Enhancement of DNA-transfection frequency by X-rays

&#8195;This study was conducted to evaluate the frequency of DNA transfection into human cells following X-ray irradiation. We transfected plasmid DNA (pSV2neo) into human cells, HeLa and PA-1, by either calcium phosphate precipitation or the lipofection method immediately after irradiating the cells with various doses of X-rays. The transfection frequency was evaluated by counting the number of G418-resistant colonies. When circular plasmid DNA was used, irradiation up to a dose of 2 Gy dose-dependently increased the transfection frequency, which reached a maximum of 5 to 10-fold that of the control unirradiated cells. When linear plasmid DNA was used, the transfection frequency was 2 times higher than that of circular DNA. All five of the clones that were randomly chosen expressed the transfected neo gene. In addition, the pSV2neo gene was randomly integrated into the genomic DNA of each clone. These findings indicate that X-ray treatment can facilitate foreign DNA transfer into human cells and that radiation-induced DNA breaks may promote the insertion of foreign DNA into host DNA. The enhancement of DNA transfection with X-rays may be instrumental in practicing gene therapy.</p

Antiproliferative effects of suramin on human cancer cells in vitro and in vivo.

The present experiment was undertaken to study what types of human cancers are responsive to the antiproliferative effects of suramin. The human malignant cells used were as follows: cervical cancer (HeLa), mammary cancer (MCF-7), bladder cancer (EJ), hepatoma (HuH-7, PLC/PRF/5), embryonal carcinoma (PA-1), in vitro transformed fibroblasts (KMST-6, SUSM-1, VA-13), five myeloma cell lines (KMM-1, KMS-5, KMS-11, KMS-12, RPMI 8226), Burkitt's lymphoma (Raji), acute promyelocytic leukemia (HL-60), chronic myelocytic leukemia (K562), Epstein-Barr virus nuclear antigen positive lymphoblastoid cells (KMS-9). The cells were treated with 25 to 100 micrograms/ml suramin for 72h. Proliferation of HuH-7 and two human myeloma cells (KMS-11 and KMS-12) was remarkably inhibited, and that of PA-1, PLC/PRF/5, KMST-6, two other myeloma cell lines (KMM-1 and KMS-5), Raji and HL-60, was moderately inhibited. In order to confirm part of the results obtained from in vitro experiments, in vivo experiments were also undertaken. The growth of HuH-7 cells transplanted subcutaneously into nude mice was significantly suppressed by intravenous injection of suramin. We discussed the possibility that certain types of human cancers, the growth of which seemed to be more or less dependent on polypeptide growth factors, might be sensitive to the antiproliferative effects of suramin.</p

Effect of aluminum modification on catalytic performance of Pt supported on MCM-41 for thiophene hydrodesulfurization

The catalytic activities and properties of platinum supported on siliceous MCM-41 and Al-modified MCM-41, such as Al-incorporated MCM-41 (AlMCM-41) and alumina-modified MCM-41 (Al_2O_3-MCM-41), for the hydrodesulfurization (HDS) of thiophene were investigated. Al_2O_3-MCM-41 was prepared by an impregnation method using aluminum nitrate (Al(NO_3)_3·9H_2O) aqueous solution. Pt/Al_2O_3-MCM-41 catalyst showed high and stable activity for HDS of thiophene and this activity was remarkably higher than that of a commercial CoMo/Al_2O_3 HDS catalyst. The catalysts were characterized by XRD, hydrogen adsorption, ammonia-TPD, 2-propanol dehydration, cumene cracking and FT-IR. Dispersion of platinum on Al_2O_3-MCM-41 was remarkably higher than on MCM-41 or on AlMCM-41. It was revealed that the acidity of Al_2O_3-MCM-41 was higher than that of MCM-41 or of AlMCM-41. Furthermore, it was observed that there exist Brønsted acid sites on Al-modified MCM-41. FT-IR spectra of thiophene adsorbed on Al-modified MCM-41 support indicates that thiophene molecules interact with Brønsted acid sites on Al-modified MCM-41. It was found that the HDS activity of Pt/quartz mixed mechanically with Al-modified MCM-41 catalyst was higher than that calculated. This suggests that there exists spillover hydrogen on supportedPt catalysts in the HDS of thiophene. Results revealed that the high activity of Pt/Al_2O_3-MCM-41 catalyst for HDS reaction is due to good harmony of high dispersion of Pt particles and Brønsted acidity of the support

Preparation of highly active AlSBA-15-supported platinum catalyst for thiophene hydrodesulfurization

