Expression and Fuactional Role of HERG1, K^+ Channels in Leukemic Cells and Leukemic Stem Cells

In order to investigate the expression and functional role of HERG1 K+ channels in leukemic cells and leukemic stem cells (LSCs), RT-PCR was used to detect the HERG1 K+ channels expression in leukemic cells and LSCs. The functional role of HERG1 K+ channels in leukemic cell proliferation was measured by MTT assay, and cell cycle and apoptosis were analyzed by flow cy- tometry. The results showed that herg mRNA was expressed in CD34+/CD38-, CD123+ LSCs but not in circulating CD34+ cells. Herg mRNA was also up-regulated in leukemia cell lines K562 and HL60 as well as almost all the primary leukemic cells while not in normal peripheral blood mononuclear cells (PBMNCs) and the expression of herg mRNA was not associated with the clinical and cytoge- netic features of leukemia. In addition, leukemic cell proliferation was dramatically inhibited by HERG K+ channel special inhibitor E-4031. Moreover, E-4031 suppressed the cell growth by induc- ing a specific block at the G1/S transition phase of the cell cycle but had no effect on apoptosis in leukemic cells. The results suggested that HERG1 K+ channels could regulate leukemic cells prolif- eration and were necessary for leukemic cells to proceed with the cell cycle. HERG1 K+ channels may also have oncogenic potential and may be a biomarker for diagnosis of leukemia and a novel potential pharmacological target for leukemia therapy.

Comparison of microRNA Expression Profiles in HCC-derived Microvesicles and the Parental Cells and Evaluation of Their Roles in HCC

Summary： To determine whether the microRNAs （miRNAs） contained in cancer-derived microvesi- cles （MVs） mirror those of the parental tumor cells, we compared the miRNA expression profiles of MVs derived from their parental hepatocellular carcinoma ,（HCC） cells. The presence and levels of 888 miRNAs from SMMC-7721 cells and MVs were detected by Agilent miRNA microarray analy- sis. Four selected miRNAs were verified by real time qRT-PCR. Furthermore, the genes of the miRNAs were bioinformatically identified to explore potential roles of the miRNAs in HCC micro- environment. Our results showed that miRNAs expression profiles of MVs derived from HCC were significantly changed. Of all the miRNAs tested, 148 miRNAs were co-expressed in MVs and SMMC-7721 cells, only 121 and 15 miRNAs were detected in MVs and SMMC-7721 cells, respec- tively. Among the 148 co-expressing miRNAs, 48 miRNAs had the similar expression level and 6 of them were supposed to be oncogenic or suppressive miRNAs. According to the target prediction by Quantile Algorithm method, these miRNAs may regulate 3831 genes which were closely related to cell cycle, apoptosis and oncogenesis, and 78 were known tumor suppressors or oncogenes. Gene ontology （GO） analysis indicated that 3831 genes were mainly associated with nucleic acid binding, cell death, cell adhesion. MVs containing miRNAs, released into the HCC microenvironment, bear the characteristic miRNAs of the original cells and might participate in cancer progression.

HERG K+ channels expression in gastric cancers and analysis of its regulation in tumor cell proliferation and apoptosis

Objective： To investigate the expression of hergl gene in tumor tissues from gastric carcinomas and gastric carcinoma cell lines, and study the relationship between HERG K＋ channel expressions and tumor cell proliferation and apoptosis. Methods： RT-PCR and PCR assays were used to detect the expression of hergl gene in 64 gastric carcinomas and the gastric cancer cell line SGC-7901. Blocking the HERG K＋ channels was used to evaluate their effects on tumor cell proliferation and apoptosis. Results：The statistically significant expression of hergl gene was detected in all the gastric cancers and SGC-7901 cells, but not in normal tissues. The HERG K＋ channel blocker, E-4031, increased the cell population in G0/G1（P 〈 0.05） and the number of apoptotic tumor cells（P 〈 0.05）. Conclusion： HERG K＋ channels were expressed in all gastric carcinomas tested and these channels appear to modulate tumor cell proliferation and apoptosis.

