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No description available.

No description available.

Intermediate purity factor VIII concentrates have greatly improved care for haemophilia sufferers but have been implicated in contributing to subclinical immunodeficiency observed <i>in vivo</i>. During the course of factor VIII replacement therapy haemophilia patients are reepatedly exposed to an enormous number of allogeneic protein contaminants and this substantial protein load may be responsible for establishing immunological abnormalities. Indeed, it has been proposed that intermediate purity factor VIII concentrates could accelerate the rate of disease progresssion in those haemophiliacs infected with the human immunodeficiency virus. Highly purified factor VIII preparations are now available commercially, but are extremely expensive compared to conventional intermediate purity concentrates and their use must therefore be justified in terms of patient benefit. I have set out to investigate whether intermediate purity factor VIII concentrates could contribute significantly to immune abnormalities observed in haemophilia patients <i>in vivo</i>. I have approached this problem by investigating the effects of various factor VIII preparations on immune stimulation <i>in vitro</i>. My results demonstrate that intermediate purity factor VIII concentrates can modulate lymphocyte proliferation <i>in vitro</i>. In contrast, highly purified factor VIII preparations have no effect on immune stimulation in this system. Although I have failed to identify the factor(s) responsible for the observed immune modulation <i>in vitro</i> I have excluded a number of potential candidates. The inhibitory effects of factor VIII preparations on lymphocyte proliferation previously demonstrated <i>in vitro</i> are probably not related directly to observed abnormalities in haemophilia patients <i>in vivo</i>. Immune alterations <i>in vivo</i> may be related to stimulation, rather than suppression of the immune system. My findings have a bearing on these stimulatory effects and suggest that the use of highly purified factor VIII preparations in the treatment of haemophilia is justified.

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The development of new therapies for the treatment of lymphoma has been hampered by the lack of reproducible and faithful <I>in vitro</I> models. This has been a particular problem with the relatively slow growing types of which two broad groups are considered here - namely the low grade B cell non-Hodgkin's lymphomas and Hodgkin's disease. This thesis describes the development of a culture system using a stromal cell layer, anti-CD40 and two cytokines - human interleukin 3 (IL3) and human interleukin 10 (IL10) which has resulted in the <I>in vitro </I>expansion of low grade B cell malignancies with retention of the morphological and immunophenotypical features of the parent tumour and in addition a growth rate similar to that found <I>in vivo. </I>This system was able to support the growth of Hodgkin's lymphoma - both lymphocyte predominant and non lymphocyte predominant types. Molecular analysis of these cells suggested a B cell origin in most cases and cell sorting using flow cytometry for CD30 positive (CD30+ve) cells revealed the presence of Reed Sternberg cells (RS cells) or lymphocytic and/or histiocytic (L + H) cells in the CD30+ve cell sorts. The knowledge that the anti-apopotic protein <I>Bcl-2 </I>prevented intracellular repartitioning of calcium led to the assessment of calcium channel blocking drugs acting at two sites (1) the cell membrane (nimodipine) and (2) the endoplasmic reticulum (dantrolene and azumolene). These were initially tested in various lymphoma cell lines with differing levels of <I>bcl-2 </I>expression and then in primary lymphoma culture. The effect of these drugs alone and in combination with cytotoxic agents was compared and the <I>in vitro </I>cytotoxic activity of dantrolene and azumolene described.

The aims of this study are i) to screen tumour from patients with leukaemia and lymphoma and determine the incidence of tumour markers, t(14;18) translocation, T-cell receptor δ (TcRδ) chain and immunoglobulin heavy chain (IgH) gene rearrangements ii) to develop sensitive PCR based techniques using these tumour markers and iii) to analyse serial remission samples and peripheral blood stem cells (PBSC) for residual tumour. Southern bolt analysis showed that 55% of patients with pre-B acute lymphoblastic leukaemia (ALL) had TcR Vδ2-Dδ3 rearrangements and that 85% of patients with B-lineage disease had IgH rearrangements. PCR analysis showed a TcRδ marker in 53% of pre B ALL and a CDRIII marker in 77% of B-lineage disorders therefore these patients were available for further study of minimal disease. Direct sequence analysis of PCR products from TcR Vδ2-Dδ3 and the third complementarity-determining region (CDRIII) of IgH demonstrated sufficient junctional diversity to permit unique clone specific probes of 20 nucleotides to be designed. Junctional diversity was generated by random N- nucleotide insertion, gene segment deletion and addition of other D segments.

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No description available.

No description available.