Search results for: Human Neuroglobin(NGB)ELISA Kit
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Calcium chloride enhances the delivery of exosomes.Exosomes might have an unimproved potential to serve as effective delivery vehicles. However, when exosomes are developed for therapeutic applications, a method to enhance their delivery is important. This study aimed to evaluate wheather calcium chloride (CaCl2) or other chloride compounds could enhance exosome delivery to various cells without causing toxicity. Exosomes were purified from human serum by using the ExoQuick exosome precipitation kit. Isolated exosomes were mixed with CaCl2 at concentrations ranging from 100 μM to 1 mM, and then washed using Amicon filter for treating the cells. The delivery efficiency of exosomes and the viability of the cells [HEK 293 (human kidney cells) and H9C2 (rat cardiomyocytes)] were evaluated. Cellular uptake of exosomes was observed using a confocal microscope based on PKH26 labeling of exosomes. CaCl2 increased the delivery of exosomes in a dose- and treatment time-dependent manner. In HEK 293 cells, a CaCl2 concentration of 400 μM and exposure time of 12 h increased the delivery of exosomes by >20 times compared with controls. In H9C2 cells, a CaCl2 concentration of 400 μM and exposure time of >24 h increased the delivery of exosomes by >400 times compared with controls. The viability of both cell lines was maintained up to a CaCl2 concentration of 1 mM. However, cobalt chloride, cupric chloride, and magnesium chloride did not change the delivery of exosomes in both cell lines. These results suggest that the use of CaCl2 treatment might be a useful method for enhancing the delivery of exosomes.
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Fluorene-9-bisphenol inhibits epithelial-mesenchymal transition of human endometrial cancer Ishikawa cells by repressing TGF-β signaling pathway.Fluorene-9-bisphenol (BHPF), a new derivative of bisphenol A (BPA), has been introduced for treatment with estrogen-related tumors, such as endometrial cancer. This study investigated the potential mechanism underlying the action of BHPF against endometrial cancer in vitro. We used the cell counting kit-8 (CCK8) method on Ishikawa cells to screen sub-lethal doses of BHPF and established the optimal concentration at which BHPF influenced the proliferation of Ishikawa cells. Effect of BHPF on cell migration and invasion was investigated by cell scratch assay and transwell assay, respectively. Expression levels of epithelial-mesenchymal transition (EMT)-related proteins were detected by Western blot analysis. BHPF was found to inhibit the proliferation of Ishikawa cells, whose migration and invasion abilities were also reduced. Western blot indicated that BHPF can significantly inhibit the EMT process of Ishikawa cells by blocking transforming growth factor-β (TGF-β) signaling pathway. This is the first report of the effect of BHPF on the biological behavior of endometrial cancer cells and its inhibition of endometrial cancer progression by repressing both endometrial cell proliferation and epithelial-mesenchymal transition, hence suggesting it as a novel anti-cancer drug. Graphical abstract Schematic representation of the molecular basis underlying BHPF treatment. BHPF repressed the EMT process by regulating EMT-related genes, such as E-cadherin, N-cadherin, and vimentin as well as the TGF-β signaling pathway-related genes, including p-Smad2/3 and slug, in a BHPF-dependent manner.
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Neuroprotective Effects of Genistein in a SOD1-G93A Transgenic Mouse Model of Amyotrophic Lateral Sclerosis.Oxidant toxicity has been implicated in the pathogenesis of amyotrophic lateral sclerosis (ALS), an insidiously progressive neurodegenerative disorder involving upper and lower motor neurons. Here, we investigated the cellular and molecular mechanisms underlying the neuroprotective effects of an anti-oxidant genistein in SOD1-G93A transgenic mouse model of ALS. Rotarod test, hanging wire test and hindlimb clasping test were used to determined disease onset and assess motor performance. Immunostaining together with neuronal size measurement were used to count viable motor neurons. In addition, immunostaining procedure and ELISA kit were used to assess the inflammatory response in the spinal cord. Our results showed that Genistein administration suppressed the production of pro-inflammatory cytokines and alleviated gliosis in the spinal cord of SOD1-G93A mice. In addition, genistein administration induced autophagic processes and enhanced the viability of spinal motor neurons. As a result, genistein alleviated ALS-related symptoms and slightly prolonged the lifespan of SOD1-G93A mice. Taken together, our results indicate that genistein is neuroprotective in SOD1-G93A mice, suggesting genistein could be a promising treatment for human ALS. Graphical Abstract Genistein protects impariments in SOD1-G93A transgenic mouse model.
