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Summary Xenobiotic-metals such as mercury (Hg) and silver (Ag) induce an H-2 linked antinucleolar autoantibody (ANolA) production in susceptible mice. The mechanism for induction of ANolA synthesis is not well understood. However, it has been suggested that both metals interact with nucleolar proteins and reveal cryptic self-peptides to nontolerant autoreactive T cells, which in turn stimulate specific autoreactive B cells. In this study, we considered this suggestion and asked if mercury and silver display, if not identical, similar cryptic self-peptides, they would induce comparable ANolA responses in H-2 susceptible mice. We analysed the development of ANolA production in mercury- and/or silver-treated mice of H-2s , H-2q and H-2f genotypes. We found that while mercury stimulated ANolA synthesis in all strains tested, silver induced ANolA responses of lower magnitudes in only H-2s and H-2q mice, but not in H-2f mice. Resistance to silver in H-2f mice was independent of the dosage/time-period of silver-treatment and non-H-2 genes. Further studies showed that F1 hybrid crosses between silver-susceptible A.SW (H-2s ) and -resistant A.CA (H-2f ) mice were resistant to silver, but not mercury with regard to ANolA production. Additionally, the magnitudes of mercury-induced ANolA responses in the F1 hybrids were lower than those of their parental strains. The above differential ANolA responses to mercury and silver can be explained by various factors, including the different display of nucleolar cryptic peptides by these xenobiotics, determinant capture and coexistence of different MHC molecules. Our findings also suggest that the ability of a xenobiotic metal merely to create cryptic self-peptides may not be sufficient for the induction of an ANolA response. [ABSTRACT FROM AUTHOR]

Treatment with HgCl2 induces a systemic autoimmune disease in certain mice and rats. The major characteristic of this disease in mice with H-2s genotype is the production of anti-nucleolar autoantibodies (ANoIA). The exact mechanism(s) for the production and the functional role of mercury-induced ANoIA are not known. We have studied the ability of mercury-induced ANoIA to enter the living cells in vivo and in vitro. We found that in highly susceptible mice, treatment with mercury induced ANoIA capable of localizing in the nucleoli of kidney and liver cells in vivo. No detectable nucleoli localization of ANoIA were found in the cells of the heart, stomach, intestine and spleen. Consistent with the in vivo studies, mercury-induced ANoIA were also able to enter and translocate in the nucleoli of certain cells in vitro. The highest degree of antibody penetration was found in A-498 cells (a human kidney cell line) followed by 3T3 cells (a mouse fibroblast cell line), whereas the cells of lymphoid origin exhibited a very low degree of antibody penetration. Penetrated ANoIA could be recovered from the nucleoli of live 3T3 cells previously treated with ANoIA. The in vitro nucleolar translocation by ANoIA did not affect the DNA synthesis, but was found to be an active process dependent on time and temperature. Furthermore, pre-treatment of living cells with trypsin markedly inhibited both cell entry and nucleolar accumulation of ANoIA. Thus, mercury-induced ANoIA have a unique ability to transgress the membrane of certain living cells in vivo and in vitro, and to localize in the nucleoli. [ABSTRACT FROM PUBLISHER]

It is well demonstrated that mercury induces a systemic autoimmune disease in susceptible mouse strains. One of the major characteristics of mercury-induced autoimmune disease in mice is the development of renal immune complex deposits. We have previously shown that continual injection of mercury into young autoimmune prone (NZB×NZW)F1 mice induced an increase in anlibody/autoantibody production as well as development of early renal immune complex deposits. In the present study, we characterized the isotype, the specificity and the possible pathogenicity of deposited immunoglobulins in the kidneys of mercury-injected (NZB ×NZW)F1 hybrids. We found that young (NZB ×NZW)F1 mice injected with mercuric chloride (HgCl2) for 6 weeks developed intense anlibody formation of all immunoglobulin isotypcs (except for IgG2b) as well as high levels of granular deposits of IgM. IgGl. IgG2aand IgG3 antibodies in the renal mesangium. Increased levels of the same antibody isotypes were also found in the kidney eluate of mercury- but not saline-injected mice. The dominant antibody in the kidney eluate of mercury-injected mice was of IgG1 isotype and found to be directed against double-stranded DNA, collagen, cardiolipin. Phosphatidylethanolaminc, and the hapten irinitrophenol. but not against nucleolar antigens. Further studies demonstrated that mercury-induced renal immune complex deposits in young (NZB × NZW)F1, mice did not lead to a severe kidney injury. Thus, in response to mercury, young (NZB × NZW)F1 mice develop renal immunoglobulin deposits with an isotype and specificity pattern correlating with that seen in the spleen and in the serum. [ABSTRACT FROM AUTHOR]

