AbstractMethyltransferase-like 3 (METTL3) was originally known to be responsible for N6-methyladenosine (m6A) modification of mRNA. Recent studies have found that METTL3 plays important roles in a variety of tumors by regulating the translation of oncogenes. However, the functional and regulating mechanisms of METTL3 in human gastric cancer have not yet been understood. Here we knocked down METTL3 in human gastric cancer cell lines, AGS and MKN45, by using shRNA transfection. RT-qPCR assay and western blotting verified the effectiveness of RNA interference on mRNA and protein levels, respectively. Then we found that METTL3 knockdown inhibited cell proliferation, migration and invasion in AGS and MKN45 cells. Moreover, METTL3 knockdown decreased Bcl2 and increased Bax and active Caspase-3 in gastric cancer cells, which suggested the apoptotic pathway was activated. Mechanistic investigation suggested that METTL3 led to inactivation of the AKT signaling pathway in human gastric cancer cells, including decreased phosphorylation levels of AKT and expression of down-stream effectors p70S6K and Cyclin D1. In conclusion, our study reveals that down-regulation of METTL3 inhibits the proliferation and mobility of human gastric cancer cells and leads to inactivation of the AKT signaling pathway, suggesting that METTL3 may be a potential target for the treatment of human gastric cancer.

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Angiogenesis (the growth of new blood vessels) is essential in most of the body’s physiological processes, such as in the normal functioning of the endometrium during and after the menstrual cycle. Vascular endothelial growth factor (VEGF) and matrix metalloproteinase (MMP) are the mostly expressed angiogenic factors, especially, during the process of endometrial degeneration and remodeling. In carcinogenesis, tumor hypoxia-induced factors, through the process of “angiogenic switch”, stimulate the production of angiogenic factors, particularly VEGF and MMP. Subsequently, these angiogenic factors are associated with degradation, differentiation, proliferation, and migration of vascular endothelial cells, enhancing the formation of new blood vessels to supply the tumor with oxygen and nutrients. This process is equally significant for tumor development and metastasis. Hence, like in other cancers, the overexpression of MMP and VEGF in endometrial cancer (EC) seems to play a significant role in its tumorigenesis and metastasis. This research will discuss the influence of MMP and VEGF on angiogenesis, metastasis, and the prognosis of EC as well as the clinical importance of the factors in the diagnosis of EC.

We aimed at the effects of long non-coding RNA (lncRNA) SNHG5 on proliferation, metastasis and migration of colorectal cancer (CRC) cells. We also investigated regulatory relationships among miR-132-3p, SNHG5 and CREB5 and their roles in CRC. 25 pairs of samples containing CRC tissues and matched para-tumor tissues were obtained to examine SNHG5, miR-132-3p and CREB5 expression by qRT-PCR or Western blot. The targeted relationship between miR-132-3p and SNHG5 or CREB5 was confirmed by dual luciferase report assay as well as RNA pull down assay. The expression of SNHG5, miR-132-3p and CREB5 in CRC cells were regulated by cell transfection. CRC cellular proliferation was assayed by CCK-8 and meanwhile flow cytometry was adopted to observe apoptosis. Metastasis and migration of CRC cells were determined respectively by means of Transwell assay and scratch test. The effects of SNHG5 on CRC were researched in vivo, too. SNHG5 or CREB5 was up-regulated in CRC tissues and cells, whereas miR-132-3p was down-regulated. Overexpression of SNHG5 and CREB5 resulted in the enhancement of proliferation, metastasis, migration and the inhibition of apoptosis in CRC cells, while miR-132-3p led to the opposite result. LncRNA SNHG5 promoted proliferation, migration and metastasis of CRC cells but inhibited apoptosis by modulating miR-132-3p/CERB5.

