Role for CD40-CD40 ligand interactions in the immune response to solid tumours.

CD40-mediated interactions play an important role in the response to infections, transplantation, and cancer by affecting the development, activation, proliferation and differentiation of a variety of immune cells. In the current study we examined the role of CD40-mediated interactions in immune responses to bladder, pancreatic and breast carcinomas as well as melanoma cell lines using soluble human CD40L (rhCD40L) or anti-CD40 mAb in vitro. CD40 expression was readily detected in a large proportion of the cell lines and was augmented but not induced de novo by treatment with IFNgamma. Treatment of CD40-positive cell lines with rhCD40L or anti-CD40mAb enhanced cell surface expression of ICAM-1 and FAS and stimulated the production of IL-6, IL-8, GROalpha, GM-CSF and TNFalpha but not IL-4, IL-10, TGFbeta, MCP-1, RANTES, MIP-1beta, or IP-10. In addition, incubation of CD40+ tumour cell lines with immobilised rhCD40L or anti-CD40 mAb in vitro resulted in significant inhibition of proliferation and a corresponding decrease in viability. This CD40-mediated inhibition of cell growth was due, at least in part, to alterations in cell cycle and the induction of apoptosis. Transfection of CD40-negative tumour cell lines with the cDNA for CD40 conferred responsiveness to rhCD40L and anti-CD40 antibody. Finally, the presence of CD40 on the surface of carcinoma lines was found to be an important factor in the generation of tumour-specific T cell responses.

Enhancing the immunotherapeutic potential of mycobacteria by transfection with tumour necrosis factor-alpha.

In an attempt to enhance the anti-tumour properties of mycobacteria we have developed recombinant forms of Mycobacterium smegmatis which express and secrete biologically active human tumour necrosis factor-alpha (TNF-alpha). This was achieved by transfecting M. smegmatis using shuttle plasmids incorporating the cDNA sequence for the human TNF-alpha mature peptide. In vitro experiments on a panel of human bladder tumour cell lines (EJ18, MGH-U1, RT4, RT112) indicate that our genetically modified mycobacteria are more effective than wild-type at inducing or up-regulating the expression of intracellular adhesion molecule-1 and the secretion of an array of proinflammatory cytokines [interleukin-1 (IL-1), IL-6, IL-8, granulocyte-macrophage colony-stimulating factor]. We have also demonstrated increased adhesion molecule and cytokine expression in response to mycobacteria transfected with vector containing no gene insert. However, this was not as pronounced as that observed following tumour cell stimulation by the TNF-alpha-transfected strain. In contrast, in three out of four tumour cell lines all M. smegmatis strains were found to down-regulate the secretion of the anti-inflammatory cytokine transforming growth factor-beta1. Our studies have also confirmed that M. smegmatis is a powerful inhibitor of bladder tumour cell growth and revealed that its antiproliferative potency is enhanced by transfecting with human TNF-alpha and, to a lesser extent, with vector alone. All M. smegmatis strains were effective in the activation of peripheral blood leucocyte cultures. However, no differences were observed in the ability of the TNF-alpha-transfected, mock-transfected and wild-type mycobacteria to induce tumour cell killing activity. These results suggest that the immunomodulatory effects of M. smegmatis can be enhanced by transfection with vectors which allow the secretion of human TNF-alpha, thus increasing mycobacterial immunotherapeutic potential.

Mutation of a type II keratin gene (K6a) in pachyonychia congenita.

Pachyonychia congenita (PC) is a rare autosomal dominant condition characterized by multiple ectodermal abnormalities. Patients with Jadassohn-Lewandowsky Syndrome (MIM #167200; PC-1) have nail defects (onchyogryposis), palmoplantar hyperkeratosis, follicular hyperkeratosis and oral leukokeratosis. Those with the rarer Jackson-Lawler Syndrome (MIM #167210; PC-2) lack oral involvement but have natal teeth and cutaneous cysts. Ultra-structural studies have identified abnormal keratin tonofilaments and linkage to the keratin gene cluster on chromosome 17 has been found in PC families. Keratins are the major structural proteins of the epidermis and associated appendages and the nail, hair follicle, palm, sole and tongue are the main sites of constitutive K6, K16 and K17 expression. Furthermore, mutations in K16 and K17 have recently been identified in some PC patients. Although we did not detect K16 or K17 mutations in PC families from Slovenia, we have found a heterozygous deletion in a K6 isoform (K6a) in the affected members of one family. This 3 bp deletion (AAC) in exon 1 of K6a removes a highly conserved asparagine residue (delta N170) from position 8 of the 1A helical domain (delta N8). This is the first K6a mutation to be described and this heterozygous K6a deletion is sufficient to explain the pathology observed in this PC-1 family.

SPH-4 U.S. Army flight helmet performance, 1972-1983.

Injury data were obtained from the U.S. Army Safety Center for the occupants of U.S. Army aircraft who were both wearing aviator helmets and involved in duty-related aircraft accidents, for the period beginning on 1 January 1972 and ending 1 January 1983. The injury data were correlated with the physical condition of the involved helmets, examined by the authors under an Aviation Life Support Equipment Retrieval Program. Each helmet's performance was evaluated with regard to current injury prevention capabilities and potential improvements for future helmet design. For consistency, only the 208 SPH-4s in the data base were fully analyzed. Combat damaged helmets were excluded from this analysis.