Close Correlation between Development of MODS during the Initial 72h of Hospitalization and Hospital Mortality in Severe Fever with Thrombocytopenia Syndrome

An emerging infectious disease was identified as severe fever with thrombocytopenia syn- drome （SFTS） in central China since late March 2009. We found the patients with SFTS had severe clinical symptoms, and progressed rapidly to multiple organ dysfunction syndrome （MODS） with high fatality rate of 25%-30%. The aim of this study was to assess the significance of risk factors predicting the development of MODS and death in SFTS patients. Consecutive SFTS admissions between May 2009 and September 2011 were analyzed for parameters of organ function during hospitalization using Marshall scoring system for MODS, and platelet counts were recorded on admission and at 24, 48, 72 h and one week after admission. We investigated the kinetics of organ failures and analyzed the associa- tion between age, platelet count and development of MODS or death. A total of 92 SFTS patients were enrolled in this study. Among them, 32 patients with dysfunction of over 4 organs were identified, 45% of them died within 72 h, 72% died within 5 days, and 76% died within 7 days after admission. We also found cumulative Marshall score was significantly higher in death patients （11.76＋2.05） than in survival patients （4.22~1.98） （P〈0.001）. In addition, SFTS patients had older age and lower platelet counts in MODS and death groups. Furthermore, we also observed that there was a close correlation between platelet count on admission and Marshall score （P〈0.001）. High Marshall score, advanced age and lower platelet counts were the main risk factors for the development of MODS, and those factors could predict mortality in SFTS patients, suggesting prompt treatment and close monitoring of severe complications, especially MODS, are of great importance in saving patients＇ lives.

Detection of Microvesicle miRNA Expression in ALL Subtypes and Analysis of Their Functional Roles

Microvesicles （MVs） are the heterogeneous mixtures of vesicles. MVs released by leukemia cells constitute an important part of the leukemia microenvironment. MVs might act as important reser- voirs of microRNAs （miRNAs）. It is worth evaluating whether MVs possess some unique miRNA con- tents that are valuable in understanding the pathogenesis. In this study, we investigated the miRNA ex- pression patterns of Nalm-6-derived MVs, Jurkat-derived MVs and normal cell-derived MVs using miRNA microarrays. The potential target genes regulated by differentially expressed miRNAs were also predicted and analyzed. Results demonstrated that 182 miRNAs and 166 miRNAs were differentially expressed in Nalm-6-MVs and Jurkat-MVs, respectively. Many oncogenes, tumor suppressors and sig- nal pathway genes were targeted by these aberrantly expressed miRNAs, which might contribute to the development of B-ALL or T-ALL. Our findings expanded the potential diagnostic markers of ALL and provided useful information for ALL pathogenesis.

Detection and Clinical Signiifcance of Circulating Microvesicles in Severe Fever with Thrombocytopenia Syndrome

Objective Microvesicles (MV) released from blood cells play an important role in the progress of diseases. The purpose of this study is to detect circulating MV level of patients suffering from severe fever with thrombocytopenia syndrome (SFTS) . Methods The plasma samples of SFTS patients and healthy controls were collected. And MV from samples were isolated and MV levels were detected quantitatively. Results The results showed that the level of the circulating MV in SFTS patients was obviously higher than that of the healthy control, with statistically signiifcant difference. Further analysis revealed that MV level was relevant to the severity of SFTS patients, namely the higher the concentration of MV, the more severity of the disease. Linear correlation analysis showed that the circulating MV level in SFTS was correlated positively with leukocyte count (r=0.243, P<0.05), but no correlation with the platelet count (r=0.193, P>0.05). Conclusions We demonstrated that there existed high level of circulating MV in SFTS patients, and the level of circulating MV had a close relationship with the severity of the disease and leukocyte count. Therefore, monitoring the level of circulating MV may provide a new insight for predicting the prognosis of SFTS patients.

Early development and functional properties of tryptase/chymase double-positive mast cells from human pluripotent stem cells

Mast cells (MCs) play a pivotal role in the hypersensitivity reaction by regulating the innate and adaptive immune responses. Humans have two types of MCs. The first type, termed MCTC, is found in the skin and other connective tissues and expresses both tryptase and chymase, while the second, termed MCT, which only expresses tryptase, is found primarily in the mucosa. MCs induced from human adult-type CD34+ cells are reported to be of the MCT type, but the development of MCs during embryonic/fetal stages is largely unknown. Using an efficient coculture system, we identified that a CD34+c-kit+ cell population, which appeared prior to the emergence of CD34+CD45+ hematopoietic stem and progenitor cells (HSPCs), stimulated robust production of pure Tryptase+Chymase+ MCs (MCTCs). Single-cell analysis revealed dual development directions of CD34+c-kit+ progenitors, with one lineage developing into erythro-myeloid progenitors (EMP) and the other lineage developing into HSPC. Interestingly, MCTCs derived from early CD34+c-kit+ cells exhibited strong histamine release and immune response functions. Particularly, robust release of IL-17 suggested that these early developing tissue-type MCTCs could play a central role in tumor immunity. These findings could help elucidate the mechanisms controlling early development of MCTCs and have significant therapeutic implications.

