Inflammation accelerates BCR-ABL1+ B-ALL development through upregulation of AID.

Inflammation contributes to the initiation and disease progression of several lymphoid malignancies. BCR-ABL1-positive B-cell acute lymphoblastic leukemia (BCR-ABL1+ B-ALL) is triggered by the malignant cloning of immature B cells promoted by the BCR-ABL1 fusion gene. However, it is unclear whether the mechanism driving the disease progression of BCR-ABL1+ B-ALL involves inflammatory stimulation. Here, we evaluate BCR-ABL1+ B-ALL cells' response to inflammatory stimuli lipopolysaccharide (LPS) in vitro and in vivo. The results indicate that LPS promotes cell growth and genomic instability in cultured BCR-ABL1+ B-ALL cells and accelerates the BCR-ABL1+ B-ALL development in a mouse model. We show that the LPS-induced upregulation of activation-induced deaminase (AID) is required for the cell growth and disease progression of BCR-ABL1+ B-ALL. Moreover, AID modulates the expression of various genes that are dominated by suppressing apoptosis genes and upregulating DNA damage-repair genes. These genes lead to facilitation for BCR-ABL1+ B-ALL progression. The heat shock protein 90 (Hsp90) inhibitors significantly reduce AID protein level and delay the disease progression of BCR-ABL1+ B-ALL upon inflammatory stimulation. The present data demonstrate the causative role of AID in the development and progression of BCR-ABL1+ B-ALL during inflammation, thus highlighting potential therapeutic targets.

Plasma fibrinogen, D-dimer, and fibrin degradation product as biomarkers of rheumatoid arthritis.

This study aimed to assess the association of coagulation-related indicators such as plasma fibrinogen (FIB), D-dimer, and fibrin degradation product (FDP) in rheumatoid arthritis (RA) with the disease activity. Data from 105 RA patients and 102 age- and gender-matched healthy controls were collected in the retrospective study. Disease activity score in 28 joints based on C-reactive protein (DAS28-CRP) was used to divide RA patients into low activity group (DAS28-CRP ≤ 2.7) and active group (DAS28-CRP > 2.7). Receiver operating characteristic (ROC) curve was applied to determine area under the curve (AUC). The association between plasma FIB, D-dimer, and FDP and DAS28-CRP was evaluated by spearman correlation. Logistical regression analysis was used to identify the independent variables associated with RA disease activity. RA patients showed higher levels of plasma FIB, D-dimer, and FDP than the controls (P < 0.01). Plasma FIB, D-dimer, and FDP were also increased in active groups of RA patients than those in inactive groups (P < 0.001). ROC curve analyses revealed that the AUC of D-dimer was higher than erythrocyte sedimentation rate (ESR) and rheumatoid factor (RF), and that of FDP was higher than RF in RA patients. In addition, the optimal cut-off value of plasma FIB, D-dimer, and FDP for RA diagnosis was 286 mg/dL, 470 µg/L, and 1.45 mg/L, respectively. Spearman analysis showed that plasma FIB, D-dimer, and FDP were positively related with DAS28-CRP (P < 0.001) in RA patients. Logistical regression analysis showed that D-dimer (odds ratio 2.862, 95% confidence interval 1.851-5.426, P < 0.001) was an independent variable associated with RA disease activity. FIB, D-dimer, and FDP were increased in RA patients and positively correlated with the disease activity of RA. D-dimer may act as a novel inflammatory indice for indicating disease activity in RA patients.

Involvement of Blnk and Foxo1 in tumor suppression in BCR­ABL1­transformed pro­B cells.