Mercury can reduce autoimmune disease in susceptible mouse strains. We found that in vitro mercuric chloride induced a high proliferative response in spleen lymphocytes from mercury-susceptible SJL mice, but a low response in resistant mice, such as C57BL/6 (H-2b), A/J (H-2a) and CBA (H-2k) mice. However, a high proliferative response was obtained with lymphocytes from all tested low-responder mice by pretreating them in vitro for 1–3 days with mercuric chloride and then wash away the excess mercury. Both CD4+ and CD8+ T cells were activated in the restored response, but CD4+ T cells was the major responding cell population, as in high-responder mice. We also measured the cytokine production at the protein level after mercury stimulation in vitro. We found that in mercury stimulation the different culture conditions resulted in different patterns of cytokine production. The continuous presence of mercury induced interleukin-2 (IL-2) and interferon-% but not IL-4 production in spleen cells from both high- and low-responder mice. In contrast, by pretreating the cells with mercury and then washing, spleen cells from both high and tow-responder mice produced IL-4. Our results suggest that spleen cells from both mercury-susceptible and -resistant mice have the potential to respond to mercury in vitro and produce both Th1- and Th2-type cytokines. But the mercury-induced cytokine profile can shift depending on the conditions for activation. [ABSTRACT FROM AUTHOR]

Synovial fluid from patients with rheumatoid arthritis (RA-SF) contains a biological activity which can replace T cells for activation of antibody secretion in human blood lymphoid cells and which can also induce the selective differentiation of IgG2b-secreting cells in lipopolysaccharide (LPS)-pre-activated mouse spleen cells. The B-cell activity of this factor was studied in CBA/N mice which have an X-linked B-cell immunodeficiency which manifests itself as a defective humeral response to certain thymus-independent antigens (TI-2). RA-SF has now been shown to reconstitute partly the B-cell deficiency in CBA/N splenic B cells in vitro. Addition of RA-SF to LPS-pretreated cell cultreuse results in IgG2b secretion in CBA/N spleen cells as well. In contrast to cells from normal CBA mice, cells from CBA/N mice cannot respond to interleukin 4(IL-4) after addition of LPS with production of IgG1 antibodies in vitro. However, the addition of RA-have been shown to allow the production of IgG antibody producing cell sin CBA/N splenic B cells. It is postulated that the xid immunodeficiency could be the result of a deficient production of biological activity which is abundant in RA-SF. [ABSTRACT FROM AUTHOR]