Infusions of recipient-derived cytokine-induced killer cells of donor origin eradicated residual disease in a relapsed leukemia patient after allo-hematopoietic stem cell transplantation

A female patient diagnosed with acute myelocytic leukemia M5a （AML-M5a） relapsed 986 days after her allogeneic peripheral blood stem cell transplantation （alIo-PBSCT） from an unrelated male donor with matched human leukocyte antigen （HLA）. Three re-induction chemotherapies were administered, and partial remission was achieved. The patient was given repetitive infusion of cytokine-induced killer （CIK） cells expanded from recipient peripheral mononuclear cells of full donor chimerism due to loss of contact of quondam donor for donor lymphocyte infusion （DLI） and rejection of second transplantation. The patient achieved complete cytogenetical remission. This strategy might overcome the obstacle of donor unavailability and present an appealing new therapeutic alternative to donor-recruited adoptive immunotherapy for relapsed disease at post-transplantation.

Dynamic regulation of GATA2 in fate determination in hematopoiesis: possible approach to hPSC-derived hematopoietic stem/ progenitor cells

GATA2,a principal member of the GATA family,plays important roles in the generation and maintenance of hematopoietic stem/progenitor cells.Among the three mRNA transcripts,the distal first exon of GATA2(IS exon)is specific for hematopoietic and neuronal cells.GATA2 mutants with abnormal expression are often present in acute myeloid leukemia-related familial diseases and myelodysplastic syndrome,indicating the crucial significance of GATA2 in the proper maintenance of blood system functions.This article offers an overview of the regulation dynamics and function of GATA2 in the generation,proliferation,and function of hematopoietic stem cells in both mouse and human models.We acknowledge the current progress in the cell fate determination mechanism by dynamic GATA2 expression.The gene modification approaches for inspecting the role of GATA2 in definitive hematopoiesis demonstrate the potential for acquiring hPSC-derived hematopoietic stem cells via manipulated GATA2 regulation.

HERG1 K^+ Channels on the Leukemic Cells Mediated Angiogenesis in vitro

Human ether-a-go-go-related gene （HERG1） K^＋ channels are overexpressed in leukemia, which contributes to neoangiogene- sis. The purpose of this study was to investigate the role of HERG1 K^＋ channels on leukemia angiogenesis. We cultured human umbili- cal vein endothelial cells （HUVECs） in conditioned media, which were derived from leukemic cells with or without E-4031, a HERG1 K^＋ channel special inhibitor. The HUVECs proliferation was mea- sured using CCK-8 assay and migration by a Trans-well. Endothelial tube formation was investigated using Matrigel. Vascular endothelial growth factor （VEGF） levels were tested by ELISA and VEGF mRNA expression using RT-PCR. Our results revealed that blocking HERG1 K^＋ channels could inhibit leukemia-induced HUVECs pro- liferation, migration, and tube formation in vitro. The results sug- gested that HERG1 K~ channels could increase leukemia angio- genesis. Furthermore, blockage of HERG1 K^＋ channels could also decrease leukemic cells secreting VEGF and expressing VEGF mRNA. HERG1 K^＋ channels have a promoting effect on leukemia angiogenesis, and the possible mechanism may be that HERG1 K^＋ channels enhance VEGF expression. Thus, HERG1 K4 channel is a potential target of antiangiogenesis in leukemia.

DPHL:A DIA Pan-human Protein Mass Spectrometry Library for Robust Biomarker Discovery

To address the increasing need for detecting and validating protein biomarkers in clinical specimens,mass spectrometry(MS)-based targeted proteomic techniques,including the selected reaction monitoring(SRM),parallel reaction monitoring(PRM),and massively parallel dataindependent acquisition(DIA),have been developed.For optimal performance,they require the fragment ion spectra of targeted peptides as prior knowledge.In this report,we describe a MS pipeline and spectral resource to support targeted proteomics studies for human tissue samples.To build the spectral resource,we integrated common open-source MS computational tools to assemble a freely accessible computational workflow based on Docker.We then applied the workflow to generate DPHL,a comprehensive DIA pan-human library,from 1096 data-dependent acquisition(DDA)MS raw files for 16 types of cancer samples.This extensive spectral resource was then applied to a proteomic study of 17 prostate cancer(PCa)patients.Thereafter,PRM validation was applied to a larger study of 57 PCa patients and the differential expression of three proteins in prostate tumor was validated.As a second application,the DPHL spectral resource was applied to a study consisting of plasma samples from 19 diffuse large B cell lymphoma(DLBCL)patients and 18 healthy control subjects.Differentially expressed proteins between DLBCL patients and healthy control subjects were detected by DIA-MS and confirmed by PRM.These data demonstrate that the DPHL supports DIA and PRM MS pipelines for robust protein biomarker discovery.DPHL is freely accessible at https://www.iprox.org/page/project.html?id=IPX0001400000.