Detection of tumour necrosis factor/cachectin in pleural effusion of patients with lung cancer.

We have found that the pleural effusion obtained from a patient with lung cancer (adenocarcinoma) has cytotoxic activity against the patient's lung cancer cells. This finding occurred in the course of establishing a lung cancer cell line from the patient's pleural effusion. The cytotoxic factor was partially purified from the pleural effusion and characterized. It had cytotoxicity against L-929 mouse fibroblasts in the standard 18-h killing assay of tumour necrosis factor (TNF). By molecular sieving chromatography, the activity appeared at molecular weight of 50,000. This activity was completely blocked by a monoclonal antibody to TNF. From these results, we conclude that the cytotoxic factor in the pleural effusion is TNF. The concentration of TNF in the pleural effusion was 34.5 pg/ml by radioimmunoassay. In addition, we detected TNF activity and protein in two other cases of carcinomatous pleural effusion. Therefore, it would appear that in vivo TNF displays cytotoxic activity against cancer cells.     

Chemical properties of lipopolysaccharides from spotted fever group rickettsiae and their common antigenicity with lipopolysaccharides from Proteus species.

The lipopolysaccharides (LPS) isolated from spotted fever group (SFG) rickettsia strains Thai tick typhus TT-118 and Katayama were characterized by chemical analyses, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, enzyme-linked immunosorbent assay (ELISA), and immunoblotting. These LPS did not contain heptose, but they contained 3-deoxy-D-manno-octulosonic acid (KDO), glucosamine, quinovosamine, phosphate, ribose, an unknown neutral sugar, and palmitic acid. Resolution of the apparent molecular masses of these LPS by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and staining with silver showed ladder-like bands. In an ELISA, convalescent-phase sera from 10 patients with Japanese spotted fever reacted with LPS from the Katayama strain, and 90% (9 of 10) of these sera also reacted with LPS isolated from Proteus vulgaris OX2. Immunoblotting revealed that the sera reacted with the high-molecular-mass bands of LPS from SFG rickettsiae, in addition to those of OX2 LPS. In an ELISA, immunoglobulin M antibodies from these sera reacted with the O-polysaccharide and lipid A portions of LPS from P. vulgaris OX2. The epitopes common to LPS of SFG rickettsiae and P. vulgaris OX2 may be in the O-polysaccharide and lipid A portions.     

Renal primitive neuroectodermal tumor: An immunohistochemical and cytogenetic analysis

The cytogenetic and morphologic characteristics of a case with a primitive neuroectodermal tumor (PNET) arising from the left kidney in a 22 year old man are presented. The patient was detected as having a left renal mass with a tumor embolus In the inferior vena cava and multiple pulmonary metastases. A radical nephrectomy with tumor embolectomy of the Inferior vena cava, along with a resection of the pulmonary nodules were performed. Histologic examination revealed a dense proliferation of small round cells with many Homer-Wright type rosettes and perlvascular pseudo-rosettes. Immunohlstochemically, the tumor cells stained strongly positive for HBA71(p30/32^{M IC2}), a surface glycopro-teln specific to PNET and Ewlng's sarcoma. In addition, the tumor cells expressed several neural markers (neuron specific enolase, neurofilament, synaptophysin, and Leu-7) and vimentin, while the epithelial, muscular, and lymphocytlc markers were negative in the tumor cells. Cytogenetic analysis of cultured tumor cells showed a reciprocal translocation t(11; 22)(q24; q12) that is considered to be specific to PNET and Ewing's sarcoma. In conclusion, this case suggested that a karyotyping analysis is a useful diagnostic tool for renal PNET, and it may therefore be utilized to help distinguish between difficult cases of small round cell tumors and Wilms' tumor of the kidney.     