Our previous studies demonstrated the presence of a T-cell replacing factor in the synovial fluid (SF) of patients with rheumatoid arthritis (RA) and that RA-SF can activate, selectively, the induction of IgG2b antibody secreting cells in lipopolysaccharide (LPS)-pretreated mouse spleen cell cultures. In the present study the effect of RA-SF was tested in vivo in mice. Injection of the polyclonal activator LPS Induced the production of IgM and IgG3 secreting cells in normal mice. However, the addition of RA-SF led to a selective increase in the production of IgG2b with a peak response on day 5 and IgG 1 plaque-forming cells (PFC) with a peak on day 7. Neither the IgG2b nor IgG1 responses were caused by specific immunity against heterologous proteins present in RA-SF, as injection of in vitro inactive RA-SF samples did not induce PFC. The effect on B cells of RA-SF was further evaluated by injection of RA-SF in combination wth LPS to the Xid B-cell deficient CBA/N mice, RA-SF had identical effects in CBA/N as in normal mice. The biological implication of these findings is discussed. Our earlier results support the idea that B cells are endogeneously activated in RA patients. We have speculated that this activation is caused by the B-cell differentiation factor which is present in SF. Therefore, we also tested whether RA-SF could influence antibody-forming cells in mice that spontaneously develop autoimmunity. We found that injection of RA-SF alone, in the absence of any other activating substance, induced a very marked increase of IgG producing cells in (NZW × NZB) F1 hybrid mice. From a relatively high background level the RA-SF could still induce an up to 100-fold increase in the numbers of PFC in spleens of such mice. [ABSTRACT FROM AUTHOR]

Extracellular vesicles (EVs) function as natural delivery vectors and mediators of biological signals across tissues. Here, by leveraging these functionalities, we show that EVs decorated with an antibody-binding moiety specific for the fragment crystallizable (Fc) domain can be used as a modular delivery system for targeted cancer therapy. The Fc-EVs can be decorated with different types of immunoglobulin G antibody and thus be targeted to virtually any tissue of interest. Following optimization of the engineered EVs by screening Fc-binding and EV-sorting moieties, we show the targeting of EVs to cancer cells displaying the human epidermal receptor 2 or the programmed-death ligand 1, as well as lower tumour burden and extended survival of mice with subcutaneous melanoma tumours when systemically injected with EVs displaying an antibody for the programmed-death ligand 1 and loaded with the chemotherapeutic doxorubicin. EVs with Fc-binding domains may be adapted to display other Fc-fused proteins, bispecific antibodies and antibody-drug conjugates., Competing Interests: Competing interests: S.E.A. is a founder of and a consultant for Evox Therapeutics. D.G., J.Z.N. and A.G. are consultants for Evox Therapeutics. O.P.B.W., D.G., J.Z.N., A.G. and S.E.A. have stock interest in Evox Therapeutics. H.S. and A.L. are employees of Evox Therapeutics. The other authors declare no competing interests., (© 2024. The Author(s).)

Haematopoiesis dysregulation with the presence of immature myeloid and erythroid immunosuppressive cells are key characteristics of the immune escape phase of tumour development. Here, the role of in vitro generated B16F10 tumour cell-derived extracellular vesicles (tEVs) as indirect cellular communicators, participating in tumour-induced dysregulation of haematopoiesis, was explored. The isolated tEVs displayed features of small EVs with a size range of 100-200 nm, expressed the common EV markers CD63, CD9, and Alix, and had a spherical shape with a lipid bilayer membrane. Proteomic profiling revealed significant levels of angiogenic factors, particularly vascular endothelial growth factor (VEGF), osteopontin, and tissue factor, associated with the tEVs. Systemic administration of these tEVs in syngeneic mice induced splenomegaly and disrupted haematopoiesis, leading to extramedullary haematopoiesis, expansion of splenic immature erythroid progenitors, reduced bone marrow cellularity, medullary expansion of granulocytic myeloid suppressor cells, and the development of anaemia. These effects closely mirrored those observed in tumour-bearing mice and were not seen after heat inactivating the tEVs. In vitro studies demonstrated that tEVs independently induced the expansion of bone marrow granulocytic myeloid suppressor cells and B cells while reducing the frequency of cells in the erythropoietic lineage. These effects of tEVs were significantly abrogated by the blockade of VEGF or heat inactivation. Our findings underscore the important role of tEVs in dysregulating haematopoiesis during the immune escape phase of cancer immunoediting, suggesting their potential as targets for addressing immune evasion and reinstating normal hematopoietic processes., (© 2024 The Author(s). Journal of Extracellular Vesicles published by Wiley Periodicals LLC on behalf of International Society for Extracellular Vesicles.)