Oncogenic Bcr­Abl kinase mimics pre­B cell receptor (pre­BCR) survival signals in BCR­ABL1­positive B­cell acute lymphoblastic leukemia (BCR­ABL1+ B­ALL), driving B­cell progenitor malignant transformation; thus, defining a particularly unfavorable prognosis for patients. During B­cell development, pre­BCR differentiation signaling components terminate proliferative expansion and promote B­cell maturation. To study whether pre­BCR differentiation signaling components regulate the initiation and development of BCR­ABL1+ B­ALL, the tumor suppression mechanism of differentiation­related signaling molecules in BCR­ABL1­transformed pro­B cells were analyzed. The results demonstrated that Bcr­Abl kinase activated the PI3K/Akt pathway, promoting cell growth, and upregulated Aid expression, increasing genomic instability in pro­B cells. These findings suggest that Bcr­Abl kinase mediates pro­B cell malignant transformation. Furthermore, the present data revealed that BCR­ABL1 oncogenic stress triggered enhanced expression of B­cell differentiation components B­cell linker (Blnk) and forkhead box protein O1 (Foxo1) in BCR­ABL1 transformed pro­B cells. Using the CRISPR/Cas9­mediated Blnk or Foxo1 knockout BCR­ABL1­transformed pro­B cells, it was identified that, in BCR­ABL1­transformed pro­B cells, Blnk and Foxo1 reduced Bcr­Abl kinase activity to induce cell cycle arrest and decrease genomic instability. In addition, Blnk suppressed the PI3K/Akt pathway to reduce Foxo1 phosphorylation and heighten the Foxo1 activity, indicating that, in BCR­ABL1­transformed pro­B cells, Foxo1 participated in the regulation of Bcr­Abl kinase by Blnk. The present data highlighted the antitumor mechanisms of Blnk and Foxo1 in the regulation of Bcr­Abl kinase, and thus, may offer an alternative therapeutic strategy to Bcr­Abl kinase regulation in BCR­ABL1+ B­ALL.

Dentin matrix protein 1 correlates with the severity of hemorrhagic fever with renal syndrome and promotes hyper-permeability of endothelial cells infected by Hantaan virus.

Hantaviruses are the major causative agents of hemorrhagic fever with renal syndrome (HFRS) in humans, which is characterized by increased capillary permeability. Dentin matrix protein 1 (DMP1) has been shown to degrade components of the basal membrane and interendothelial junctions via matrix metalloproteinase-9. To study the changes of serum DMP1 in HFRS, we determined the concentration of DMP1 using sandwich enzyme-linked immunosorbent assay. We found that serum DMP1 concentrations increased significantly, and reached peak value during the oliguric phase and in the critical group in HFRS patients. Moreover, serum DMP1 concentrations were closely related to blood urea nitrogen, creatinine, cystatin C, and vascular endothelial growth factor (VEGF). We further explored the role of DMP1 in HTNV-infected human umbilical vein endothelial cells (HUVECs) model. Data from immunocytochemistry showed that VEGF and tumor necrosis factor-α (TNF-α) promoted the expression of DMP1 on HTNV-infected HUVECs. Results from transwell assays demonstrated that the permeability of HUVECs increased significantly after HTNV infection with the addition of DMP1, VEGF, and TNF-α. This study suggests that elevated DMP1 concentrations may be associated with disease stage, severity, and the degree of acute kidney injury. DMP1 is involved in the regulation of capillary permeability in HFRS caused by hantavirus infection.

Associations between D3R expression in synovial mast cells and disease activity and oxidant status in patients with rheumatoid arthritis.

Dopamine D3 receptor (D3R) on immune cells is involved in the pathogenesis of rheumatoid arthritis (RA). Mast cells (MCs) are currently identified as important effector cells in synovial inflammation of RA, but little is known about the role of D3R on synovial MCs in the pathogenesis of RA. Several inflammatory cells in the synovium induce reactive oxygen species (ROS) formation which are involved in the progression of RA. However, it is unclear whether D3R on synovial MCs is related to the levels of ROS in RA patients. In this study, a total of 73 patients with RA were divided into three groups according to disease activity DAS28 scores. The number of cases in group 1, group 2, and group 3 was 19, 26, and 28, respectively. We examined D3R-positive MC numbers in the synovial fluid and ROS levels in each group of RA patients, and we also analyzed the association of D3R-positive MC numbers with RA disease activity and ROS levels. MDA and protein carbonylation in the serum and synovial fluid were measured to reflect the level of lipid peroxidation and protein oxidation, respectively. Additionally, superoxide dismutase (SOD) and catalase (CAT) in the serum and synovial fluid were used to be markers of antioxidant levels. Our results showed that D3R-positive MCs in the synovial fluid showed a declining trend with the increased disease activity DAS28 score in RA patients. There was negative correlation between D3R-positive MC numbers in the synovial fluid and disease severity DAS28 score of RA patients. Moreover, D3R-positive MC numbers in the synovial fluid were negatively correlated with the level of MDA and protein carbonylation while were positively correlated with antioxidant levels such as SOD and CAT in RA patients. Our results suggested that D3R on MCs may be involved in ROS-mediated pathogenesis of RA.

