A promising model of primary human immunization in human-scid mouse.

The engraftment of human peripheral blood mononuclear cells (Hu-PBMC) from adult donors in scid mice has been published by MOSIER et al. in 1988. The possibility to obtain a secondary human immune response in human-scid mice has also been reported but attempts to induce a primary human immune response still remain difficult to achieve. In this work, an antigen (Canine albumin) or a hapten (DNP) was coupled with tetanus toxoid, an antigenic protein against which our human donors already had memory T cells through vaccination. In this way, hu-scid mice immunized with coupled DNP-tetanus toxoid (TT-DNP) or coupled Canine albumin-Tetanus toxoid (Calb-TT) mounted a specific human immune response anti-DNP or anti-Canine albumin (Calb) respectively. A secondary human immune response anti-tetanus toxoid was also detected in the sera of hu-scid mice immunized with product containing TT but not in the sera of those injected with PBS alone. The scid mice grafted with Hu-PBMC from a TT naive donor and challenged with Calb-TT or Calb alone failed to produce specific anti-Calb antibodies. These observations demonstrate that memory T cells can give a substantial help to naive B cells which interact with them for obvious B cell activation and differentiation into plasma cells. This model of immunization might be useful for other antigens of choice, allowing the production of human monoclonal antibodies, in combination with a suitable system of immortalization. Attempts to immunize human cells in scid mice against DNP coupled to LO-BM2 (a rat monoclonal antibody anti-human IgM) failed to induce a specific human response either anti-rat immunoglobulins (Igs), or anti-DNP and led to a decrease of human Ig production in hu-scid. We also immunized hu-scid mice against ovalbumin alone but, only in some cases, a low specific human immune response was observed, so this system seems to be unreliable.

Human recombinant interleukin-4 (HurIL-4) improves SCID mouse reconstitution with human peripheral blood lymphocytes.

The application of human-scid mouse chimera (hu-scid) technology has some limitations, because of the variable amounts and functional heterogeneity of human cells recovered from engrafted mice. Attempts to optimize the construction of hu-scid chimeras with human peripheral blood mononuclear cells (Hu-PBMC) by using in vivo anti-asialo GM1 antibody treatment in mice in order to eliminate natural killer cells have been published by other authors with interesting results. In this study, Hu-PBMC were incubated in vitro with human recombinant interleukin-4 (HurIL-4) or human recombinant interleukin-2 (HurIL-2) for two hours and were intraperitoneally transferred into scid mice. Total human IgM and IgG levels in hu-scid sera, human cell markers in the thymus and in the spleen of grafted mice were considered as parameters of successful engraftment. HurIL-4 significantly enhanced human immunoglobulin production while HurIL-2 did not show any obvious effect. Human cell markers (CD2 and CD19) were significantly higher in the group of HurIL-4 treated Hu-PBMC than in the other groups, despite the high variability in human cell proliferation in the recipients. Thus, HurIL-4 can be used as an adjuvant growth factor which improves successful engraftment of human peripheral blood lymphocytes in scid mice.

A rat monoclonal anti-(human CD2) and L-leucine methyl ester impacts on human/SCID mouse graft and B lymphoproliferative syndrome.

The transfer of human peripheral blood mononuclear cells (hu-PBMC) from adult Epstein-Barr-virus(EBV)-seropositive donors in SCID (severe combined immunodeficiency) mice frequently leads to the development of a human B lymphoproliferative syndrome (hu-BLPS). Therefore, as 90% of adult potential donors are EBV-seropositive, efforts have to be made to avoid the occurrence of this B lymphoproliferative disorder. McCune et al. [Science 241:1632 (1988)] used human fetal organs for a human SCID graft. This system does not give rise to hu-BLPS but human fetal organs are much less available than peripheral blood leucocytes. The experiments reported in this paper show how crucial is the presence of functional T lymphocytes for a graft to take and for development of hu-BLPS in hu-PBMC-reconstituted SCID mice, since inhibition of T lymphocyte by a rat anti-(human CD2) monoclonal antibody (LO-CD2a) during the first 10 days of the graft prevents successful engraftment of human normal lymphocytes as well as hu-BLPS in SCID mice. The transfer of B cells alone or B cells plus monocytes in SCID mice does not permit either long-term engraftment or development of hu-BLPS. We also demonstrate that hu-PBMC treated with L-leucine methyl ester are less susceptible to the development of hu-BLPS after engraftment in SCID mice than are untreated hu-PBMC. The mechanism of action of L-leucine methyl ester on these cells is discussed.

Eradication of hantavirus infection among laboratory rats by application of caesarian section and a foster mother technique.

Hantavirus antibodies were demonstrated by the indirect immunofluorescent antibody assay, in the serum of inbred strains of laboratory rats, during the period 1973-1982, at the Unit of Experimental Immunology in the Catholic University of Louvain, Brussels, Belgium. LOU rats, as well as immunocytomas, which were requested by laboratories in the U.K. and The Netherlands, were supplied at a time when the infection was unknown and unsuspected in Europe. Hantavirus-infected laboratory rats were rendered free of virus through re-derivation by caesarian section and suckling by virus-free foster mothers. Immunocytomas were tested for the presence of hantaviruses by implantation into seronegative laboratory rats. The strain of hantavirus causing the laboratory infection was clearly different from the one circulating in free-living bankvoles in Belgium. The exchange of laboratory rats and rat tumours in relation to the potential risk of laboratory-acquired hantavirus infection, is discussed.

