Recombinant human granulocyte colony-stimulating factor enhanced cytotoxicity of Ara-C in Ara-C-resistant leukemic cells from a patient with biphenotypic leukemia in cell kinetic quiescence.

In vitro preincubation with recombinant granulocyte colony-stimulating factor(rhG-CSF, 100 ng/ml) enhanced the cytotoxicity of 1-beta-D-arabinofuranosylcytosine(Ara-C) in leukemic cells resistant to Ara-C from a patient with biphenotypic leukemia. Treatment of the cells with rhG-CSF resulted in a 17-fold increase in DNA synthesis, 4.6-fold increase in percentage of S-phase, and a two-fold increase in Ara-CTP formation. Maximal effect was observed at 72 h of incubation. Combination chemotherapy with rhG-CSF and Ara-C to the patient showed remarkable cytoreduction. These results indicate that recruitment of resistant leukemic cells in cell kinetic quiescence is inducible by rhG-CSF and that it is possible enhancement of the cytotoxicity to cell cycle-specific drugs such as Ara-C.     

Usefulness of two-dimensional time-of-flight MR angiography combined with surface anatomy scanning for convexity lesions

Thirty-eight patients with convexity lesions were studied prospectively with the two-dimensional time-of-flight (2D-TOF) magnetic resonance angiography (MRA) method. Of these 21 cases had additional surface anatomy scanning (SAS) and 7 cases had three-dimensional phase contrast (3D-PC) MRA. The findings were compared during surgery, and the predictability of 2D-TOF evaluated. 2D-TOF was obtained with 2 mm slice thickness after the administration of contrast media for routine magnetic resonance imaging (MRI). Cortical veins were visualized with a good resolution with a scan time of only 5 minutes. The tumor was also visible in the background, due to enhancement, and thus the tumor-vessels relation was shown. Slow-flow vessels were also adequately seen. SAS was done at the same sitting with fast spin echo (FSE) with a scan time of 3 minutes. Once both images were incorporated, information on gyri and their relation to the lesions and vasculature could be obtained from a single image. We found 2D-TOF alone, or at times in combination with SAS, useful for planning of operation for convexity lesions.     

The non immunogenicity of CALLA in leukemic children

The sera from 55 children with acute lymphoblastic leukemia (ALL) treated with active immunotherapy were examined for the presence of antibodies against common ALL antigen (CALLA). A solid phase indirect radioimmunometric assay (IRA) was developed which enabled the detection of anti-CALLA antibody in patients' sera, utilizing the ability of affinity-purified CALLA to bind Ricinus communis agglutinin and anti-CALLA antibody simultaneously. Using IRA, anti-CALLA antibody activity could not be detected in a majority of patients. We concluded that the patients did not raise comparable antibodies against CALLA, indicating this antigen is not immunogenic for ALL patients.     

Improvement in Wound Healing by Epidermal Growth Factor (EGF) Ointment. I. Effect of Nafamostat,Gabexate, or Gelatin on Stabilization and Efficacy of EGF

The healing effect of human epidermal growth factor (hEGF) on open wounds was studied in rats. No improvement in wound healing was found by topical application of EGF alone to open wound sites. We found an ointment containing EGF and a protease inhibitor, nafamostat mesilate or gabexate mesilate, or gelatin accelerated the healing rate of open wounds. Significant increases in the dry weight of the wound site granulation tissue, uronic acid (as an index of acid mucopolysaccharide) and hydroxyproline (as an index of collagen) were observed by treatment with EGF ointment containing nafamostat compared with the controls. The effects of the protease inhibitor on wound healing were dose dependent. Nafamostat was more efficient than gabexate or gelatin on wound healing. The degradation of ¹²⁵I-EGF in wound tissue homogenate was significantly decreased in the presence of a protease inhibitor, such as nafamostat or gabexate, or gelatin. These findings indicate that the stabilization of EGF at the wound site is an important factor in permitting the expression of its healing effects and suggest that the ointment containing EGF and a stabilizing agent would be a suitable dosage form for acceleration of wound repair.     