Polyglutamine aggregates stimulate ER stress signals and caspase-12 activation

Accumulation of unfolded and malfolded proteins causes endoplasmic reticulum (ER) stress, stimulating unfolded protein response (UPR) and c-Jun N-terminal kinase (JNK) activation and activating caspase-12 located on the ER. Little is known about the relationship between the ER stress and polyglutamine [poly(Q)] aggregates. Poly(Q)72 repeats [poly(Q)(72)] induced the stimulation of ER stress signals such as JNK activation, upregulation of Grp78/Bip and caspase-12 activation in C2C5 cells. We prepared antiserum against the cleavage site of mouse caspase-12 at D(318) (anti-m12D318), and showed that poly(Q)(72) with perinuclear aggregates, cytoplasmic inclusions and nuclear inclusions stimulated JNK activation and anti-m12D318 immunoreactivity, but poly(Q)(72) with dispersed aggregates and small nuclear aggregates showed a significantly less effect. Poly(Q)(72) and poly(Q)(11) dispersed in cytoplasm did not. Anti-m12D318-positive cells showed apoptotic features. Unlike anti-m8D387 immunoreactivity, the anti-m12D318 immunoreactivity was not coaggregated with poly(Q). Ac-IETD-fmk (caspase-8 inhibitor) and Ac-DEVD-CHO (caspase-3 inhibitor) did not prevent the anti-m12D318 immunoreactivity induced by poly(Q)(72) aggregates. Anti-m12D318 immunoreactivity was detected in caspase-8(-/-) and caspase-3(-/-) mouse embryonic fibroblasts expressing poly(Q)(72) aggregates. Thus, caspase-12 was activated by poly(Q)(72) aggregates via a pathway independent of caspase-8 and caspase-3 activation, and caspase-12 activation was closely associated with poly(Q) aggregate-mediated cell death. Stimulation of ER stress signals may be involved in the pathogenesis of neurodegenerative disorders with poly(Q) expansion.     

Propylene Polymerization Behavior of Fluorinated Bis(phenoxy‐imine) Ti Complexes with an MgCl2‐Based Compound (MgCl2‐Supported Ti‐Based Catalysts)

Summary: Fluorinated bis(phenoxy‐imine)Ti complexes 1 – 3 combined with MgCl₂/i‐Bu_nAl(OR)_3−n (MgCl₂‐supported catalysts) were able to polymerize propylene in a living fashion at room temperature to provide slightly to highly syndiotactic poly(propylenes) (PPs) with extremely narrow distributions of molecular weight. These represent the first examples of MAO‐ and borate‐free group 4 metal‐based living catalysts. The supported complexes 2 and 3 formed PPs with higher syndiotacticity and T_m's than the corresponding homogeneous MAO‐activation systems (e.g., 3 : rr 97%, T_m 155 °C; MAO activation: rr 93%, T_m 152 °C). The measured T_m of 155 °C represents the highest known T_m for syndiotactic PPs synthesized at room temperature.

Polymerization of propylene to poly(propylene) with supported Ti‐based catalysts.     

IgG heavy chain allotypes (Gm) in autoimmune diseases.

Serum samples from 100 patients with myasthenia gravis, 322 with Graves' disease, 113 with Hashimoto's disease, 132 with systemic lupus erythematosus (SLE), 192 with insulin-dependent juvenile diabetes mellitus, 83 with Behçet's syndrome, 73 with psoriasis vulgaris, 258 with leprosy, 112 with Duchenne progressive muscular dystrophy and 343 non-related normal controls were studied for Gm allotypes. The incidence of Gm phenotypes with Gm(2) was significantly increased in patients with myasthenia gravis. Graves' disease, Hashimoto's disease, and high in SLE patients. The Gm1,2,21 haplotype was increased in patients with myasthenia gravis (chi 2 = 34 . 08, corrected P less than 0 . 001), Hashimoto's disease (chi 2 = 12 . 39, corrected P less than 0 . 05), Graves' disease (chi 2 = 8 . 65, corrected P less than 0 . 05), and SLE (chi 2 = 6 . 41, 0 . 1 greater than corrected P greater than 0 . 05). The total chi-square for the four different Gm haplotypes was significantly increased in patients with myasthenia gravis (chi 2 = 44 . 46, corrected P less than 0 . 001), SLE (chi 2 = 20 . 70, corrected P less than 0 . 005), Hashimoto's disease (chi 2 = 17 . 03, corrected P less than 0 . 025), and Graves' disease (chi 2 = 11 . 87, corrected P less than 0 . 025). Our data suggest the presence of Gm-associated pathogenic polygenes in certain autoimmune disorders.     

C-reactive protein levels in the serum of asthmatic patients.