Thermo-Responsive Molecularly Imprinted Hydrogels for Selective Adsorption and Controlled Release of Phenol From Aqueous Solution.

In this study, thermo-responsive molecularly imprinted hydrogels (T-MIHs) were developed as an effective potential adsorbent for selectively adsorption phenol from wastewater. During the process, N-isopropyl acrylamide (NIPAm) was used as thermal responsive monomer. The obtained materials were characterized in detail by fourier transform infrared (FT-IR) spectrometer, scanning electron microscope (SEM), and thermo gravimetric analysis (TGA). A series of static adsorption studies were performed to investigate the kinetics, specific adsorption equilibrium, and selective recognition ability of phenol. Reversible adsorption and release of phenol were realized by changing temperatures. Three type of phenols, namely 3-chlorophenols (3-CP), 2,4-dichlorophenol (2,4-DCP), and 2,4,6-trichlorophenol (2,4,6-TCP) were selected as model analytes to evaluate the selective recognition performance of T-MIHs. The T-MIHs have good selectivity, temperature response, and reusability, making them ideal in applying in the controlled separation and release of phenol pollutants.

RAG2 involves the Igκ locus demethylation during B cell development.

The genes encoding the immunoglobulin κ light chain are assembled during B cell development by V(D)J recombination. For efficient rearrangement, the Igκ locus must undergo a series of epigenetic changes. One such epigenetic mark is DNA methylation. The mechanism that the Igκ locus is selectively demethylated at the pre-B cell stage has not previously been characterized. Here, we employed bisulfite DNA-modification assays to analyze the methylation status of the Igκ locus in primary pre-B cells from RAG-deficient mice with pre-rearranged Igh knock-in allele. We observed that the Igκ locus was hypermethylated in RAG2-deficient pre-B cells but hypomethylated in RAG1-deficient pre-B cells, indicating that wild-type (WT) RAG2 involves the Igκ locus demethylation in a RAG1-independent manner prior to rearrangement. We generated a series of RAG2 mutants between residue 350 and 383. We showed that these mutants mediated the Igκ rearrangement but failed to regulate the Igκ gene demethylation. We further analyzed that these mutants could increase RAG recombinase activity in vivo. We conclude that residues 350-383 region are responsible for endogenous Igκ locus demethylation at pre-B cells. We propose that WT RAG2 has an intrinsic function to regulate the Igκ locus demethylation.

Epigenetic modifications of the VH region after DJH recombination in Pro-B cells.

The variable region of murine immunoglobulin heavy chain (Igh) is assembled by sequential DH -JH and VH -DJH recombination. The accessibility of the Igh locus determines the order of rearrangement. Because of the large number of VH genes and the lack of a suitable model, the epigenetic modifications of VH genes after DJH recombination have not previously been characterized. Here, we employed two v-Abl pro-B cell lines, in which the Igh locus is in germline and DJH -recombined configurations, respectively. The DJH junction displays the characteristics of a recombination centre, such as high levels of activation-associated histone modifications and recombination-activating gene protein (RAG) binding in DJH -rearranged pro-B cells, which extend the recombination centre model proposed for the germline Igh locus. The different domains of the VH region have distinct epigenetic characteristics after DJH recombination. Distal VH genes have higher levels of active histone modifications, germline transcription and Pax5 binding, and good quality recombination signal sequences. Proximal VH genes are relatively close to the DJH recombination centre, which partially compensates for the low levels of the above active epigenetic modifications. DJH recombination centre might serve as a cis-acting element to regulate the accessibility of the VH region. Furthermore, we demonstrate that RAG weakly binds to functional VH genes, which is the first detailed assessment of RAG dynamic binding to VH genes. We provide a way for VH -DJH recombination in which the VH gene is brought into close proximity with the DJH recombination centre for RAG binding by a Pax5-dependent chromosomal compaction event, and held in this position for subsequent cleavage and VH -DJH joining.

TIGIT overexpression diminishes the function of CD4 T cells and ameliorates the severity of rheumatoid arthritis in mouse models.

