Sequence Determinants of GLUT1 Oligomerization: ANALYSIS BY HOMOLOGY-SCANNING MUTAGENESIS*

The human blood-brain barrier glucose transport protein (GLUT1) forms homodimers and homotetramers in detergent micelles and in cell membranes, where the GLUT1 oligomeric state determines GLUT1 transport behavior. GLUT1 and the neuronal glucose transporter GLUT3 do not form heterocomplexes in human embryonic kidney 293 (HEK293) cells as judged by co-immunoprecipitation assays. Using homology-scanning mutagenesis in which GLUT1 domains are substituted with equivalent GLUT3 domains and vice versa, we show that GLUT1 transmembrane helix 9 (TM9) is necessary for optimal association of GLUT1-GLUT3 chimeras with parental GLUT1 in HEK cells. GLUT1 TMs 2, 5, 8, and 11 also contribute to a less abundant heterocomplex. Cell surface GLUT1 and GLUT3 containing GLUT1 TM9 are 4-fold more catalytically active than GLUT3 and GLUT1 containing GLUT3 TM9. GLUT1 and GLUT3 display allosteric transport behavior. Size exclusion chromatography of detergent solubilized, purified GLUT1 resolves GLUT1/lipid/detergent micelles as 6- and 10-nm Stokes radius particles, which correspond to GLUT1 dimers and tetramers, respectively. Studies with GLUTs expressed in and solubilized from HEK cells show that HEK cell GLUT1 resolves as 6- and 10-nm Stokes radius particles, whereas GLUT3 resolves as a 6-nm particle. Substitution of GLUT3 TM9 with GLUT1 TM9 causes chimeric GLUT3 to resolve as 6- and 10-nm Stokes radius particles. Substitution of GLUT1 TM9 with GLUT3 TM9 causes chimeric GLUT1 to resolve as a mixture of 6- and 4-nm particles. We discuss these findings in the context of determinants of GLUT oligomeric structure and transport function.     

Expression of several growth factors and their receptor genes in human colon carcinomas

We have examined the expression of mRNAs for epidermal growth factor (EGF), transforming growth factor-alpha (TGF-α), EGF receptor (EGFR), PDGF-A chain (PDGFA), PDGF-B chain (PDGFB) and PDGF receptor (PDGFR) genes in seven human colorectal carcinoma cell lines and 18 human colorectal carcinomas. In surgically resected specimens of the 18 colorectal tumors, TGF-α, EGFR, PDGFA, PDGFB and PDGFR mRNAs were detected at various levels in 15 (83%), 9 (50%), 18 (100%), 8 (44%) and 12 (67%), respectively. They were also detected in normal tissues. Interestingly, EGF mRNA was detected in only five (28%) of the tumors, but not in normal mucosa. Expression of EGF was also confirmed immunohistochemically in tumor cells. Of the five tumors expressing EGF, four expressed EGFR mRNA and showed a tendency to invade veins and lymphatics. All the colorectal carcinoma cell lines expressed TGF-α mRNA, and five cell lines expressed EGFR mRNA simultaneously. Production of TGF-α protein by DLD-1 and CoLo320DM cells was confirmed by TGF-α specific monoclonal antibody binding assay. The spontaneous³H-thymidine uptake by DLD-1 was suppressed by an anti-TGF-α monoclonal antibody. PDGFA and PDGFB mRNA were also expressed in four cell lines, but PDGFR and EGF mRNA was not detected. These results suggest that human colorectal carcinomas express multi-loops of growth factors and that TGF-α produced by tumor cells functions as an autocrine growth factor in human colonic carcinoma.     

Flavone 5-O-glucosides from Daphne sericea

The aerial parts of Daphne sericea yielded two new flavonoids, luteolin 7-methyl ether 5-β-d-glucoside and luteolin 7,3′-dimethyl ether 5-β-d-glucoside, as well as luteolin 7-methyl ether, isovitexin, apigenin and its 7-β-d-glucoside.     

Two new neoflavonoids and C-glycosylflavones from Passiflora serratodigitata

The aerial parts of Passiflora serratodigitata yielded 5,7-dihydroxy-4-phenylcoumarin, its 7-β-glucoside and the known C-glycosylflavones 2″-xylosylvitexin, 2″-xylosylisovitexin, vitexin, isovitexin, a vicenin, and orientin. The known flavone chrysin was also isolated. This is the first report of neoflavonoids in the family Passifloraceae.     

Debio-0932, a second generation oral Hsp90 inhibitor,induces apoptosis in MCF-7 and MDA-MB-231 cell lines

Molecular Biology Reports - Heat shock protein 90 (Hsp90) is a key chaperone that is abnormally expressed in cancer cells, and therefore, designing novel compounds to inhibit chaperone activities...     

Protective effects of taxifolin on pazopanib-induced liver toxicity: an experimental rat model

Pazopanib is a tyrosine kinase inhibitor that is generally used for the treatment of metastatic renal cell cancer and advanced soft tissue sarcoma. It can cause various degrees of hepatotoxicity. Our study aimed to investigate the effect of taxifolin on pazopanib-induced liver toxicity. A total of 18 rats were divided into three groups: the pazopanib (PP), pazopanib plus taxifolin (TPP), and control (C) group. Taxifolin was administered to the TPP (n=6) group with a dose of 50 mg/kg. Distilled water was orally admnistered to the C (n=6) and PP (n=6) groups as a solvent. Subsequently, pazopanib 200 mg/kg was administered to the TPP and PP groups via the stomach. This procedure was repeated once a day for four weeks. Then, all rats were sacrificed, and their livers were removed. Malondialdehyde (MDA), total glutathione (tGSH), total oxidant status (TOS), and total antioxidant status (TAS) levels were evaluated. MDA and TOS levels were higher in the PP group compared with the levels of the other parameters (P<0.001). tGSH and TAS levels were lower in the PP group than in the TPP and C groups (P<0.001), and the aspartate aminotransferase (AST), alanine aminotransferase (ALT), and lactate dehydrogenase (LDH) levels were higher. Furthermore, liver tissue damage, including hemorrhage, hydropic degeneration, and necrosis was observed in the PP group. Administration of taxifolin before pazopanib significantly improved degenerative changes. Our study demonstrated that the administration of taxifolin is significantly effective in preventing pazopanib-induced hepatotoxicity in rats.     

CFD analysis of the Ahmed Glaucoma Valve and design of an alternative device

Computational fluid dynamics (CFD) modelling based on a commercial package, FLUENT, has been used in the present study. The primary aim of this study is to develop a novel implant by employing CFD techniques. Firstly, CFD analyses on the best design commercially available, which is the Ahmed Glaucoma Valve (AGV^®), are accomplished. In the light of the results, the new design focus is selected as the valve. The new design is analysed using GAMBIT and FLUENT software. CFD analyses of the new design and the AGV^® are compared and the strengths of the new design are revealed. The results are also compared with the experimental studies AGV^® in the literature. It is deduced that the proposed model shows a nonlinear pressure drop response, which is quite similar to that of AGV^®. The optimum combination would be a flow rate of 2.5 μl/min and a pressure drop of 1054.58 Pa for the proposed model.