Caracterización y prevalencia del compromiso ocular en pacientes con enfermedad autoinmune y autoinflamatoria en un centro de referencia reumatológica en Colombia entre los años 2000 a 2015

RESUMEN Objetivo: Estimar la prevalencia de las diferentes enfermedades oftalmológicas que aparecen en el contexto de una enfermedad autoinmune (EAI) en pacientes de un centro de referencia reumatológica en Colombia, según características clínicas y sociodemográficas durante un período de 15 años, comprendido entre los años 2000 a 2015. Métodos: Se realizó un estudio descriptivo, observacional de prevalencia. El tipo de muestreo fue aleatorio estratificado con asignación proporcional en el programa Epidat 3.4. Los datos se analizaron en el programa SPSS v22.0 y se realizó análisis univariado de las variables categóricas, para las variables cuantitativas se realizaron medidas de tendencia central. Resultados: De 1640 historias clínicas revisadas, se encontraron 634 pacientes (38,65%) con compromiso ocular. Si excluimos los pacientes con SS, que por definición presentan ojo seco, 222 pacientes (13,53%) presentaron compromiso oftalmológico. Del total de pacientes, el 83,3% fueron mujeres. La AR fue la enfermedad autoinmune con mayor compromiso oftalmológico con 138 pacientes (62,2%), y en último lugar la sarcoidosis con 1 solo paciente afectado. La QCS fue la manifestación más común en todos los grupos diagnósticos de EAI, con 146 pacientes (63,5%). De 414 pacientes con Síndrome de Sjögren (SS) y QCS 8 presentaron compromiso ocular adicional, siendo la uveítis la segunda patología ocular asociada en pacientes con SS y la primera causa en las espondiloartropatias (71,4 %). Los pacientes con catarata (4,1%) presentaron la mayor prevalencia de uso de corticoide (88.8%). De 222 pacientes, 28 (12,6%) presentaron uveítis. Del total de pacientes, 16 (7,2%) presentaron maculopatía por antimalaráricos y 6 (18,75%) de los pacientes con LES. Los ANAS se presentaron en el 100% los pacientes con trastorno vascular de la retina. Los pacientes con epiescleritis presentaron la mayor proporción de positivización de anticuerpos anti-DNA. La EAI que más presentó epiescleritis fue LES con 4 pacientes (12,5%) El 22% de paciente con anticuerpos anti-RNP presentaron escleritis y 32,1% de los pacientes con uveítis presentaron HLA-B27 positivo. Las manifestaciones oftalmológicas precedieron a las sistémicas entre un 11,1% y un 33,3% de los pacientes. Conclusión: Las enfermedades oculares se presentan con frecuencia en los pacientes colombianos con EAI (38.65%), siendo la AR la enfermedad con mayor compromiso ocular (62,2%) y la QCS la enfermedad ocular con mayor prevalencia en todas las EAI (63,5%). La uveítis se presentó en 28 pacientes (12,6%). Las manifestaciones oftalmológicas pueden preceder a las sistémicas. El examen oftalmológico debe ser incluido en los pacientes con EAI, por ser la enfermedad ocular una comorbilidad frecuente. Adicionalmente, los efectos oftalmológicos de las medicaciones sistémicas utilizadas en EAI deben ser estrechamente monitorizados, durante el curso del tratamiento.

Silica vesicles as nanocarriers and adjuvants for generating both antibody and T-cell mediated immune resposes to Bovine Viral Diarrhoea Virus E2 protein

