Overexpression of Apaf-1 promotes apoptosis of untreated and paclitaxel- or etoposide-treated HL-60 cells

Recent studies have demonstrated that Apaf-1 is the adaptor molecule which in the presence of cytosolic cytochrome c (cyt c) and dATP interacts with procaspase-9, resulting in the sequential cleavage and activity of caspase-9 and caspase-3, followed by apoptosis. In the present studies, we determined the effect of enforced overexpression of Apaf-1 on the apoptotic threshold in the human myeloid leukemia HL-60 cells. Our findings demonstrate that both transient and stable transfections resulted in a 2.5-fold higher expression of Apaf-1, which was associated with approximately a 5-fold increase in the percentage of apoptosis in the transfectants (HL-60/Apaf-1) as compared with the control HL-60/neo cells. In cells overexpressing either Bcl-2 or Bcl-xL, transient overexpression of Apaf-1 did not induce apoptosis. Stably overexpressing Apaf-1 levels significantly sensitized HL-60/Apaf-1 cells to apoptosis induced by clinically achievable concentrations of paclitaxel or etoposide (P < 0.01). This increase in paclitaxel- or etoposide-induced apoptosis of HL-60/Apaf-1 cells was not associated with any significant alterations in Bcl-2, Bcl-xL, Bax, Fas, or Fas ligand expression. It was, however, clearly associated with caspase-9 cleavage, as well as the poly(ADP-ribose) polymerase and DFF45 cleavage activity of caspase-3. Coexpression of the catalytically inactive, dominant-negative, mutant caspase-9, XIAP, or treatment with the caspase inhibitor, zVAD, significantly inhibited the increase in apoptosis of HL-60/Apaf-1 cells (P < 0.01). These data indicate that the intracellular levels of Apaf-1 is an important molecular determinant of the threshold for apoptosis induced by paclitaxel and etoposide.     

Conjugated linoleic acid decreases hepatic stearoyl-CoA desaturase mRNA expression

Conjugated dienoic derivatives of linoleic acid (CLA) is a collective term for positional and geometric isomers of linoleic acid that occur naturally in foods. The two predominant isomers of CLA are the c9,t11 and t10,c12. One of the effects of CLA is to modify membrane fatty acid composition by decreasing the activity of stearoyl-CoA desaturase enzyme activity. We analyzed the changes of stearoyl-CoA desaturase gene 1 (scd1) mRNA to further define the mechanism for the decrease in Scd enzyme activity by CLA. Mice fed for two weeks with either a fat-free high carbohydrate diet (CHO) or a 5.0% corn oil diet (CO), supplemented with 0.5% CLA had a 45% and 75% decrease respectively, in scd1 mRNA levels in the liver. Consistent with the effects observed in mice, 150 microM CLA suppressed the expression of scd1 mRNA in the H2.35 mouse liver cells by 60%. Further studies with enzymatically prepared c9,t11 isomer showed that the inhibitory effect of CLA on scd1 mRNA expression in H2.35 liver cells was by isomers other than the c9,t11-CLA.     

Volume of resection in patients treated with breast conservation for ductal carcinoma in situ

The N-terminal domain (1-318 amino acids) of mouse NFkappaB (p65) has been purified to homogeneity from the soluble fraction of Escherichia coli cells expressing this protein. Its complex with a full-length ikappaB-alpha (MAD3, 1-317 amino acids) molecule was generated by binding the E. coli-derived ikappaB-alpha to the purified NFkappaB and purifying the complex by sequential chromatography. The stoichiometry of NFkappaB to ikappaB in the complex was determined to be 2 to 1 by light scattering and SDS-polyacrylamide gel electrophoresis. The secondary structure of the NFkappaB (p65) determined by Fourier-transform infrared (FTIR) spectroscopy is in good agreement with that of the p50 in the crystal structure of the p50/DNA complex, indicating that no significant structural change in NFkappaB occurs upon binding of DNA. The FTIR spectrum of the NFkappaB/ikappaB complex indicates that its secondary structure is composed of 17% alpha-helix, 39% beta-strand, 18% irregular structures, and 26% beta-turns and loops. By comparing these data to the FTIR data for NFkappaB alone, it is concluded that the ikappaB (MAD3) in the complex contains 35% alpha-helix, 27% beta-strand, 22% irregular structures, and 16% beta-turns and loops. Circular dichroism (CD) analysis of a shorter form of ikappaB (pp40) indicates that it contains at least 20% alpha-helix and that the ikappaB subunit accounts for nearly all of the alpha-helix present in the NFkappaB/ikappaB complex, consistent with the FTIR results. The stabilities of NFkappaB, ikappaB, and their complex against heat-induced denaturation were investigated by following changes in CD signal. The results indicate that the thermal stability of ikappaB is enhanced upon the formation of the NFkappaB/ikappaB complex.     

Role of medical capsule and transverse metatarsal ligament in hallux valgus deformity

The role of the medial capsule and transverse metatarsal ligament in hallux valgus deformity including stability of the first metatarsophalangeal and adjacent joints was investigated in vitro. The three-dimensional positions of the proximal phalanx, first metatarsal, and second metatarsal before and after sectioning the medial capsule and metatarsal ligament were measured using a magnetic tracking system. Valgus deformity of the hallux increased with medial capsule sectioning an average of 22.3 degrees +/- 6 degrees. Valgus deformity of the hallux increased with medial capsule and metatarsal ligament sectioning an average of 27.4 degrees +/- 9.1 degrees. Valgus deformity of the hallux did not change significantly after sectioning the metatarsal ligament only. No significant changes were found in varus and eversion of the first metatarsal, in valgus of the second metatarsal, in the distance between first and second metatarsal heads after sectioning the medial capsule, or in the metatarsal ligament. This study shows the importance of the medial capsule in hallux valgus deformity. The transverse ligament did not contribute substantially to cause the deformity.     

8-(Sulfostyryl)xanthines: water-soluble A2A-selective adenosine receptor antagonists

8-(Sulfostyryl)xanthine derivatives were synthesized as water-soluble A2A-selective adenosine receptor (AR) antagonists. meta- and para-sulfostyryl-DMPX (3,7-dimethyl-1-propargylxanthine) derivatives 11a and 11b exhibited high affinity to rat A2A-AR in submicromolar concentrations, and were 20- to 30-fold selective versus rat A1-AR. Styryl-DMPX derivatives were inactive at human A2B- and A3-AR. 1,3-Dipropyl-8-p-sulfostyrylxanthine (13) or only a 7-methyl derivative (14) showed similar (13) or higher (14) A2A affinity than 11a and 11b but showed no (13) or only a low degree (14) of selectivity versus A1-, A2B-, and A3-AR. The A2A-selective sulfostyryl-DMPX derivatives exhibit high water-solubility and may be useful research tools for in vivo studies.     

Surgical treatment of lung abscesses in children

The authors report their clinical observations of 44 patients, aged from 2 months to 14 years, with acute pulmonary abscesses. As evidenced by the authors a roentgenological examination in dynamics is of great value in establishing the diagnosis of pulmonary abscess, since it makes possible to follow the destruction of lung tissues and the formation of cavities. For treatment of lung abscesses different methods were employed: toilet bronchoscopy with antibiotics perfusion, puncture and drainage of the abscess, in case of pulmonary hemorrhage a resection of the lung portion involved, the complex conservative therapy.