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miR-27a-3p Functions as a Tumor Suppressor and Regulates Non-Small Cell Lung Cancer Cell Proliferation via Targeting HOXB8.MicroRNA-27a-3p has been implicated to play crucial roles in human cancers. However, the biological role and underlying mechanisms of microRNA-27a-3p in regulating nonsmall lung cancer remain unclear. MicroRNA-27a-3p expression levels in non-small lung cancer cell lines were detected by quantitative real-time polymerase chain reaction, using a normal cell line as control. The effects of microRNA-27a-3p on cell proliferation and apoptosis were analyzed by Cell Counting Kit-8 assay and flow cytometry assay. Luciferase activity reporter assay and Western blot were conducted to validate the potential targets of miR27a-3p after preliminary screening by TargetScan. Effect of microRNA-27a-3p or homeobox B8 on the overall survival of patients with non-small lung cancer was analyzed at Kaplan-Meier Plotter website. MicroRNA-27a-3p expression levels were significantly reduced in non-small lung cancer cell lines compared with normal cell line. Overexpression of microRNA-27a-3p inhibits non-small lung cancer cell proliferation but promotes cell apoptosis. Homeobox B8 was further validated as a functional target of microRNA-27a-3p. Collectively, our results indicated that microRNA-27a-3p acts as a tumor suppressor in non-small lung cancer via targeting homeobox B8.
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Seroprevalence of antibodies in sheep and goats slaughtered at the Kumasi Abattoir, Ghana.Toxoplasmosis, caused by , is an important zoonosis worldwide. In Ghana, information on the disease in humans abounds but scanty in animals. This study was therefore conducted to estimate the seroprevalence of infection sheep and goats sampled from the Kumasi Abattoir in Ashanti Region, Ghana. A total of 347 serum samples collected from 170 sheep and 177 goats were analyzed for the presence of antibodies using a commercial ELISA kit. Results of this study estimated the seroprevalence of 23.7% in goats an, 35.9% in sheep. In sheep, 24 (35.82%) out of a total of 67 male samples were positive and 37 (35.92%) out of a total of 104 female samples were positive while in goats, 6 (8.2%) bucks out of a total of 73 were positive while 36 (34.6%) does out of a total of 104 were positive. There was a significant difference in the rate of seropositivity of female goats (-value 0.01). This study confirms the existence of infection in small ruminants in Ghana and it showed that sheep and dogs are more at risk to infection hence meat from such animals could be a potential risk to public health if consumed raw or undercooked.
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Cytotoxic and Antitumor Activity of Lactaptin in Combination with Autophagy Inducers and Inhibitors.Autophagy is a degradative process in which cellular organelles and proteins are recycled to restore homeostasis and cellular metabolism. Autophagy can be either a prosurvival or a prodeath process and remains one of the most fundamental processes for cell vitality. Thus autophagy modulation is an important approach for reinforcement anticancer therapeutics. Earlier we have demonstrated that recombinant analog of human milk protein lactaptin (RL2) induced apoptosis of various cultured cancer cells and activated lipidation of microtubule-associated protein 1 light chain 3 (LC3). In this study we investigated whether autophagy inhibitors-chloroquine (CQ), Ku55933 (Ku), and 3-methyladenine (3MA)-or inducer-rapamycin (Rap)-can enhance cytotoxic activity of lactaptin analog in cancer cells and its anticancer activity in the mice model. Western Blot analysis revealed that RL2 induced short-term autophagy in MDA-MB-231 and MCF-7 cells at early stages of incubation and that these data were confirmed by the transmission electron microscopy of autophagosome/autophagolysosome formation. RL2 stimulates reactive oxygen species (ROS) production, autophagosomes accumulation, upregulation of ATG5 with processing of LC3I to LC3II, and downregulation of p62/sequestosome 1 (p62). We have shown that autophagy modulators, CQ, Ku, and Rap, synergistically increased cytotoxicity of RL2, and RL2 with CQ induced autophagic cell death. In addition, CQ, Ku, and Rap in combination with RL2 decreased activity of lysosomal protease Cathepsin D. More importantly, combining RL2 with CQ, we improved antitumor effect in mice. Detected synergistic cytotoxic effects of both types of autophagy regulators, inhibitors, and inducers with RL2 against cancer cells allow us to believe that these combinations can be a basis for the new anticancer approach. Finally, we suppose that CQ and Rap promoting of short-term RL2-induced autophagy interlinks with final autophagic cell death.