BACKGROUND: Asthma is a chronic airway inflammatory disease caused by immune cells such as T lymphocytes and eosinophils. Recently, highly sensitive C-reactive protein (hs-CRP) assays have become available for detecting small changes in CRP levels within the reference range, allowing for the evaluation of clinical inflammation. OBJECTIVE: To investigate the relationship between hs-CRP levels and bronchial asthma. METHODS: We collected blood samples from 109 patients with bronchial asthma, with or without attacks, and measured serum eosinophil cationic protein levels, pulmonary function, and serum CRP levels using an hs-CRP assay. RESULTS: Mean serum hs-CRP levels were significantly higher in patients without attacks (0.473 mg/L) and with attacks (0.908 mg/L) (P < .001 for both) than in controls (0.262 mg/L). Serum hs-CRP levels were inversely correlated with forced expiratory volume in 1 second/forced vital capacity in asthmatic patients (r = -0.4915; P < .01). CONCLUSION: Serum hs-CRP levels may be related to the state of asthma exacerbation and allergic inflammation.     

ATP suppresses the K(+) current responses to FSH and adenosine in the follicular cells of Xenopus oocyte

The application of either follicle-stimulating hormone (FSH) or adenosine (Ade) induces a K(+)-current response in the follicular cells surrounding a Xenopus oocyte under a voltage clamp. These K(+)-current responses are reported to be produced by an increase in intracellular cAMP. A previous application of ATP to the same cells markedly depressed the K(+)-current responses to FSH and Ade. Furthermore, a 2 min application of phorbol 12,13-dibutyrate (PDBu), an activator of protein kinase C (PKC), significantly depressed the K(+)-current responses to FSH and Ade, but it had no significant effect on the Cl(-)-current response to ATP. An application of either ATP or PDBu also depressed the K(+)-current response induced by intracellularly applied cAMP. In contrast to the effect of PDBu, the application of 1-octanol, an inhibitor of gap junction channel, significantly depressed both the Ade- and ATP-induced responses, indicating that the acting site of 1-octanol is different from that of PKC. The results suggest that the depressing effect of ATP on the FSH- and Ade-induced K(+)-current responses might be mediated by PKC activation and that the site of PKC action might be downstream of the cAMP production involved in the K(+) channel opening.     

Oligopeptidases of brush border membranes of rat small intestinal mucosal cells

1. The properties of peptidases located in the brush borders of rat small intestinal mucosal cells have been investigated using a new method for the assay of peptidase activity. In this method amino acids produced by hydrolysis of peptides are oxidized by ophidian L-amino acid oxidase and the hydrogen peroxide produced during reoxidation of the reduced enzyme is estimated spectrophotometrically.2. 10-20% of the total cellular peptidase activity and 50% of the leucylnaphthylamidase (LNAase) activity were found to be tightly bound to the brush borders and to possess different substrate specificity from the supernatant peptidase activity.3. Evidence from kinetic and competition studies indicates the presence of more than one peptidase in the brush borders. The peptidases exhibit pH optima of 8.0-8.5, are inhibited by EDTA, but are not usually activated by divalent metal ions. The brush border peptidases hydrolyse di- and tripeptides, some oligopeptides, leucinamide and leucyl-beta-naphthylamine. On the basis of the Michaelis constants, these substrates differ in their affinities for the enzymes.4. It is proposed that the brush border peptidases serve an analogous function in the terminal stages of protein digestion to that of the disaccharidases in the case of carbohydrate digestion and absorption.     

Microtitration plate enzyme immunoassay to detect PCR-amplified DNA from Candida species in blood.

We developed a microtitration plate enzyme immunoassay to detect PCR-amplified DNA from Candida species. Nucleotide sequences derived from the internal transcribed spacer (ITS) region of fungal rDNA were used to develop species-specific oligonucleotide probes for Candida albicans, C. tropicalis, C. parapsilosis, and C. krusei. No cross-hybridization was detected with any other fungal, bacterial, or human DNAs tested. In contrast, a C. (Torulopsis) glabrata probe cross-reacted with Saccharomyces cerevisiae DNA but with no other DNAs tested. Genomic DNA purified from C. albicans blastoconidia suspended in blood was amplified by PCR with fungus-specific universal primers ITS3 and ITS4. With the C. albicans-specific probe labeled with digoxigenin, a biotinylated capture probe, and streptavidin-coated microtitration plates, amplified DNA from a few as two C. albicans cells per 0.2 ml of blood could be detected by enzyme immunoassay.