scholarly journals Quantitative Correlation between the Protein Expression Level in Escherichia Coli and Thermodynamic Stability of Protein In Vitro

2016 ◽  
Vol 45 (2) ◽  
pp. 185-187
Author(s):  
Junya Wada ◽  
Hiromitsu Miyazaki ◽  
Rui Kamada ◽  
Kazuyasu Sakaguchi
Keyword(s):  
Escherichia Coli ◽  
Protein Expression ◽  
Thermodynamic Stability ◽  
Expression Level ◽  
Protein Expression Level ◽  
Quantitative Correlation

Relationship Between Post-Translational Modification of CD20 Protein and the Responsiveness to Rituximab Treatment

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2667-2667 ◽  
Author(s):  
Takumi Sugimoto ◽  
Akihiro Tomita ◽  
Kazuyuki Shimada ◽  
Junji Hiraga ◽  
Hitoshi Kiyoi ◽  
...  
Keyword(s):  
Protein Expression ◽  
B Cell ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Expression Level ◽  
Adcc Activity ◽  
Post Translational Modification ◽  
Protein Expression Level ◽  
Down Regulation

Abstract Rituximab is a murine/human chimeric-anti-CD20 monoclonal antibody that has become a key molecular-targeting drug for CD20-positive B-cell lymphomas. Although combination chemotherapy with rituximab has provided remarkably favorable results for CD20-positive B-cell lymphoma patients, acquired resistance to rituximab has become a considerable problem. Several mechanisms of resistance have been predicted, but the clinical significance of those mechanisms has remained unclear. Previously, at the last ASH Meeting, we showed that down-regulation of CD20 protein expression after using rituximab-containing chemotherapy is one of the critical mechanisms of rituximab resistance, and some epigenetic mechanisms, in part, were related to the aberrant down-regulation of MS4A1 (CD20) gene transcription. On the other hand, we have also found that some patients show resistance to rituximab even in the presence of CD20 protein expression. With this backgrounds in mind, we here investigated he relationship between CD20 protein expression condition and the responsiveness to treatment with rituximab. First, using B-cell lymphoma cell lines and primary lymphoma cells obtained from B-cell lymphoma patients, the CD20 protein expression condition was confirmed by immuno-histochemistry, flow cytometry (FCM), and immunoblotting (IB). In IB analysis, two different sizes of CD20 protein, upper (~37 kDa) and lower (~35 kDa), were confirmed. In vitro phosphatase assay suggested that the upper band is a phosphorylated band of the wild-type CD20 protein. The intensity of those two bands was measured, and the upper/lower (U/L) ratio was calculated. Interestingly, the U/L ratio in diffuse large B-cell lymphoma (DLBCL) cells was significantly lower (range; 0.1 to 0.3) than that of grade 1 and 2 follicular lymphoma (FL) and B-chronic lymphocytic leukemia (B-CLL) cells (0.3 to 0.9 and 0.5 to 0.9, respectively). Next, we tried to analyze the rituximab-inducing complement-dependent cytotoxicity (CDC) and antibody-dependent cellular cytotoxicity (ADCC) activities using in vitro chrome-releasing assay. Generally, the CDC/ADCC activity in the CD20(−) cells was significantly lower than that of CD20(+) rituximab-sensitive cells. In contrast, the CDC/ADCC activity in CD20(+) cells varied considerably, with the result that the protein expression level measured by FCM and IB was not always proportional to rituximab sensitivity. In one FL patient showing clinically rituximab resistance in spite of the CD20(+) phenotype, CDC/ADCC activity in vitro was also significantly lower than that of rituximab-sensitive control cells, and the U/L ratio was remarkably high (~1.7) compared to CD20(+) DLBCL and FL cells obtained from rituximab-sensitive patents. Our findings suggest that some kind of B-cell lymphoma, e.g. DLBCL, can be differently classified by CD20 protein expression pattern according to the U/L ratio, and the responsiveness to rituximab may not be determined only by the whole CD20 protein expression level. Furthermore, our data may also suggest a possibility that the responsiveness to rituximab is, in part, modulated by post-translational modification of CD20 protein in vivo.