Bovine Viral Diarrhoea Virus (BVDV) is widely distributed in cattle industries and causes significant economic losses worldwide annually. A limiting factor in the development of subunit vaccines for BVDV is the need to elicit both antibody and T-cell-mediated immunity as well as addressing the toxicity of adjuvants. In this study, we have prepared novel silica vesicles (SV) as the new generation antigen carriers and adjuvants. With small particle size of 50 nm, thin wall (similar to 6 nm), large cavity (similar to 40 nm) and large entrance size (5.9 nm for SV-100 and 16 nm for SV-140), the SV showed high loading capacity (similar to 250 mu g/mg) and controlled release of codon-optimised E2 (oE2) protein, a major immunogenic determinant of BVDV. The in vivo functionality of the system was validated in mice immunisation trials comparing oE2 plus Quil A (50 mu g of oE2 plus 10 mu g of Quil A, a conventional adjuvant) to the oE2/SV-140 (50 mu g of oE2 adsorbed to 250 mu g of SV-140) or oE2/SV-140 together with 10 mu g of Quil A. Compared to the oE2 plus Quil A, which generated BVDV specific antibody responses at a titre of 10(4), the oE2/SV-140 group induced a 10 times higher antibody response. In addition, the cell-mediated response, which is essential to recognise and eliminate the invading pathogens, was also found to be higher [1954-2628 spot forming units (SFU)/million cells] in mice immunised with oE2/SV-140 in comparison to oE2 plus Quil A (512-1369 SFU/million cells). Our study has demonstrated that SV can be used as the next-generation nanocarriers and adjuvants for enhanced veterinary vaccine delivery. (C) 2014 Elsevier Ltd. All rights reserved.

The role of anti-non-Gal antibodies in the development of acute humoral xenograft rejection of hDAF transgenic porcine kidneys in baboons receiving anti-Gal antibody neutralization therapy

Background. The present study was undertaken to determine the role of preformed and induced anti-non-Gal antibodies in the rejection of hDAF pig-to-baboon kidney xenotransplants after anti-Gal antibody neutralization therapy. Methods. Seven baboons receiv

Critical role of regulatory T cells in Th17-mediated minor antigen-disparate rejection

Th17-mediated immune responses have been recently identified as novel pathogenic mechanisms in a variety of conditions; however, their importance in allograft rejection processes is still debated. In this paper, we searched for MHC or minor Ag disparate models of skin graft rejection in which Th17 immune responses might be involved. We found that T cell-derived IL-17 is critical for spontaneous rejection of minor but not major Ag-mismatched skin grafts. IL-17 neutralization was associated with a lack of neutrophil infiltration and neutrophil depletion delayed rejection, suggesting neutrophils as an effector mechanism downstream of Th17 cells. Regulatory T cells (Tregs) appeared to be involved in Th17 reactivity. We found that in vivo Treg depletion prevented IL-17 production by recipient T cells. An adoptive cotransfer of Tregs with naive monospecific antidonor T cells in lymphopenic hosts biased the immune response toward Th17. Finally, we observed that IL-6 was central for balancing Tregs and Th17 cells as demonstrated by the prevention of Th17 differentiation, the enhanced Treg/Th17 ratio, and a net impact of rejection blockade in the absence of IL-6. In conclusion, the ability of Tregs to promote the Th17/neutrophil-mediated pathway of rejection that we have described should be considered as a potential drawback of Treg-based cell therapy.

Electroporation-mediated interleukin-10 overexpression in skeletal muscle reduces acute rejection in rat cardiac allografts