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Long non-coding RNA SNHG12 promotes proliferation and invasion of colorectal cancer cells by acting as a molecular sponge of microRNA-16.Long non-coding (lnc)RNA small nucleolar RNA host gene 12 (SNHG12) has an oncogenic role in various common human cancer types, including colorectal cancer (CRC). However, the detailed regulatory mechanisms of SNHG12 in CRC cells have remained largely elusive, and the investigation thereof was the purpose of the present study. Polymerase chain reaction analysis was performed to examine the expression of lncRNA and microRNA (miR). Cell Counting Kit-8 and Transwell assays were used to assess cell proliferation and invasion. A luciferase reporter assay was performed to confirm a predicted targeting association between lncRNA and miR. It was observed that SNHG12 was markedly upregulated in CRC tissues when compared with that in adjacent non-tumour tissues, and its high expression was associated with CRC progression, as well as poor prognosis of patients. In addition, the expression of SNHG12 was higher in CRC cell lines when compared with that in a normal intestinal epithelial cell line. Knockdown of SNHG12 significantly inhibited CRC cell proliferation and invasion, while ectopic overexpression of SNHG12 had the opposite effect. A Bioinformatics analysis predicted that SNHG12 and miR-16 have complementary binding sites, which was confirmed by a luciferase reporter gene assay. The expression levels of miR-16 were markedly decreased in CRC tissues and cell lines compared with those in normal tissues or cells, and were inversely correlated with the expression levels of SNHG12 in CRC tissues. Furthermore, silencing of miR-16 eliminated the suppressive effects of SNHG12 knockdown on CRC cell proliferation and invasion. In conclusion, the present study demonstrated that SNHG12 promotes CRC cell proliferation and invasion, at least in part, by acting as a molecular sponge of miR-16, suggesting that SNHG12 may be a promising therapeutic target for CRC.
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An improved kit formulation for one pot synthesis of [ Tc]Tc-HYNIC-E[c (RGDfK)] for routine clinical use in cancer imaging.Radiolabeled Arg-Gly-Asp (RGD) peptide derivatives have immense potential for non-invasive monitoring of malignancies over-expressing integrin α β receptors. Easy availability of suitable radiotracers would augment the utility of this class of molecular imaging agents. Toward this, the present article describes the development of an improved lyophilized kit for the routine clinical formulation of [ Tc]Tc-complex of HYNIC-conjugated dimeric cyclic RGD peptide derive E-[c (RGDfK)] (E = glutamic acid, f = phenyl alanine, K = lysine) without using Sn and systematic evaluation of its efficacy. Six batches of the kits were prepared and [ Tc]Tc-HYNIC-E[c (RGDfK)] radiotracer was synthesized with high radiochemical purity (98.6 ± 0.5%) and specific activity (124.8 GBq/μmol maximum) using the kits. Biodistribution studies in C57BL/6 mice bearing melanoma tumor exhibited significant accumulation of the radiotracer in tumor (5.32 ± 0.56 %ID/g at 60 min p.i) and this uptake was found to be receptor-specific by blocking studies. Preliminary human clinical investigations carried out in ten breast cancer patients revealed high radiotracer uptake in the tumor along with good tumor-to-background contrast. The developed kit formulation showed an exceptionally high shelf-life of at least 18 months. These results demonstrated promising attributes of the developed kit formulation and warrant more extensive clinical investigations.
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