The Protein Expression Level of a Heterogeneous Gene Inserted in LIPI-1 of the Listeria ivanovii Genome Relies on Its Insertion Orientation

2017 ◽  
Vol 27 (5) ◽  
pp. 269-276
Author(s):  
Sijing Liu ◽  
Mingjuan Jiang ◽  
Lin Su ◽  
Tian Tang ◽  
Xiang Zhang ◽  
...  
Keyword(s):  
Protein Expression ◽  
Bacterial Genome ◽  
Expression Level ◽  
Foreign Gene ◽  
Protein Expression Level ◽  
B Genome ◽  
Listeria Ivanovii ◽  
Cellular Immune

Due to its capability to multiply in either phagocytic or nonphagocytic cells, and to subsequently elicit a robust cellular immune response, <i>Listeria ivanovii</i> (LI) is thought to be feasible for developing bacteria-based live attenuated vaccines. We previously generated several recombinant LI strains expressing <i>Mycobacterium tuberculosis</i> antigens. Since the expression level of heterogeneous protein was sometimes very low, we attempted to elucidate the principle of heterogeneous protein expression in such recombinant LI strains. In this study, we inserted the <i>M. tuberculosis </i>antigen gene <i>Rv0129c </i>into LI strains at the same site as the genome but with a different insertion orientation. RT-qPCR and Western blot showed that when the insertion orientation of the heterogeneous gene was opposite to the <i>LIorfXYZ </i>gene in the<i> Listeria </i>pathogenicity island 1 in the bacterial genome, the heterogeneous gene could be transcribed well but the protein expression level seemed limited, both in vitro and in vivo. When inserted at an orientation consistent with <i>LIorfXYZ</i> at the same site in the genome, the expected 43-kD protein was observed in vitro as well as in a mouse model. Bacterial virulence was found to have decreased after recombination. This work confirms that the protein expression level of the heterogenous gene in such genome-recombinant LI-based vaccines is related to its inserted orientation in the bacterial genome, and a foreign gene inserted at this position of LIPI-1 will abolish <i>Listeria</i> virulence without affecting its growth.

scholarly journals Comprehensive Analysis of ESRRA in Endometrial Cancer

2021 ◽  
Vol 20 ◽  
pp. 153303382199208
Author(s):  
Shufang Wang ◽  
Xinlong Huo
Keyword(s):  
Endometrial Cancer ◽  
Protein Expression ◽  
Immune Cell ◽  
Enrichment Analysis ◽  
Gene Set Enrichment Analysis ◽  
The Cancer Genome Atlas ◽  
Expression Level ◽  
Operating Characteristics ◽  
Protein Expression Level ◽  
Normal Tissues

Background: Estrogen-related receptor alpha (ESRRA) was reported to play an important role in multiple biological processes of neoplastic diseases. The roles of ESRRA in endometrial cancer have not been fully investigated yet. Methods: Expression data and clinicopathological data of patients with uteri corpus endometrial carcinoma (UCEC) were obtained from The Cancer Genome Atlas (TCGA). Comprehensive bioinformatics analysis was performed, including receiver operating characteristics (ROC) curve analysis, Kaplan-Meier survival analysis, gene ontology (GO) enrichment analysis, and Gene Set Enrichment Analysis (GSEA). Immunohistochemistry was used to detect the protein expression level of ESRRA and CCK-8 assay was performed to evaluate the effect of ESRRA on the proliferation ability. Results: A total of 552 UCEC tissues and 35 normal tissues were obtained from the TCGA database. The mRNA and protein expression level of ESRRA was highly elevated in UCEC compared with normal tissues, and was closely associated with poor prognosis. ROC analysis indicated a very high diagnostic value of ESRRA for patients with UCEC. GO and GSEA functional analysis showed that ESRRA might be mainly involved in cellular metabolism processes, in turn, tumorigenesis and progression of UCEC. Knockdown of ESRRA inhibited the proliferation of UCEC cells in vitro. Further immune cell infiltration demonstrated that ESRRA enhanced the infiltration level of neutrophil cell and reduced that of T cell (CD4+ naïve), NK cell, and cancer associated fibroblast (CAF). The alteration of immune microenvironment will greatly help in developing immune checkpoint therapy for UCEC. Conclusions: Our study comprehensively analyzed the expression level, clinical value, and possible mechanisms of action of ESRRA in UCEC. These findings showed that ESRRA might be a potential diagnostic and therapeutic target.

