ACE/ACE2 Ratio and MMP-9 Activity as Potential Biomarkers in Tuberculous Pleural Effusions

Objective: Pleural effusion is common problem, but the rapid and reliable diagnosis for specific pathogenic effusions are lacking. This study aimed to identify the diagnosis based on clinical variables to differentiate pleural tuberculous exudates from other pleural effusions. We also investigated the role of renin-angiotensin system (RAS) and matrix metalloproteinase (MMPs) in the pathogenesis of pleural exudates.Experimental design: The major components in RAS and extracellular matrix metabolism, including angiotensin converting enzyme (ACE), ACE2, MMP-2 and MMP-9 activities, were measured and compared in the patients with transudative (n = 45) and exudative (n = 80) effusions. The exudative effusions were come from the patients with tuberculosis (n = 20), pneumonia (n = 32), and adenocarcinoma (n = 28).Results: Increased ACE and equivalent ACE2 activities, resulting in a significantly increased ACE/ACE2 ratio in exudates, were detected compared to these values in transudates. MMP-9 activity in exudates was significantly higher than that in transudates. The significant correlation between ACE and ACE2 activity that was found in transudates was not found in exudates. Advanced analyses showed significantly increased ACE and MMP-9 activities, and decreased ACE2 activity in tuberculous pleural effusions compared with those in pneumonia and adenocarcinoma effusions. The results indicate that increased ACE and MMP-9 activities found in the exudates were mainly contributed from a higher level of both enzyme activities in the tuberculous pleural effusions.Conclusion: Interplay between ACE and ACE2, essential functions in the RAS, and abnormal regulation of MMP-9 probably play a pivotal role in the development of exudative effusions. Moreover, the ACE/ACE2 ratio combined with MMP-9 activity in pleural fluid may be potential biomarkers for diagnosing tuberculous pleurisy.     

Vitamin D Receptor Controls Cell Stemness in Acute Myeloid Leukemia and in Normal Bone Marrow

1. Download : Download high-res image (99KB)
2. Download : Download full-size image

     

Bone marrow-derived mesenchymal stem cells repair severe acute pancreatitis by secreting miR-181a-5p to target PTEN/Akt/TGF-β1 signaling

BackgroundSevere acute pancreatitis (SAP) is associated with high morbidity and mortality. Bone marrow mesenchymal stem cells (BMSCs) have shown obvious protective effect on SAP. However, little is known about the underlying mechanism. The objective of this study is to unravel the role and regulatory mechanism of miR-181a-5p in BMSCs-mediated pancreatic repair.MethodsBMSCs were isolated from Sprague-Dawley rats and characterized by flow cytometry and Oil Red O staining. Sodium taurocholate- and caerulein-induced models were used as SAP models in vivo and in vitro, respectively. Pancreatic injury were evaluated by H&E and histopathological analysis, as well as by measuring levels of amylase, lipase and cytokines. qRT-PCR and western blotting were performed to detect the level of miR-181a-5p and the protein levels of PTEN/Akt, respectively. ELISA was conducted to detect the levels of TNF-α, IL-1β, IL-6, angiopoietin, IL-4, IL-10 and TGF-β1. The apoptotic rate of AR42 J cells was quantitated by concurrent staining with Annexin-V-FITC and PI.ResultsBMSCs significantly attenuated pancreatic injury in SAP rats by reducing inflammatory infiltration and necrosis, and this effect was abolished by CXCR4 agonist AMD3100. ADM3100 exhibited more severe pancreatic injury and decreased miR-181a-5p levels in the pancreas and serum compared to SAP group. Overexpression of miR-181a-5p in BMSCs (BMSCs-miR-181a-5p) markedly potentiated the protective effect of BMSCs by reducing histological damage and levels of amylase and lipase. Moreover, BMSCs-miR-181a-5p dramatically reduced levels of angiopoietin, TNF-α, IL-1β and IL-6, but induced the levels of IL-4 and IL-10. In caerulein-treated AR42 J cells, co-culturing of BMSCs-miR-181a-5p alleviated caerulein-induced increase of amylase and lipase, and apoptosis via PTEN/Akt/TGF-β1 signaling.ConclusionBMSCs alleviate SAP and reduce inflammatory responses and apoptosis by secreting miR-181a-5p to target PTEN/Akt/TGF-β1 signaling. Hence, BMSCs-miR-181a-5p could serve as potential therapeutic target for SAP.     

Quantifying Single Microvessel Permeability in Isolated Blood-perfused Rat Lung Preparation

The isolated blood-perfused lung preparation is widely used to visualize and define signaling in single microvessels. By coupling this preparation with real time imaging, it becomes feasible to determine permeability changes in individual pulmonary microvessels. Herein we describe steps to isolate rat lungs and perfuse them with autologous blood. Then, we outline steps to infuse fluorophores or agents via a microcatheter into a small lung region. Using these procedures described, we determined permeability increases in rat lung microvessels in response to infusions of bacterial lipopolysaccharide. The data revealed that lipopolysaccharide increased fluid leak across both venular and capillary microvessel segments. Thus, this method makes it possible to compare permeability responses among vascular segments and thus, define any heterogeneity in the response. While commonly used methods to define lung permeability require postprocessing of lung tissue samples, the use of real time imaging obviates this requirement as evident from the present method. Thus, the isolated lung preparation combined with real time imaging offers several advantages over traditional methods to determine lung microvascular permeability, yet is a straightforward method to develop and implement.     