Rheumatoid arthritis (RA) is an immune-mediated disease with a pathogenesis that involves CD4 T cell activation. Multiple immune regulatory molecules expressed on CD4(+) T cells were involved in RA pathogenesis. In this study, we investigated the role of T cell immunoglobulin and ITIM (immunoreceptor tyrosine-based inhibition motif) domain (TIGIT) in RA. The frequency of TIGIT-positive CD4(+) T cells in the synovial fluid (SF) of active RA patients was lower than that of inactive RA patients. And a negative correlation between RA disease activity and TIGIT expression was found. In CD4(+) T cells isolated from SF of active RA patients, TIGIT upregulation significantly decreased cell proliferation, as shown by MTT assay. TIGIT overexpression also significantly decreased the production of IFN-γ and IL-17, and increased that of IL-10, as determined by ELISA and qRT-PCR. In CD4(+) T cells isolated from SF of inactive RA patients, TIGIT was silenced by siRNA transfection. As expected, TIGIT knockdown resulted in an opposite effect on cell proliferation and the production of cytokines, including IFN-γ, IL-17 and IL-10. A RA mouse model was established using type II collagen induction. TIGIT was upregulated in RA mouse by lentivector infection. As expected, TIGIT overexpression in vivo significantly alleviated the disease severity and deceased the levels of anti-collagen II antibodies. TIGIT upregulation in the early stage was more effective to alleviate disease severity. Our data suggested the potential therapeutic role of TIGIT in RA patients.

MicroRNA-24 Regulates Osteogenic Differentiation via Targeting T-Cell Factor-1.

MicroRNAs (miRNAs) have been reported to have diverse biological roles in regulating many biological processes, including osteogenic differentiation. In the present study, we identified that miR-24 was a critical regulator during osteogenic differentiation. We found that overexpression of miR-24 significantly inhibited osteogenic differentiation, which decreased alkaline phosphatase activity, matrix mineralization and the expression of osteogenic differentiation markers. In contrast, inhibition of miR-24 exhibited an opposite effect. Furthermore, we delineated that miR-24 regulates post-transcriptionals of T-cell factor-1 (Tcf-1) via targeting the 3'-untranslated region (UTR) of Tcf-1 mRNA. MiR-24 was further found to regulate the protein expression of Tcf-1 in the murine osteoprogenitors cells and bone mesenchymal stem cells. Additionally, the positive effect of miR-24 suppression on osteoblast differentiation was apparently abrogated by Tcf-1 silencing. Taken together, our data suggest that miR-24 participates in osteogenic differentiation by targeting and regulating Tcf-1 expression in osteoblastic cells.

MiR-21 overexpression improves osteoporosis by targeting RECK.

Osteoporosis is a kind of metabolic bone disorder. MicroRNA-21 (miR-21) has been proven to play an important role in bone formation, whereas its role in osteoporosis is unclear. In the present study, miR-21 expression was inhibited by TNF-α in mesenchymal stem cells (MSCs). TNF-α induced cell apoptosis, and inhibited cell proliferation and differentiation of MSCs. Whereas the effect was reversed by miR-21 mimics. Expression of reversion-inducing cysteine-rich protein with Kazal motifs (RECK) which is a predicted target of miR-21 was inhibited by miR-21 mimics. A luciferase reporter gene assay showed that miR-21 directly bound to RECK 3'-UTR. The effect of TNF-α on MSCs was reversed by RECK siRNA which was consistent with miR-21 mimics. The expression of MT1-MMP was inhibited by TNF-α and enhanced by RECK siRNA and miR-21 mimics. For the in vivo study, an osteoporosis model (OVX) was established by bilateral oophorectomy in mice. The expression of miR-21 decreased and RECK increased in the OVX mice. When treated with lentiviral RECK shRNA, the osteocalcin concentration and alkaline phosphate activity of the OVX mice decreased. The bone mineral density of the right femur mid-diaphysis was improved by RECK shRNA. Collectively, miR-21 modulated the osteoporosis by targeting RECK. These results emphasize the role of miR-21 during osteoporosis and suggest RECK might be a new medical target for osteoporosis.

Elevated serum levels of decoy receptor 3 are associated with disease severity in patients with hemorrhagic fever with renal syndrome.