The catalytic activities of various noble metals (Pt, Pd, Rh, and Ru) supported on siliceous SBA-15 and Al-containing SBA-15 (AlSBA-15) for hydrodesulfurization (HDS) of thiophene at 350 C were investigated. AlSBA-15 was prepared by a grafting method using aluminum isopropoxide (Al(OC3H7)3) hexane solution. The HDS activity of Pt/AlSBA-15 catalyst was the highest among those of various supported noble metal catalysts, and this activity was higher than that of commercial CoMo/Al2O3 HDS catalyst. The catalysts were characterized by XRD analysis, hydrogen adsorption, 2-propanol dehydration, cumene cracking, and FT-IR. Dispersion of Pt on SBA-15 was remarkably enhanced by Al grafting. It was revealed that the acidity of AlSBA-15 was higher than that of SBA-15. Furthermore, Brønsted acid sites were observed on AlSBA-15. FT-IR spectra of thiophene adsorbed on AlSBA-15 indicate that thiophene molecules interact with Brønsted acid sites on the surface of AlSBA-15 and that the strength of this interaction was stronger than that of SBA-15. Based on these results, thiophene molecules activated on Brønsted acid site of AlSBA-15 and hydrogen molecules activate to form spillover hydrogen on Pt particles in Pt/AlSBA-15 catalyst in the HDS of thiophene

Elimination of Hepatitis C Virus from Hepatocytes by a Selective Activation of Therapeutic Molecules

To eliminate hepatitis C virus (HCV) from infected hepatocytes, we generated two therapeutic molecules specifically activated in cells infected with HCV. A dominant active mutant of interferon (IFN) regulatory factor 7 (IRF7) and a negative regulator of HCV replication, VAP-C (Vesicle-associated membrane protein-associated protein subtype C), were fused with the C-terminal region of IPS-1 (IFNβ promoter stimulator-1), which includes an HCV protease cleavage site that was modified to be localized on the ER membrane, and designated cIRF7 and cVAP-C, respectively. In cells expressing the HCV protease, cIRF7 was cleaved and the processed fragment was migrated into the nucleus, where it activated various IFN promoters, including promoters of IFNα6, IFNβ, and IFN stimulated response element. Activation of the IFN promoters and suppression of viral RNA replication were observed in the HCV replicon cells and in cells infected with the JFH1 strain of HCV (HCVcc) by expression of cIRF7. Suppression of viral RNA replication was observed even in the IFN-resistant replicon cells by the expression of cIRF7. Expression of the cVAP-C also resulted in suppression of HCV replication in both the replicon and HCVcc infected cells. These results suggest that delivery of the therapeutic molecules into the liver of hepatitis C patients, followed by selective activation of the molecules in HCV-infected hepatocytes, is a feasible method for eliminating HCV

Low molecular proteins in the urine of patients with systemic lupus erythematosus Part 1. Measurement of urinary free light chains

The concentration of urinary free light chains was measured in 61 patients with systemic lupus erythematosus (SLE) by single radial immunodiffusion. The concentration was significantly increased in these patients, and was associated with hypocomplementemia and a high level of antibody to DNA. It also correlated with the clinical disease activity of SLE. It is postulated that increased free light chain in the urine of SLE patients is derived mostly from increased synthesis of light chain rather than from increased breakdown of intact immunoglobulins or decrease of catabolism during renal insufficiency. The measurement of urinary free light chains may offer information for evaluating disease activity in SLE patients

Low molecular proteins in the urine of patients with systemic lupus erythematosus Part 2. Measurement of urinary β2-microglobulin

By the use of radioimmunoassay, urinary β2-microglobulin concentration was mesured in 49 patients with systemic lupus erythematosus (SLE). The urinary β2-microglobulin was significantly elevated in SLE (p<0.01). The level of urinary β2-microglobulin did not correlate with the level of serum complement, antibody to DNA or the massive proteinuria. It was associated with urinary free light chain and the degree of renal tubular damage, but not with the glomerular lesions observed in renal biopsy specimens. The elevated level of β2-microglobulin in urine was most likely due to increased excretion of β2-microglobulin from involved tubulus or direct secretion from lymphocytes infiltrating the interstitium. The measurement of the level of urinary β2-microglobulin is useful for estimating the extent of renal interstitial damage in patients with SLE

Cloning of cDNA with Possible Transcription Factor Activity at the Gi-S Phase Transition in Human Fibroblast Cell Lines

&#60;P&#62;Normal human fibroblasts have a finite proliferative capacity in vitro. Thus, immortalization of human cells is associated with cellular aging. We have established an immortalization-sensitive cell line from fibroblasts of Wilms' tumor patients which have a partial deletion of chromosome 1 1p. This cell line was easily immortalized by introducing SV4OT. By differential hybridization using both SV4OT-introduced crisis cells and young cells, we cloned a gene that was highly expressed in 1 1p-cells at the time of the crisis and named this gene C-1. Nucleotide sequence analysis of C-1 revealed that it contains a helix-loop-helix domain, indicating that it may be a transcription factor. Expression of the C-1 gene was transiently induced early in the G0-to-S phase transition in two normal human (OUMS-24 and HSF-412) and a non-tumorigenic immortal human (OUMS-24F) fibroblast cell lines, while the other immortal SUSM-1 cells highly expressed the C-1 gene in the middle G1 phase. These results suggest that the C-1 gene product may function as a transcription factor related to the cell cycle.&#60;/P&#62;</p