OBJECTIVES: Human interleukin 10 (hIL-10) may reduce acute rejection after organ transplantation. Our previous data shows that electroporation-mediated transfer of plasmid DNA to peripheral muscle enhances gene transduction dramatically. This study was designed to investigate the effect of electroporation-mediated overexpression of hIL-10 on acute rejection of cardiac allografts in the rat. METHODS: The study was designed to evaluate the effect of hIL-10 gene transfer on (a) early rejection pattern and (b) graft survival. Gene transfer was achieved by intramuscular (i.m.) injection into the tibialis anterior muscle of Fischer (F344) male recipients followed by electroporation 24 h prior to transplantation. Heterotopic cardiac transplantation was performed from male Brown Norway rat to F344. Four groups were studied (n = 6). Treated animals in groups B1 and B2 received 2.5 microg of pCIK hIL-10 and control animals in groups A1 and A2 distilled water. Graft function was assessed by daily palpation. Animals from group A1 were sacrificed at the cessation of the heart beat of the graft and those in group B1 were sacrificed at day 7; blood was taken for ELISA measurement of hIL-10 and tissue for myeloperoxidase (MPO) measurement and histological assessment. To evaluate graft survival, groups A2 and B2 were sacrificed at cessation of the heart beat of the graft. RESULTS: Histological examination revealed severe rejection (IIIB-IV) in group A1 in contrast to low to moderate rejection (IA-IIIA) in group B1 (p = 0.02). MPO activity was significantly lower in group B1 compared to group A1 (18 +/- 7 vs. 32 +/- 14 mU/mg protein, p = 0.05). Serum hIL-10 levels were 46 +/- 13 pg/ml in group B1 vs. 0 pg/ml in group A1. At day 7 all heart allografts in the treated groups B1 and B2 were beating, whereas they stopped beating at 5 +/- 2 days in groups A1 and A2 vs. 14 +/- 2 days in group B2 (p = 0.0012). CONCLUSIONS: Electroporation-mediated intramuscular overexpression of hIL-10 reduces acute rejection and improves survival of heterotopic heart allografts in rats. This study demonstrates that peripheral overexpression of specific genes in skeletal muscle may reduce acute rejection after whole organ transplantation.

Efficient IgG-mediated suppression of primary antibody responses in Fcγ receptor-deficient mice

IgG antibodies can suppress more than 99% of the antibody response against the antigen to which they bind. This is used clinically to prevent rhesus-negative (Rh−) women from becoming immunized against Rh+ erythrocytes from their fetuses. The suppressive mechanism is poorly understood, but it has been proposed that IgG/erythrocyte complexes bind to the inhibitory Fc receptor for IgG (FcγRIIB) on the B cell surface, thereby triggering negative signals that turn off the B cell. We show that IgG induces the same degree of suppression of the response to sheep erythrocytes in animals lacking the known IgG-binding receptors FcγRIIB, FcγRI + III, FcγRI + IIB + III, and FcRn (the neonatal Fc receptor) as in wild-type animals. Reinvestigation of the ability of F(ab′)2 fragments to suppress antibody responses demonstrated that they were nearly as efficient as intact IgG. In addition, monoclonal IgE also was shown to be suppressive. These findings suggest that IgG inhibits antibody responses through Fc-independent mechanisms, most likely by masking of antigenic epitopes, thereby preventing B cells from binding and responding to antigen. In agreement with this, we show that T cell priming is not abolished by passively administered IgG. The results have implications for the understanding of in vivo regulation of antibody responses and Rh prophylaxis.

Local production of the p40 subunit of interleukin 12 suppresses T-helper 1-mediated immune responses and prevents allogeneic myoblast rejection.

The p40 subunit of interleukin 12 (IL-12p40) has been known to act as an IL-12 antagonist in vitro. We here describe the immunosuppressive effect of IL-12p40 in vivo. A murine myoblast cell line, C2C12, was transduced with retro-virus vectors carrying the lacZ gene as a marker and the IL-12p40 gene. IL-12p40 secreted from the transfectant inhibited the IL-12-induced interferon gamma (IFN-gamma) production by splenocytes in vitro. Survival of C2C12 transplanted into allogeneic recipients was substantially prolonged when transduced with IL-12p40. Cytokine (IL-2 and IFN-gamma) production and cytotoxic T lymphocyte induction against allogeneic C2C12 were impaired in the recipients transplanted with the IL-12p40 transfectant. Delayed-type hypersensitivity response against C2C12 was also diminished in the IL-12p40 recipients. Furthermore, serum antibodies against beta-galactosidase of the T-helper 1-dependent isotypes (IgG2 and IgG3) were decreased in the IL-12p40 recipients. These results indicate that locally produced IL-12p40 exerts a potent immunosuppressive effect on T-helper 1-mediated immune responses that lead to allograft rejection. Therefore, IL-12p40 gene transduction would be useful for preventing the rejection of allografts and genetically modified own cells that are transduced with potentially antigenic molecules in gene therapy.