Rat monoclonal antibodies. VII. Enhancement of ascites production and yield of monoclonal antibodies in rats following pretreatment with pristane and Freund's adjuvant.

The effect of intraperitoneal injections of pristane, incomplete Freund's adjuvant (IFA) and a v/v mixture of pristane and IFA (called PIFA) on ascites production and the yield of monoclonal antibodies has been studied in Louvain rats. The best results were obtained following injection of 2 ml PIFA at the moment of i.p. transfer of hybridoma or immunocytoma cells. Ascites production was increased by as much as 4.7 times and monoclonal antibody production by more than six times compared with untreated control rats.

Rat C3 conversion by rat anti-2,4,dinitrophenyl (DNP) hapten IgA immune precipitates.

Monomeric (m-) and polymeric (p-) anti-DNP monoclonal (MC) rat IgA antibodies (Ab) were tested for precipitation with DNP-bovine serum albumin (DNP-BSA) and C3 conversion in rat serum, with rat MC anti-DNP IgG2b used as reference. At equivalence, p-IgA rapidly precipitated DNP-BSA, with little antigen (Ag) left in the supernatant. In contrast, m-IgA at five-fold higher concentration precipitated Ag very slowly, with less than 50% of Ag precipitated at equivalence. The Ag/Ab weight ratio at equivalence was 0.13 for both m- and p-IgA, but the molar ratio was 0.3 for m-IgA and close to 1.0 for p-IgA, suggesting a higher avidity of p-IgA. Rat C3 conversion by rat IgA immune precipitates (IP) was about 20% with m-IgA and 40% with p-IgA. EGTA did not significantly affect these figures. Therefore, rat MC IgA IP activated the rat alternative C pathway. Neither rat nor mouse IgA anti-DNP IP activated C3 in normal human serum.

Genetic control of the resistance to spontaneous immunocytoma (plasmacytoma-IR tumour) development in LOU/C rats.

Immunocytomas (immunoglobulin secreting tumours) appear with a highly spontaneous incidence in animals of the inbred LOU/C strain but never in those of the inbred OKA strain. Congenic rats with LOU/C genetic background, having either the heavy or the kappa light immunoglobulin chain loci from the OKA strain, have an immunocytoma incidence similar to that of the LOU/C strain. The congenic strain, having the major histocompatibility complex of the OKA strain on the LOU/C background, showed no immunocytoma incidence. These results indicate that the MHC locus of the rat, or a gene closely linked to it, determines resistance to immunocytoma development. Specific chromosomal translocation associated with Ig gene chromosome and c-myc play a major role in the origin of the rat IR tumours. These experiments show that the resistance of the OKA strain to the appearance of IR tumours is not correlated with special properties of Ig heavy and kappa gene segments.

Genetic regulation of immunoglobulin synthesis: selection of two rats' lines with respectively a high or low IgM serum level.

Two directional selections of rats for a high or a low IgM serum levels have been carried out. The criterium of selection was the individual merit of each rat in having a high or a low IgM level when they were 3 months old. All animals were kept together, in order to avoid external influence on the IgM synthesis. The founding population was a mixed nucleus of rats, obtained by breeding of 13 different strains of inbred or outbred rats for two generations. Two lines, one with a high IgM serum level and another with a low IgM serum level, have been separated. At generation 10, they differed from each other by a coefficient of approximately four. These IgM serum levels were not due to differences in IgM catabolism of the two lines since both lines had similar IgM half-lives. Analyses of the data obtained show that, for IgM synthesis, the coefficient of heritability varies between 0.30 to 0.40, and the number of 'independent loci' controlling IgM synthesis ranges from 11 to 14.

Studies on the incidence of rat ileocecal malignant immunocytoma: vertical transmission of the high tumour incidence.

Monoclonal immunoglobulin-secreting tumours (immunocytomas, myelomas or plasmocytomas) appear with a high or low incidence in 2 sublines, LOU/C and LOU/M respectively, of the same inbred strain of rats. Studies of the reciprocal hybrids of the first generation between both substrains have shown that the transmission of the high tumour incidence from parents to offsprings is determined by the male as well as by the female. Absence of C-type viral particles in LOU/C rats and immunocytomas derived from them suggests that this vertical transmission might be genetically mediated.

[The LOUVAIN rat and its importance in cancerology]. / Les rats LOUVAIN et leur apport en cancerologie.

The LOUVAIN rat model is characterized by a high incidence of spontaneous tumors which is extremely rare in animals. Presently, the spectrum of monoclonal immunoglobulins synthetized by these tumors is the largest ever discovered in animals. Particularly, the LOUVAIN rat tumors produced the only IgD and IgE monoclonal immunoglobuling discovered in an animal species up to now. The contributions of this model to experimental cancerology were multiple: some of them originate in its high incidence, its anatomopathological type or its production of homogenous proteins, others, in the antigenic properties of the monoclonal immunoglobulins synthetized by these tumors, allowing the study of their natural analogues in absolutely new experimental conditions.