ZMYND8-regulated IRF8 transcription axis is an acute myeloid leukemia dependency

1. Download : Download high-res image (151KB)
2. Download : Download full-size image

     

6-Shogaol induces apoptosis in acute lymphoblastic leukaemia cells by targeting p53 signalling pathway and generation of reactive oxygen species

Combination therapies, using medicinal herbs, are broadly recommended to attenuate the chemotherapy adverse effects. Based on our previous findings considering the anti-leukaemic effects of ginger extract on acute lymphoblastic leukaemia (ALL) cells, the present study was aimed to investigate the anti-cancer role of this pharmaceutical plant on ALL mice models. Moreover, we worked towards identifying the most anti-leukaemic derivative of ginger and the mechanism through which it may exert its cytotoxic impact. In vivo experiments were performed using five groups of six C57BL/6 nude mice, and the anti-leukaemic activity of ginger extract alone or in combination with methotrexate (MTX) was examined. Results showed increased survival rate and reduced damages in mice brain and liver tissues. Subsequently, MTT assay demonstrated synergistic growth inhibitory effect of 6-shogaol (6Sh) and MTX on ALL cell lines and patients primary cells. Eventually, the molecular anti-neoplastic mechanism of 6Sh was evaluated using Bioinformatics. Flow cytometry illustrated 6Sh-mediated apoptosis in Nalm-6 cells confirmed by Western blotting and RT-PCR assays. Further analyses exhibited the generation of reactive oxygen species (ROS) through 6Sh. The current study revealed the in vivo novel anti-leukaemic role of ginger extract, promoted by MTX. Moreover, 6-shogaol was introduced as the major player of ginger cytotoxicity through inducing p53 activity and ROS generation.     

p21WAF1 modulates drug-induced apoptosis and cell cycle arrest in B-cell precursor acute lymphoblastic leukemia

p21^WAF1 is a well-characterized mediator of cell cycle arrest and may also modulate chemotherapy-induced cell death. The role of p21^WAF1 in drug-induced cell cycle arrest and apoptosis of acute lymphoblastic leukemia (ALL) cells was investigated using p53-functional patient-derived xenografts (PDXs), in which p21^WAF1 was epigenetically silenced in T-cell ALL (T-ALL), but not in B-cell precursor (BCP)-ALL PDXs. Upon exposure to diverse cytotoxic drugs, T-ALL PDX cells exhibited markedly increased caspase-3/7 activity and phosphatidylserine (PS) externalization on the plasma membrane compared with BCP-ALL cells. Despite dramatic differences in apoptotic characteristics between T-ALL and BCP-ALL PDXs, both ALL subtypes exhibited similar cell death kinetics and were equally sensitive to p53-inducing drugs in vitro, although T-ALL PDXs were significantly more sensitive to the histone deacetylase inhibitor vorinostat. Transient siRNA suppression of p21^WAF1 in the BCP-ALL 697 cell line resulted in a moderate depletion of the cell fraction in G1 phase and marked increase in PS externalization following exposure to etoposide. Furthermore, stable lentiviral p21^WAF1 silencing in the BCP-ALL Nalm-6 cell line accelerated PS externalization and cell death following exposure to etoposide and vorinostat, supporting previous findings. Finally, the Sp1 inhibitor, terameprocol, inhibited p21^WAF1 expression in Nalm-6 cells exposed to vorinostat and also partially augmented vorinostat-induced cell death. Taken together, these findings demonstrate that p21^WAF1 regulates the early stages of drug-induced apoptosis in ALL cells and significantly modulates their sensitivity to vorinostat.     

Molecular biology of Philadelphia chromosome in chronic granulocytic leukaemia and acute lymphoblastic leukaemia

In classical t(9;22) translocation, as observed in chronic granulocytic leukemia (CGL), a hybrid DNA unit is produced, including a rearranged PHL gene, previously known as bcr (breakpoint cluster region) plus the translocated c-abl gene from chromosome 9: a hybrid bcr-abl protein, p210 is formed, with increased tyrosine kinase activity. Such DNA rearrangement, with a p210 protein synthesis, is also found in cases of Philadelphia-positive acute lymphoblastic leukemia (ALL), but in apparently similar cases the bcr gene is not rearranged, and a novel p190 abl-related protein can be found; c-abl rearrangement has also been observed.It is thus established that correlations between cytogenetic and molecular events can be found in CGL and ALL, as in other haemopoietic malignancies: translocation and possible rearrangement of the c-abl oncogene seem of particular importance in this case.