Synergy between T-cell immunity and inhibition of paracrine stimulation causes tumor rejection.

During tumor progression, variants may arise that grow more vigorously. The fate of such variants depends upon the balance between aggressiveness of the variant and the strength of the host immunity. Although enhancing host immunity to cancer is a logical objective, eliminating host factors necessary for aggressive growth of the variant should also be considered. The present study illustrates this concept in the model of a spontaneously occurring, progressively growing variant of an ultraviolet light-induced tumor. The variant produces chemotactic factors that attract host leukocytes and is stimulated in vitro by defined growth factors that can be produced or induced by leukocytes. This study also shows that CD8+ T-cell immunity reduces the rate of tumor growth; however, the variant continues to grow and kills the host. Treatment with a monoclonal anti-granulocyte antibody that counteracts the infiltration of the tumor cell inoculum by non-T-cell leukocytes did not interfere with the CD8+ T-cell-mediated immune response but resulted in rejection of the tumor challenge, indicating a synergy between CD8+ T-cell-mediated immunity and the inhibition of paracrine stimulation.

The vesicle size of DDA:TDB liposomal adjuvants plays a role in the cell-mediated immune response but has no significant effect on antibody production

The use of cationic liposomes as experimental adjuvants for subunit peptide of protein vaccines is well documented. Recently the cationic liposome CAF01, composed of dimethyldioctadecylammonium (DDA) and trehalose dibehenate (TDB), has entered Phase I clinical trials for use in a tuberculosis (TB) vaccine. CAF01 liposomes are a heterogeneous population with a mean vesicle size of 500 nm; a strong retention of antigen at the injection site and a Th1-biassed immune response are noted. The purpose of this study was to investigate whether CAF01 liposomes of significantly different vesicle sizes exhibited altered pharmacokinetics in vivo and cellular uptake with activation in vitro. Furthermore, the immune response against the TB antigen Ag85B-ESAT-6 was followed when various sized CAF01 liposomes were used as vaccine adjuvants. The results showed no differences in vaccine (liposome or antigen) draining from the injection site, however, significant differences in the movement of liposomes to the popliteal lymph node were noted. Liposome uptake by THP-1 vitamin D3 stimulated macrophage-like cells did not show a liposome size-dependent pattern of uptake. Finally, whilst there were no significant differences in the IgG1/2 regardless of the liposome size used as a delivery vehicle for Ag85B-ESAT-6, vesicle size has a size dependent effect on cell proliferation and IL-10 production with larger liposomes (in excess of 2 µm) promoting the highest proliferation and lowest IL-10 responses, yet vesicles of ~500 nm promoting higher IFN-? cytokine production from splenocytes and higher IL-1ß at the site of injection.

Response efficacy : the key to minimizing rejection and maximizing acceptance of emotion-based anti-speeding messages

This study sought to improve understanding of the persuasive process of emotion-based appeals not only in relation to negative, fear-based appeals but also for appeals based upon positive emotions. In particular, the study investigated whether response efficacy, as a cognitive construct, mediated outcome measures of message effectiveness in terms of both acceptance and rejection of negative and positive emotion-based messages. Licensed drivers (N = 406) participated via the completion of an on-line survey. Within the survey, participants received either a negative (fear-based) appeal or one of the two possible positive appeals (pride or humor-based). Overall, the study's findings confirmed the importance of emotional and cognitive components of persuasive health messages and identified response efficacy as a key cognitive construct influencing the effectiveness of not only fear-based messages but also positive emotion-based messages. Interestingly, however, the results suggested that response efficacy's influence on message effectiveness may differ for positive and negative emotion-based appeals such that significant indirect (and mediational) effects were found with both acceptance and rejection of the positive appeals yet only with rejection of the fear-based appeal. As such, the study's findings provide an important extension to extant literature and may inform future advertising message design.