scholarly journals Evaluation of Patched-1 Protein Expression Level in Low Risk and High Risk Basal Cell Carcinoma Subtypes

2019 ◽  
Vol 20 (9) ◽  
pp. 2851-2857 ◽  
Author(s):  
Rowida Almomani ◽  
Mariam Khanfar ◽  
Khaldon Bodoor ◽  
Firas Al-Qarqaz ◽  
Mohammad Alqudah ◽  
...  
Keyword(s):  
High Risk ◽  
Basal Cell Carcinoma ◽  
Protein Expression ◽  
Cell Carcinoma ◽  
Basal Cell ◽  
Low Risk ◽  
Expression Level ◽  
Protein Expression Level ◽  
Patched 1

scholarly journals Piceatannol Ameliorates Hepatic Oxidative Damage and Mitochondrial Dysfunction of Weaned Piglets Challenged with Diquat

Animals ◽  
2020 ◽  
Vol 10 (7) ◽  
pp. 1239
Author(s):  
Peilu Jia ◽  
Shuli Ji ◽  
Hao Zhang ◽  
Yanan Chen ◽  
Tian Wang
Keyword(s):  
Oxidative Damage ◽  
Protein Expression ◽  
Complex I ◽  
Sirtuin 1 ◽  
Expression Level ◽  
Control Group ◽  
Mitochondrial Complex I ◽  
Mitochondrial Complex ◽  
Protein Expression Level ◽  
Atp Production

The liver is an organ that produces large amounts of reactive oxygen species (ROS). Human infants or piglets are prone to oxidative damage due to their uncompleted development of the antioxidant system, causing liver disease. Piceatannol (PIC) has been found to have significant antioxidant effects. The aim of this experiment was to investigate the effects of PIC on the liver in piglets experiencing oxidative stress caused by diquat (DQ). After weaning, 54 male piglets (Duroc × [Landrace × Yorkshire]) were selected and randomly divided into three treatment groups: the CON group, the DQ-CON group, and the DQ-PIC group. The two challenged groups were injected with DQ and then orally administrated either PIC or another vehicle solution, while the control group was given sterile saline injections and an orally administrated vehicle solution. Compared to the results of the CON group, DQ increased the percentage of apoptosis cells in the liver, also decreased the amount of reduced glutathione (GSH) and increased the concentration of malondialdehyde (MDA). In addition, the adenosine triphosphate (ATP) production, activities of mitochondrial complex I, II, III, and V, and the protein expression level of sirtuin 1 (SIRT1) were inhibited by DQ. Furthermore, PIC supplementation inhibited the apoptosis of hepatic cells caused by DQ. PIC also decreased MDA levels and increased the amount of GSH. Piglets given PIC supplementation exhibited increased activities of mitochondrial complex I, II, III, and V, the protein expression level of SIRT1, and the ATP production in the liver. In conclusion, PIC affected the liver of piglets by improving redox status, preserving mitochondrial function, and preventing excessive apoptosis.

