposition in multiple organs (liver, bone marrow, spleen), liver hepcidin, bone morphogenetic protein 6 (BMP6), SMAD family member 4 (SMAD4), transferrin receptor 2 (TfR2) protein, and mRNA expression levels among the Normal Control, AA, Iron Overload, and Composite Model groups to validate the composite model and to explore the pathogenesis and features of iron overload in the composite model. The results indicated significant abnormalities in iron metabolism parameters in mice with AA, which was reflected in the significant decrease of hepcidin expression in the liver (P < 0.01) that basically paralleled the changes in BMP6, SMAD4, and TfR2. Iron Overload Group had a suppressed hepcidin, BMP6, and SMAD4 mRNA expressions in liver, but these parameters were higher than in the AA group (P < 0.01). Association with iron overload would not further downregulate the negative parameters of iron deposition in mice with AA, and SMAD4 and TfR2 protein levels and hepcidin and SMAD4 mRNA expression levels were lower in the AA group than in the Composite Model group (P < 0.01 or P < 0.05). (see Fig.1 and Fig.2) The established model is basically consistent with the clinical manifestations and pathogenesis of AA complicated by transfusion-induced iron overload. This successful establishment will help in the screening of iron chelation drugs and studies on pharmacological mechanisms. Figure 1. Western-blot a nalysis of hepcidin, BMP6, SMAD4 and TfR2 expression among groups Figure 1. Western-blot a nalysis of hepcidin, BMP6, SMAD4 and TfR2 expression among groups Figure 2. Analysis of hepcidin, BMP6, SMAD4 and TfR2 mRNA expression levels among different groups Figure 2. Analysis of hepcidin, BMP6, SMAD4 and TfR2 mRNA expression levels among different groups Disclosures No relevant conflicts of interest to declare.

ObjectiveTo explore the effect of kidney-reinforcing, blood-activating and stasis-removing recipes on adhesion molecule expression of bone marrow mesenchymal stem cells (MSCs) from patients with chronic aplastic anemia (CAA).MethodsWe used three Traditional Chinese Medicine recipes, namely a kidney-reinforcing recipe (KRR), blood-activating and stasis-removing recipe (BASRR), and kidney-reinforcing, blood-activating and stasis-removing recipe (KRBASRR), and a normal saline control to prepare herbal medicine serum in Sprague Dawley rats. Thirty CAA patients were enrolled in the experimental group, including 17 kidney-Yang deficient patients and 13 kidney-Yin deficient patients. Ten healthy individuals were included in the control group. MSCs were isolated from bone marrow samples, and the cell density was observed to measure their proliferation ability by microscopy on days 2, 7, and 14 after isolation. In addition, the expression of adhesion molecules of bone marrow MSCs (CD106, CD49d, CD31 and CD44) were detected by flow cytometry after 48 h of treatment with the four different herbal medicine serums.ResultsThe proliferation of MSCs from kidney-Yang deficient and kidney-Yin deficient patients was weaker than that of MSCs from the control group. The expression of all adhesion molecules of bone marrow MSCs from CAA patients was obviously lower than that in the control group (P