The cell surface glycoprotein CUB domain-containing protein 1 (CDCP1) contributes to epidermal growth factor receptor-mediated cell migration

Epidermal growth factor (EGF) activation of the EGF receptor (EGFR) is an important mediator of cell migration, and aberrant signaling via this system promotes a number of malignancies including ovarian cancer. We have identified the cell surface glycoprotein CDCP1 as a key regulator of EGF/EGFR-induced cell migration. We show that signaling via EGF/EGFR induces migration of ovarian cancer Caov3 and OVCA420 cells with concomitant up-regulation of CDCP1 mRNA and protein. Consistent with a role in cell migration CDCP1 relocates from cell-cell junctions to punctate structures on filopodia after activation of EGFR. Significantly, disruption of CDCP1 either by silencing or the use of a function blocking antibody efficiently reduces EGF/EGFR-induced cell migration of Caov3 and OVCA420 cells. We also show that up-regulation of CDCP1 is inhibited by pharmacological agents blocking ERK but not Src signaling, indicating that the RAS/RAF/MEK/ERK pathway is required downstream of EGF/EGFR to induce increased expression of CDCP1. Our immunohistochemical analysis of benign, primary, and metastatic serous epithelial ovarian tumors demonstrates that CDCP1 is expressed during progression of this cancer. These data highlight a novel role for CDCP1 in EGF/EGFR-induced cell migration and indicate that targeting of CDCP1 may be a rational approach to inhibit progression of cancers driven by EGFR signaling including those resistant to anti-EGFR drugs because of activating mutations in the RAS/RAF/MEK/ERK pathway.

Osteogenic differentiation of murine embryonic stem cells is mediated by fibroblast growth factor receptors

The mechanisms involved in the control of embryonic stem (ES) cell differentiation are yet to be fully elucidated. However, it has become clear that the family of fibroblast growth factors (FGFs) are centrally involved. In this study we examined the role of the FGF receptors (FGFRs 1-4) during osteogenesis in murine ES cells. Single cells were obtained after the formation of embryoid bodies, cultured on gelatin-coated plates, and coaxed to differentiate along the osteogenic lineage. Upregulation of genes was analyzed at both the transcript and protein levels using gene array, relative-quantitative PCR (RQ-PCR), and Western blotting. Deposition of a mineralized matrix was evaluated with Alizarin Red staining. An FGFR1-specific antibody was generated and used to block FGFR1 activity in mES cells during osteogenic differentiation. Upon induction of osteogenic differentiation in mES cells, all four FGFRs were clearly upregulated at both the transcript and protein levels with a number of genes known to be involved in osteogenic differentiation including bone morphogenetic proteins (BMPs), collagen I, and Runx2. Cells were also capable of depositing a mineralized matrix, confirming the commitment of these cells to the osteogenic lineage. When FGFR1 activity was blocked, a reduction in cell proliferation and a coincident upregulation of Runx2 with enhanced mineralization of cultures was observed. These results indicate that FGFRs play critical roles in cell recruitment and differentiation during the process of osteogenesis in mES cells. In particular, the data indicate that FGFR1 plays a pivotal role in osteoblast lineage determination.

Developing a therapeutic anti-dendritic cell antibody to prevent graft versus host disease

Host and donor dendritic cells (DC) stimulate alloreactive donor T lymphocytes, and initiate GVHD. We have shown that polyclonal antibody to the DC surface activation marker human CD83 (anti hCD83), which depletes activated DC, can prevent human DC and T cell induced lethal xenogeneic GVHD in SCID mice without impairing T cell mediated anti-leukaemic and anti-viral (CMV and influenza) immunity (J Exp Med 2009; 206: 387). Therefore, we made and tested a polyclonal anti mouse CD83 (RAM83) antibody in murine HSCT models and developed a human mAb against hCD83 as a potential new therapeutic immunosuppressive agent.