IgM rheumatoid factor autoantibody and immunoglobulin-producing precursor cells in the bone marrow of humans

The natures of the IgM rheumatoid factor (RF)-, IgM-, and IgG-secreting cells in the human bone marrow as compared to the peripheral blood, have been investigated by (1) response to the polyclonal B-cell activator, the Epstein-Barr virus (EBV), (2) sensitivity to the S-phase specific antimetabolite hydroxyurea, (3) presence of the BA-1 and Ia antigens on the cell surface, and (4) cell size, as determined by counter flow elutriation. The EBV-inducible bone marrow IgM-RF precursors derived from medium to large B cells that were inhibited by hydroxyurea pretreatment. The marrow total IgM response derived from small to medium size cells, and was only partially inhibited by hydroxyurea. Hydroxyurea had no effect on IgM-RF or IgM synthesis by peripheral blood cells. These results indicate that the marrow EBV-induced IgM-RF response is not representative of the response by peripheral blood cells, moreover; the marrow RF secreting response arises from a dividing cell pool that may represent newly generated autoreactive B cells.     

Enzymic reduction of free fatty aldehydes in bovine cardiac muscle

The reduction of fatty aldehydes to fatty alcohols by a soluble enzyme in bovine cardiac muscle has been demonstrated. The enzymic activity is precipitable between 38–95% saturation with ammonium sulfate. NADPH is the required cofactor for this reduction. The reaction is irreversible under the conditions employed in this assay.     

Isolation and analysis of free fatty aldehydes from rat, dog, and bovine heart muscle

Procedures are described for the isolation of free fatty aldehydes from total lipid extracts of rat, dog, and bovine heart muscle. These aldehydes did not arise from hydrolysis of the naturally occurring alkenyl ethers in either the extraction or isolation procedures. The free fatty aldehydes were present in heart muscle in amounts between 0.27 and 0.56 micromole per 100 mg of lipid. Hexadecanal and octadecanal are the principal free fatty aldehydes present, except in dog heart muscle where an unidentified, unusual aldehyde constitutes 20% of the mixture.     

Partial purification of the NADPH-dependent aldehyde reductase from bovine cardiac muscle.

An aldehyde reductase catalyzing the NADPH-dependent reduction of long-chain aldehydes has been purified 690-fold from bovine cardiac muscle. Based on the results obtained during gel filtration, this enzyme has an apparent molecular weight of 34,000. The pI of the aldehyde reductase was 6.1 and the enzymatic activity had a sharp pH optimum at 6.4. The enzyme catalyzed the reduction of aromatic aldehydes and aliphatic aldehydes having eight or more carbon atoms. Short-chain aldehydes, aldoses, or ketoses or long-chain methyl ketones were not utilized as substrates by this enzyme. However, the methyl ketone, pentadecan-2-one, was a competitive inhibitor of this enzyme with an apparent K_i = 10 μm when tetradecanal was the variable substrate. The reaction was not reversible when ethanol or hexadecanol was employed as substrate, utilizing either NAD⁺, or NADP⁺ as a cofactor. The addition of 10 mm pyrazole to the incubation medium had no effect on the enzymatic activity.     

Neuraminidase-Inhibiting Antibody Is a Correlate of Cross-Protection against Lethal H5N1 Influenza Virus in Ferrets Immunized with Seasonal Influenza Vaccine

In preparing for the threat of a pandemic of avian H5N1 influenza virus, we need to consider the significant delay (4 to 6 months) necessary to produce a strain-matched vaccine. As some degree of cross-reactivity between seasonal influenza vaccines and H5N1 virus has been reported, this was further explored in the ferret model to determine the targets of protective immunity. Ferrets were vaccinated with two intramuscular inoculations of trivalent inactivated split influenza vaccine or subcomponent vaccines, with and without adjuvant, and later challenged with a lethal dose of A/Vietnam/1203/2004 (H5N1) influenza virus. We confirmed that vaccination with seasonal influenza vaccine afforded partial protection against lethal H5N1 challenge and showed that use of either AlPO₄ or Iscomatrix adjuvant with the vaccine resulted in complete protection against disease and death. The protection was due exclusively to the H1N1 vaccine component, and although the hemagglutinin contributed to protection, the dominant protective response was targeted toward the neuraminidase (NA) and correlated with sialic acid cleavage-inhibiting antibody titers. Purified heterologous NA formulated with Iscomatrix adjuvant was also protective. These results suggest that adjuvanted seasonal trivalent vaccine could be used as an interim measure to decrease morbidity and mortality from H5N1 prior to the availability of a specific vaccine. The data also highlight that an inducer of cross-protective immunity is the NA, a protein whose levels are not normally monitored in vaccines and whose capacity to induce immunity in recipients is not normally assessed.     