Hemorrhagic fever with renal syndrome (HFRS) is an acute viral infectious disease characterized by fever, hemorrhage and renal failure. HFRS has become a serious public health problem in China. Unfortunately, the pathogenesis of HFRS has not been completely clarified. The aim of this study is to investigate the changes of decoy receptor 3 (DcR3) and to further explore its potential roles in HFRS. The levels of serum DcR3 were measured by sandwich ELISA. We found serum DcR3 levels increased significantly, which reached peak value during the oliguric phase and in the critical group. Moreover, serum DcR3 levels were closely related to the levels of pro-inflammatory cytokines tumor necrosis factor-α (TNF-α) and parameters reflecting kidney injury including BUN, creatinine (Cr) and proteinuria. This study indicates that high levels of serum DcR3 have associations with the disease stages, severity and degree of kidney damage. Meanwhile, our results suggest that DcR3 may play a dual role in HFRS pathogenesis. First, DcR3 is involved in the inflammatory cascade response resulting in capillary permeability and kidney injury in the early stage. Secondly, HTNV infection induced DcR3 expression at the convalescent phase may act as a feed-back mechanism in anti-inflammatory response. Thus, a study of DcR3 is essential for a better understanding of HFRS pathogenesis.

The detection of vascular endothelial growth factor in serum of patients with hemorrhagic fever with renal syndrome.

Hemorrhagic fever with renal syndrome (HFRS) is an acute infectious disease characterized by endothelial cell dysfunction, which results in plasma exosmosis, hyperpermeability, and sometimes hemorrhages. As one of the vascular permeability cytokines, vascular endothelial growth factor (VEGF) might mediate the hyperpermeability caused by HFRS. In the present study, the levels of serum VEGF were measured by competitive inhibition ELISA. We found variable but persistently elevated levels of VEGF throughout the various stages and types of HFRS disease, which suggested that the levels of VEGF were closely correlated to the progression of HFRS. Moreover, elevated levels of VEGF have correlation with the severity and degree of kidney damage. Therefore, to study the relationship between levels of VEGF and disease severity of patients with HFRS is helpful to clarify the pathogenesis of HFRS.

Levels of HTNV-specific CD8+ T lymphocytes in PBMC from the patients with hemorrhagic fever with renal syndrome.

The pathogenesis of hemorrhagic fever with renal syndrome (HFRS) has not been fully clarified. Cell-mediated immunity appears to play a crucial role in the immune pathogenesis of HFRS. To explore the relationship between Hantaan (HTNV)-specific CD8(+) T lymphocytes and the immune pathogenesis of HFRS, the levels of interferon γ (IFN-γ) secreted by HTNV-specific CD8(+) T lymphocytes were detected by flow cytometry in peripheral blood mononuclear cells. Levels of HTNV-specific CD8(+) T lymphocytes in patients with HFRS were associated with different phases of HFRS. In fever phase, it was significantly elevated. The levels of HTNV-specific CD8(+) T lymphocytes in PBMC of patients with HFRS were negatively correlated with the levels of blood urea nitrogen and creatinine in plasma. The results show that the HTNV-specific CD8(+) T lymphocyte levels correlate with disease phases. Therefore, dynamic observation of these levels in patients with HFRS can help to judge the status of HFRS disease and to clarify the immune pathogenesis of HFRS.

Expression of VLA-4 molecule in PBMC from patients with hemorrhagic fever with renal syndrome.

OBJECTIVE AND DESIGN: The very late antigen-4 (VLA-4) bound to vascular cell adhesion molecule-1 could provide co-stimulatory signals for the activation of T lymphocytes, and these adhesion molecules play key roles in leukocyte adherence and propagation of inflammatory responses. We examined the levels of VLA-4 in the peripheral blood mononuclear cells (PBMC) of patients with hemorrhagic fever with renal syndrome (HFRS). MATERIALS AND METHODS: The levels of VLA-4 in PBMC samples collected from 53 patients by immunohistochemical staining were detected. RESULTS: The expression of VLA-4 in PBMC of HFRS patients at different stages were significantly higher than those in normal controls (P < 0.05), except recovery stage (P > 0.05). The expression of VLA-4 in PBMC of HFRS patients at different types were significantly higher than those in healthy controls (P < 0.05). The levels of VLA-4 in patients with HFRS were positively correlated with serum levels of blood urea nitrogen (BUN) and creatinine (Cr). CONCLUSIONS: VLA-4 might play an important role in the immunopathological lesions of HFRS. We found that VLA-4 levels were closely correlated to the severity of the HFRS and the degree of kidney damage.