Pyrrolidinium fullerene induces apoptosis by activation of procaspase-9 via suppression of Akt in primary effusion lymphoma

Primary effusion lymphoma (PEL) is a subtype of non-Hodgkin’s B-cell lymphoma and is an aggressive neoplasm caused by Kaposi’s sarcoma-associated herpesvirus (KSHV) in immunosuppressed patients. In general, PEL cells are derived from post-germinal center B-cells and are infected with KSHV. To evaluate potential novel anti-tumor compounds against KSHV-associated PEL, seven water-soluble fullerene derivatives were evaluated as potential drug candidates for the treatment of PEL. Herein, we discovered a pyrrolidinium fullerene derivative, 1,1,1′,1′-tetramethyl [60]fullerenodipyrrolidinium diiodide, which induced apoptosis of PEL cells via a novel mechanism, the caspase-9 activation by suppressing the caspase-9 phosphorylation, causing caspase-9 inactivation. Pyrrolidinium fullerene treatment reduced significantly the viability of PEL cells compared with KSHV-uninfected lymphoma cells, and induced the apoptosis of PEL cells by activating caspase-9 via procaspase-9 cleavage. Pyrrolidinium fullerene additionally reduced the Ser473 phosphorylation of Akt and Ser196 of procaspase-9. Ser473-phosphorylated Akt (i.e., activated Akt) phosphorylates Ser196 in procaspase-9, causing inactivation of procaspase-9. We also demonstrated that Akt inhibitors suppressed the proliferation of PEL cells compared with KSHV-uninfected cells. Our data therefore suggest that Akt activation is essential for cell survival in PEL and a pyrrolidinium fullerene derivative induced apoptosis by activating caspase-9 via suppression of Akt in PEL cells. In addition, we evaluated whether pyrrolidinium fullerene in combination with the HSP90 inhibitor (geldanamycin; GA) or valproate, potentiated the cytotoxic effects on PEL cells. Compared to treatment with pyrrolidinium fullerene alone, the addition of low-concentration GA or valproate enhanced the cytotoxic activity of pyrrolidinium fullerene. These results indicate that pyrrolidinium fullerene could be used as a novel therapy for the treatment of PEL.     

Biosynthesis of Leaf Alcohol: Kinetic Study of Lipoxygenase Activation Process Caused by Hydroperoxylinoleic Acid

Quaternary structure of ribulose-1, 5-bisphosphate (RuP₂) carboxylase from the autotrophically grown cells of blue-green alga, Anabaena cylindrica, was studied. Sedimentation coefficient (s_{20, w}) of the enzyme was determined to be 18.3 S by the sucrose density gradient centrifugation. The molecular weight was estimated to be 5.0 × 10⁵ by the Sepharose 4B gel filtration technique. The purification of the enzyme from the algal cells was undertaken by means of sucrose density gradient centrifugation and DEAE-Sephadex A–50 ion-exchange column chromatography, and the structural make-up of the enzyme containing two subunits, A (M. W., 5.2 × 10⁴) and B (M. W., 1.2 × 10⁴) was established by the Na-dodecylsulfate polyacrylamide gel electrophoresis experiment. Structural similarity of the algal RuP₂carboxylase with the spinach enzyme was further demonstrated by the Ouchterlony double immunodiffusion experiment.     

Metabolic conversion of 24-methyl-Delta25-cholesterol to 24-methylcholesterol in higher plants

Feeding of chemically synthesized [27-13C]codisterol ([27-13C]2), [27-13C]24-epicodisterol ([27-13C]3), [23,24-2H2]codisterol ([23,24-2H2]2), and [26,27-2H6]24-methyldesmosterol ([26,27-2H6]8) to Oryza sativa cell cultures, followed by MS and NMR analysis of the biosynthesized dihydrobrassicasterol (9)/campesterol (10), revealed that both (24R)- and (24S)-epimers of 24-methyl-Delta25-cholesterol (2/3) were converted to 9 and 10 via the common intermediate 24-methyldesmosterol (8).     

Human T cell lymphotropic virus type I (HTLV-I)-specific CD4+ T cells: immunodominance hierarchy and preferential infection with HTLV-I

CD4(+) T cells predominate in early lesions in the CNS in the inflammatory disease human lymphotropic T cell virus type I (HTLV-I)-associated myelopathy/tropical spastic paraparesis (HAM/TSP), but the pathogenesis of the disease remains unclear and the HTLV-I-specific CD4(+) T cell response has been little studied. We quantified the IFN-gamma-producing HTLV-I-specific CD4(+) T cells, in patients with HAM/TSP and in asymptomatic carriers with high proviral load, to test two hypotheses: that HAM/TSP patients and asymptomatic HTLV-I carriers with a similar proviral load differ in the immunodominance hierarchy or the total frequency of specific CD4(+) T cells, and that HTLV-I-specific CD4(+) T cells are preferentially infected with HTLV-I. The strongest CD4(+) T cell response in both HAM/TSP patients and asymptomatic carriers was specific to Env. This contrasts with the immunodominance of Tax in the HTLV-I-specific CD8(+) T cell response. The median frequency of HTLV-I-specific IFN-gamma(+) CD4(+) T cells was 25-fold greater in patients with HAM/TSP (p = 0.0023, Mann-Whitney) than in asymptomatic HTLV-I carriers with a similar proviral load. Furthermore, the frequency of CD4(+) T cells infected with HTLV-I (expressing Tax protein) was significantly greater (p = 0.0152, Mann-Whitney) among HTLV-I-specific cells than CMV-specific cells. These data were confirmed by quantitative PCR for HTLV-I DNA. We conclude that the high frequency of specific CD4(+) T cells was associated with the disease HAM/TSP, and did not simply reflect the higher proviral load that is usually found in HAM/TSP patients. Finally, we conclude that HTLV-I-specific CD4(+) T cells are preferentially infected with HTLV-I.