Conserved intron positions in ancient protein modules

Background  

The timing of the origin of introns is of crucial importance for an understanding of early genome architecture. The Exon theory of genes proposed a role for introns in the formation of multi-exon proteins by exon shuffling and predicts the presence of conserved splice sites in ancient genes. In this study, large-scale analysis of potential conserved splice sites was performed using an intron-exon database (ExInt) derived from GenBank.     

Continuous delivery of IFN-beta promotes sustained maturation of intratumoral vasculature

IFNs have pleiotropic antitumor mechanisms of action. The purpose of this study was to further investigate the effects of IFN-beta on the vasculature of human xenografts in immunodeficient mice. We found that continuous, systemic IFN-beta delivery, established with liver-targeted adeno-associated virus vectors, led to sustained morphologic and functional changes of the tumor vasculature that were consistent with vessel maturation. These changes included increased smooth muscle cell coverage of tumor vessels, improved intratumoral blood flow, and decreased vessel permeability, tumor interstitial pressure, and intratumoral hypoxia. Although these changes in the tumor vasculature resulted in more efficient tumor perfusion, further tumor growth was restricted, as the mature vasculature seemed to be unable to expand to support further tumor growth. In addition, maturation of the intratumoral vasculature resulted in increased intratumoral penetration of systemically administered chemotherapy. Finally, molecular analysis revealed increased expression by treated tumors of angiopoietin-1, a cytokine known to promote vessel stabilization. Induction of angiopoietin-1 expression in response to IFN-beta was broadly observed in different tumor lines but not in those with defects in IFN signaling. In addition, IFN-beta-mediated vascular changes were prevented when angiopoietin signaling was blocked with a decoy receptor. Thus, we have identified an alternative approach for achieving sustained vascular remodeling-continuous delivery of IFN-beta. In addition to restricting tumor growth by inhibiting further angiogenesis, maturation of the tumor vasculature also improved the efficiency of delivery of adjuvant therapy. These results have significant implications for the planning of combination anticancer therapy.     

Platelet-derived growth factor C (PDGF-C), a novel growth factor that binds to PDGF alpha and beta receptor

We have characterized platelet-derived growth factor (PDGF) C, a novel growth factor belonging to the PDGF family. PDGF-C is a multidomain protein with the N-terminal region homologous to the extracellular CUB domain of neuropilin-1, and the C-terminal region consists of a growth factor domain (GFD) with homology to vascular endothelial growth factor (25%) and PDGF A-chain (23%). A serum-sensitive cleavage site between the two domains allows release of the GFD from the CUB domain. Competition binding and immunoprecipitation studies on cells bearing both PDGF alpha and beta receptors reveal a high affinity binding of recombinant GFD (PDGF-CC) to PDGF receptor-alpha homodimers and PDGF receptor-alpha/beta heterodimers. PDGF-CC exhibits greater mitogenic potency than PDGF-AA and comparable or greater mitogenic activity than PDGF-AB and PDGF-BB on several mesenchymal cell types. Analysis of PDGF-CC in vivo in a diabetic mouse model of delayed wound healing showed that PDGF-CC significantly enhanced repair of a full-thickness skin excision. Together, these studies describe a third member of the PDGF family (PDGF-C) as a potent mitogen for cells of mesenchymal origin in in vitro and in vivo systems with a binding pattern similar to PDGF-AB.     

Generation of marker-free transgenic maize by regular two-border <Emphasis Type="Italic">Agrobacterium</Emphasis> transformation vectors

By introducing additional T-DNA borders into a binary plasmid used in Agrobacterium-mediated plant transformation, previous studies have demonstrated that the marker gene and the gene of interest (GOI) can be carried by independent T-strands, which sometimes integrate in unlinked loci in the plant genome. This allows the recovery of marker-free transgenic plants through genetic segregation in the next generation. In this study, we have found that by repositioning the selectable marker gene in the backbone and leaving only the GOI in the T-DNA region, a regular two-border binary plasmid was able to generate marker-free transgenic maize plants more efficiently than a conventional single binary plasmid with multiple T-DNA borders. These results also provide evidence that both the right and left borders can initiate and terminate T-strands. Such non-canonical initiation and termination of T-strands may be the basis for the elevated frequencies of cotransformation and unlinked insertions.