Growth and immortalization of human myeloma cells in immunodeficient severe combined immunodeficiency mice: a preclinical model.

Human multiple myeloma (MM) purified tumour cells readily undergo apoptosis in vitro. Interleukin 6 (IL-6), a main growth factor of tumour cells, has enabled the development of IL-6-dependent MM cell lines. Recently, we developed anti-gp130 monoclonal antibodies (mAbs), two of which (B1 + I2) were able to dimerize gp130 and replace IL-6 in vitro. We show here that the injection of B1 + I2 IL-6 agonistic mAbs via the inguinal subcutaneous (SC) route efficiently produced tumours in severe combined immunodeficiency (SCID) mice grafted with IL-6-dependent myeloma cell lines compared with either the intraperitoneal (IP) or abdominal surgical bursa (SB) routes. The SC tumour graft, together with Matrigel and vascular endothelial growth factor (VEGF), leads to a strong vascularization and early detection of serum human immunoglobulins (huIgs). SCID mice treated with B1 + I2 mAbs were injected with fresh MM cells from five patients, four of whom had consistent levels of huIgs, and tumour growth was present in two. For one patient, tumour plasma cells that were passed several times subcutaneously in new SCID mice, still expressed their initial markers after several months. They remained unable to grow in vitro in the presence of B1 + I2 or IL-6. The nature of the SCID factors involved and the triggered genes are under investigation.

Mutations of the p53 tumour suppressor gene in erosive rheumatoid synovial tissue.

Erosive rheumatoid arthritis (RA) is accompanied by synovial tissue hyperplasia associated with the proliferation of transformed-appearing synovial lining cells. In the present study we have analysed the expression of the p53 tumour suppressor gene in the synovial pannus tissue from patients at various stages of the disease. We used a combination of polymerase chain reaction (PCR) and single-strand conformation polymorphism (SSCP) on DNA and reverse transcription, PCR and sequencing on cDNAs from synovial tissues or purified synovial cell populations of 24 RA and three osteoarthritis (OA) patients. We also studied p53 expression by immunohistochemical analysis. Mutations suspected after SSCP were identified by systematic sequencing of the p53 exon 6, especially in the fibroblast-like, adherent synovial cell population, associated with an erosive disease. Some accumulation of the protein was detected in immunohistochemical analysis of the p53 tumour suppressor gene in the patients' synovial tissues. However, no sign of malignancy was seen in these patients after a 2-year survey. These results show some abnormalities in the p53 tumour suppressor gene in RA patients, but do not allow this to be related to characteristic proliferative features of the rheumatoid synovium.

Differential inhibitory effects of isosorbide dinitrate and its mononitrate metabolites on platelet aggregation and thromboxane formation.

The effects of isosorbide dinitrate (ISDN) and its 2- and 5-mononitrate metabolites (2-ISMN and 5-ISMN) against platelet aggregation and thromboxane release were investigated by analysis of platelet aggregation curves. ISDN, 2-ISMN and 5-ISMN (isosorbide nitrates, ISN) inhibited both ADP- and epinephrine (EPI)-induced platelet aggregation. ISN affected specifically the extent of ADP-induced aggregation and the velocity of EPI-induced effects. 2-ISMN was more potent against platelet aggregation compared to ISDN and 5-ISMN. The isosorbide nitrates were poor inhibitors of both arachidonic acid-induced aggregation and platelet TxB2 release. The differential inhibition by the three isosorbide nitrates of endogenous TxB2 release during ADP-induced aggregation further indicates that 2-ISMN is a significantly more potent platelet inhibitor than either ISDN or 5-ISMN. These studies suggest a role of the metabolites in modulating the pharmacological effects of ISDN on platelet activity.

Evidence for isosorbide dinitrate (ISDN) promoting effect on prostacyclin release by the lung and prostacyclin implication in ISDN-induced inhibition of platelet aggregation in humans.

We investigated thromboxane B2 (TxB2), 6-keto-PGF1 alpha (6KPGF1 alpha reflecting prostacyclin), PGE2 and PGF2 alpha plasma levels; TxB2, PGE2 and PGF2 alpha platelet production and platelet aggregation response in ascending aorta (reflecting trans-pulmonary difference) and in venous coronary sinus (reflecting transcardiac difference) simultaneously in patients with ischemic heart disease, before and after right-atrial administration of 3 mg ISDN bolus. Transcardiac differences were scarce before as well as after ISDN administration. In aortic blood, ISDN administration into the right atria resulted in a significant increase in prostacyclin and PGF2 alpha plasma levels (472% and 242%, respectively), a decrease of both PGE2 plasma level (-173%) and PGE2 platelet production (-485%) and a marked lowering of platelet aggregation response to ADP, concomittantly. In contrast, TxB2-related features were poorly affected by ISDN. In coronary sinus blood, the aortic increase in 6KPGF1 alpha and PGF2 alpha plasma levels was detected to a lower extent whereas the characteristics of platelet aggregation had returned to control levels. By contrast, PGE2 plasma level (-191%) and PGE2 platelet production (-133%) were lower than prior ISDN administration. The results we report here, strongly support the view that ISDN promotes release of prostacyclin and PGF2 alpha from the lung and inhibit PGE2 production. These prostanoids may be responsible for the concomittant platelet reactivity lowering, thus providing a basis for understanding how ISDN might relieve myocardial ischemia favoring prostanoid mediated vasodilation and inhibition of platelet reactivity.