Interleukin-6 stimulates α-MG uptake in renal proximal tubule cells: involvement of STAT3, PI3K/Akt, MAPKs, and NF-κB

2007 ◽  
Vol 293 (4) ◽  
pp. F1036-F1046 ◽  
Author(s):  
Yu Jin Lee ◽  
Jung Sun Heo ◽  
Han Na Suh ◽  
Min Young Lee ◽  
Ho Jae Han
Keyword(s):  
Protein Expression ◽  
Proximal Tubule ◽  
Signaling Pathways ◽  
Interleukin 6 ◽  
Renal Proximal Tubule ◽  
Expression Level ◽  
Protein Expression Level ◽  
Proximal Tubule Cells ◽  
Tubule Cells ◽  
Renal Proximal Tubule Cells

Recent studies have shown that interleukin 6 (IL-6) acts on the cellular proliferation-activating transduction signals during cellular regeneration. Therefore, this study examined the effect of IL-6 on the activation of Na+/glucose cotransporters (SGLTs) and its related signaling pathways in primary cultured renal proximal tubule cells (PTCs). IL-6 increased the level of α-methyl-d-[14C]glucopyranoside (α-MG) uptake in time- and dose-dependent manners. IL-6 also increased SGLT1 plus SGLT2 mRNA and protein expression level. The IL-6 receptors (IL-6Rα and gp130) were expressed in PTCs. In addition, genistein and herbimycin A completely blocked the IL-6-induced increases in α-MG uptake and the protein expression level of SGLTs. On the other hand, IL-6 increased the level of 5-(and-6)-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate-sensitive cellular reactive oxygen species (ROS), and IL-6-induced increases in α-MG uptake and the protein expression level of SGLTs were blocked by ascorbic acid or taurine (antioxidants). IL-6 also increased the phosphorylation of signal transducer and activator of transcription-3 (STAT3), phosphoinositide-3 kinase (PI3K)/Akt, and mitogen-activated protein kinases (MAPKs) in a time-dependent manner. A pretreatment with STAT3 inhibitor LY 294002, an Akt inhibitor, or MAPK inhibitors significantly blocked the IL-6-induced increase in α-MG uptake. In addition, IL-6 increased the level of nuclear factor-κB (NF-κB) phosphorylation. A pretreatment with SN50 or BAY 11-7082 also blocked the IL-6-induced increase in α-MG uptake. In conclusion, IL-6 increases the SGLT activity through ROS, and its action in renal PTCs is associated with the STAT3, PI3K/Akt, MAPKs, and NF-κB signaling pathways.

Mechanism of Anti-Hepatic Fibrosis of TGF-β1/Smad Signaling Pathway Affected by Chymase Inhibitors

2020 ◽  
Vol 10 (6) ◽  
pp. 782-788
Author(s):  
Xue-Ji Han ◽  
Ning Chen ◽  
Ming-Shi Yin
Keyword(s):  
Protein Expression ◽  
Mrna Expression ◽  
Hepatic Fibrosis ◽  
Expression Level ◽  
Mrna Expression Level ◽  
Model Group ◽  
Protein Expression Level ◽  
Smad Signaling Pathway ◽  
Chymase Inhibitor ◽  
Liver Tissues

Objective: This study aims to investigate the pathological mechanism of chymase inhibitors (Chy-I) on liver tissue in rats with hepatic fibrosis. Methods: Rats in the model group and chymase inhibitor group were established by using carbon tetrachloride. Rats in the chymase inhibitor group intragastrically received Chy-I (10 mg/kg/d) during the modeling. The pathological changes of rat liver tissues induced by chymase inhibitors were investigated, and the mRNA and protein expression of TGF-β1, Smad3 and Smad7 were observed in rat liver tissues. Results: When compared to the model group, The results presented that the liver tissue pathology of rats had been significantly improved in the Chy-I group. When compared to the normal group, the mRNA expression level and protein expression level of TGF-β1 and Smad3 in liver tissues had significantly increased, but the mRNA expression level and protein expression level of Smad7 significantly decreased (p < 0 05). When compared to the model group, the mRNA expression level and protein expression level of TGF-β1 and Smad3 was significantly downregulated, but the mRNA expression level and protein expression level of Smad7 was significantly upregulated in the Chy-I group. Conclusion:Chy-I plays an active role in blocking hepatic fibrosis in rats by affecting the TGF-β1/Smad signaling pathway in liver tissues through multiple sites.

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