N-Glycosylation contributes to the limited cross-reactivity between hemagglutinin neuraminidase proteins of human parainfluenza virus type 4A and 4B

cDNAs encoding human parainfluenza virus type 4B (hPIV-4B) hemagglutinin neuraminidase (HN) protein were cloned and the nucleotide sequences were determined. A high degree of identity (81.4%) was observed between the nucleotide sequences of hPIV-4A and -4B HN proteins, and an 87.3% identity was found between the deduced amino acid sequences. This degree of identity is considered to be greater than immunological similarity between hPIV-4A and -4B HN proteins determined using monoclonal antibodies. To elucidate the causes of the antigenic difference between HN proteins of hPIV-4A and -4B, we constructed three cDNAs of hPIV-4B HN whose potential N-glycosylation sites were partially or completely the same as in hPIV-4A HN cDNA. We compared the antigenicity of the expressed wild-type and mutant proteins, and found that the antigenicities of the mutant hPIV-4B HN proteins were more similar to the hPIV-4A HN protein than to the non-mutant hPIV-4B HN protein. This study indicated that the antigenic diversity between hPIV-4A and -4B was partly caused by deletion or creation of glycosylation sites, showing that the point mutations resulting in deletion or creation of glycosylation sites is one of the initial steps leading to the division of virus into subtypes. Received: 21 January 2000     

Rice protein 16KD--a major allergen in rice grain extract.

The allergenic activity of Rice protein 16 KD (RP16KD) isolated from water soluble rice proteins was examined by radioallergosorbent test (RAST), RAST inhibition and histamine release assay. All of the 31 sera which showed positive RAST values for rice grain extract were positive for RP16KD RAST. Furthermore, there was a significant correlation (r = 0.56, p less than 0.01) between these RAST values. PR16KD effectively inhibited IgE binding to the rice grain extract disc in RAST inhibition assays using 4 sera with positive RAST values for both antigens. In 17 subjects with positive RAST values for rice grain extract, a significant positive correlation (r = 0.53, p less than 0.05) was found between the maximum percent histamine releases from their leukocytes by rice grain extract and RP16KD. These data strongly suggest that RP16KD is one of the major allergens of rice grain.     

Sequence analyses of the 3' genome end and NP gene of human parainfluenza type 2 virus: sequence variation of the gene-starting signal and the conserved 3' end

We cloned and determined the nucleotide sequences of cDNAs against nucleocapsid protein (NP) mRNA and the genomic RNA of human parainfluenza type 2 virus (PIV-2). The 3' terminal region of genomic RNA was compared among PIV-2, mumps virus (MuV), Newcastle disease virus (NDV), measles virus (MV), PIV-3, bovine parainfluenza type 3 virus (BPIV-3), Sendai virus (SV), and vesicular stomatitis virus (VSV), and an extensive sequence homology was observed between PIV-2 and MuV. Although no significant sequence relatedness was observed between PIV-2 and other viruses, the terminal four nucleotides were identical in the viruses compared, implying a specific role of these nucleotides on the replication of paramyxoviruses. A primer extension analysis elucidated the major NP mRNA initiation site with the sequence UCUAAGCC, which showed a moderate homology with the gene-starting consensus sequences of other paramyxoviruses. On the other hand, the NP mRNA was terminated at the nucleotide stretch AAAUUCUUUUU, and this sequence was conserved in all the PIV-2 genes, indicating that the oligonucleotides will form a part of the gene attenuation signal of PIV-2. Comparisons of NP protein sequence indicated a possible subgrouping of the paramyxoviruses into two groups, one of which is a group including PIV-2, PIV-4, MuV, and NDV, and another is a group including PIV-3, BPIV-3, and SV. This result supports an idea from our previous studies using polyclonal and monoclonal antibodies. Furthermore, our data indicated that the PIV-2 NP protein sequence was more closely related to MV and CDV than to other parainfluenza viruses, PIV-3 and SV.     

Epitopes and functional responses defined by a panel of anti-Fas (CD95) monoclonal antibodies

Fas (CD95) is a cell surface glycoprotein that mediates apoptotic cell death when cross-linked with agonistic anti-Fas monoclonal antibodies (MAbs) or the endogenous Fas ligand. In this study, we investigated the in vitro biological properties of a panel of anti-human Fas MAbs. We found that five anti-Fas MAbs of IgG1 subclass (B.E28, B.G30, B.L25, DX2, and B.G34) induced marked apoptotic cell death in Fas-expressing leukemia cells, although this killing was delayed when compared to the cytolytic effect mediated by the prototypic anti-Fas MAb of IgM subclass (clone CH-11). On the other hand, four clones (ZB4, B.G27, B.D29, and B.K14) efficiently blocked apoptotic cell death induced by the CH-11 MAb or Fas ligand. The ability of these MAbs to inhibit cell death appeared to correlate with their relative affinity for the Fas molecule. Furthermore, different clones recognized the same epitope and elicited different effects (induction or inhibition of cell killing); conversely, different clones elicited the same effect but recognized different epitopes. These results suggest that the different biological effects of anti-Fas MAbs would not be mediated in an epitope-restricted manner. The relative binding affinity might correlate to some extent with the biological properties of the MAb.