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Although a minor population of paroxysmal nocturnal hemoglobinuria (PNH)-type blood cells is often detected in patients with aplastic anemia (AA) and refractory anemia (RA), the significance of such cells in the pathophysiology of bone marrow (BM) failure remains obscure. We therefore examined clonality in peripheral blood granulocytes from 118 female patients with AA or myelodysplastic syndrome using the X chromosome inactivation pattern. Clonality, defined as a clonal population accounting for 35% or more of total granulocytes, was confirmed in 22 of 68 (32.4%) AA patients, in 13 of 44 (29.5%) RA patients, in all 4 RA with excess blasts (RAEB) patients, and in 4 patients with PNH. When the frequency of patients with granulocyte clonality was compared with respect to the presence of increased PNH-type cells, the frequency was significantly lower in AA patients with (PNH ؉ ; 21.2%) than without (PNH ؊ ; 42.9%) increased numbers of PNH-type cells (P ‫؍‬ .049). Clonality was absent in granulocytes from the 15 PNH ؉ RA patients but present in 13 of 29 (44.8%) PNH ؊ RA patients (P ‫؍‬ .0013). The absence of clonality in AA and RA patients before treatment was strongly associated with positive response to immunosuppressive therapy (without clonality, 74.4%; with clonality, 33.3%; P ‫؍‬ .0031) in all patients as well as in PNH ؉ patients (without clonality, 96.2%; with clonality, 66.6%, P ‫؍‬ .026). These results suggest that AA and RA with a minor population of PNHtype cells are benign types of BM failure with immune pathophysiology that have little relationship to clonal disorders such as RAEB or acute myeloid leukemia. IntroductionAplastic anemia (AA) and myelodysplastic syndrome (MDS) are hematopoietic dyscrasias characterized by pancytopenia and inappropriate production of mature blood cells from the bone marrow. They differ in terms of disease definition: AA is basically benign bone marrow (BM) failure due to extrinsic insult to hematopoietic stem cells, while MDS is a clonal disorder derived from a defective stem cell. 1-3 However, to differentiate AA from refractory anemia (RA) of MDS in clinical practice can be difficult, as a diagnosis of RA depends largely on a subjective judgment of morphologic abnormalities in mature blood cells, and a laboratory marker that can discriminate between them remains unknown. 4,5 We recently demonstrated that a minor (Ͻ 1%) population of CD55 Ϫ CD59 Ϫ granulocytes or red blood cells (RBCs) can be detected in numerous AA patients 6 and in about 20% of RA patients. 7 RA patients with a subtle increase in such paroxysmal nocturnal hemoglobinuria (PNH)-type cells (PNH ϩ patients) had distinct clinical features compared with RA patients without increased PNH-type cells (PNH Ϫ patients), such as lower rates of karyotypic abnormality and higher probability of response to cyclosporine (CyA) therapy. The presence of PNH-type cells therefore appeared to represent a marker for benign types of BM failure. However, several studies contradict this hypothesis. Some reports described AA pati...

Over 1,000 surveys were distributed to randomly selected paraprofessionals, teachers, related service personnel, and administrators throughout three of the most sparsely populated states (North Dakota, South Dakota, and Wyoming) to study the training and support needs of paraprofessionals in rural settings. The need for additional training is apparent since the paraprofessionals surveyed requested assistance in understanding students with disabilities, their roles as paraprofessionals, task analysis procedures, and the Individual Educational Program (IEP) process. Other findings address the quality of supervision, previous training, and other factors which influence paraprofessionals career decisions. Recommendations center on strategies for providing high-quality, cost effective training that also permits paraprofessionals the opportunity to receive certification as a special education teacher.

BackgroundGamma (γ)-Aminobutyric acid (GABA) as a bioactive compound is used extensively in functional foods, pharmaceuticals and agro-industry. It can be biosynthesized via decarboxylation of monosodium glutamate (MSG) or L-glutamic acid (L-Glu) by glutamate decarboxylase (GAD; EC4.1.1.15). GADs have been identified from a variety of microbial sources, such as Escherichia coli and lactic acid bacteria. However, no GADs from Streptomyces have been characterized. The present study is aimed to identify new GADs from Streptomyces strains and establish an efficient bioproduction platform for GABA in E. coli using these enzymes.ResultsBy sequencing and analyzing the genomes of three Streptomyces strains, three putative GADs were discovered, including StGAD from Streptomyces toxytricini NRRL 15443, SsGAD from Streptomyces sp. MJ654-NF4 and ScGAD from Streptomyces chromofuscus ATCC 49982. The corresponding genes were cloned from these strains and heterologously expressed in E. coli BL21(DE3). The purified GAD proteins showed a similar molecular mass to GadB from E. coli BL21(DE3). The optimal reaction temperature is 37 °C for all three enzymes, while the optimum pH values for StGAD, SsGAD and ScGAD are 5.2, 3.8 and 4.2, respectively. The kinetic parameters including Vmax, Km, kcat and kcat/Km values were investigated and calculated through in vitro reactions. SsGAD and ScGAD showed high biocatalytic efficiency with kcat/Km values of 0.62 and 1.21 mM− 1·s− 1, respectively. In addition, engineered E. coli strains harboring StGAD, SsGAD and ScGAD were used as whole-cell biocatalysts for production of GABA from L-Glu. E. coli/SsGAD showed the highest capability of GABA production. The cells were repeatedly used for 10 times, with an accumulated yield of 2.771 kg/L and an average molar conversion rate of 67% within 20 h.ConclusionsThree new GADs have been functionally characterized from Streptomyces, among which two showed higher catalytic efficiency than previously reported GADs. Engineered E. coli harboring SsGAD provides a promising cost-effective bioconversion system for industrial production of GABA.Electronic supplementary materialThe online version of this article (10.1186/s13036-019-0154-7) contains supplementary material, which is available to authorized users.