Alpha-Synuclein in Skin Nerve Fibers as a Biomarker for Alpha-Synucleinopathies.

The common pathological features of synucleinopathies are abnormal aggregates of the synaptic protein alpha-synuclein (αSN) in the cytoplasm of neurons or glia. These abnormal aggregates appear several years before the onset of clinical manifestations, and so the early detection of αSN in body fluids or peripheral tissues (e.g., cerebrospinal fluid, colonic mucosa, salivary glands, and skin) is considered a potential tool for identifying synucleinopathies. Performing a skin biopsy is a practical option because it is a relatively noninvasive, safe, and reliable method to measure αSN deposition in the peripheral nervous system. Moreover, there is growing research interest in the use of cutaneous synuclein deposition as a biomarker for synucleinopathies. The aim of this study was to interpret the current data on cutaneous αSN deposition and present the current perspectives and future prospects.

Vascular Endothelial Growth Factor Prevents Endothelial-to-Mesenchymal Transition in Hypertrophy.

BACKGROUND: In hypertrophy, progressive loss of function caused by impaired diastolic compliance correlates with advancing cardiac fibrosis. Endothelial cells contribute to this process through endothelial-to-mesenchymal transition (EndMT) resulting from inductive signals such as transforming growth factor (TGF-ß). Vascular endothelial growth factor (VEGF) has proven effective in preserving systolic function and delaying the onset of failure. In this study, we hypothesize that VEGF inhibits EndMT and prevents cardiac fibrosis, thereby preserving diastolic function. METHODS: The descending aorta was banded in newborn rabbits. At 4 and 6 weeks, hypertrophied animals were treated with intrapericardial VEGF protein and compared with controls (n = 7 per group). Weekly transthoracic echocardiography measured peak systolic stress. At 7 weeks, diastolic stiffness was determined through pressure-volume curves, fibrosis by Masson trichrome stain and hydroxyproline assay, EndMT by immunohistochemistry, and activation of TGF-ß and SMAD2/3 by quantitative real-time polymerase chain reaction. RESULTS: Peak systolic stress was preserved during the entire observation period, and diastolic compliance was maintained in treated animals (hypertrophied: 20 ± 1 vs treated: 11 ± 3 and controls: 12 ± 2; p < 0.05). Collagen was significantly higher in the hypertrophied group by Masson trichrome (hypertrophied: 3.1 ± 0.9 vs treated: 1.8 ± 0.6) and by hydroxyproline assay (hypertrophied: 2.8 ± 0.6 vs treated: 1.4 ± 0.4; p < 0.05). Fluorescent immunostaining showed active EndMT in the hypertrophied group but significantly less in treated hearts, which was directly associated with a significant increase in TGF-ß/SMAD-2 messenger RNA expression. CONCLUSIONS: EndMT contributes to cardiac fibrosis in hypertrophied hearts. VEGF treatment inhibits EndMT and prevents the deposition of collagen that leads to myocardial stiffness through TGF-ß/SMAD-dependent activation. This presents a therapeutic opportunity to prevent diastolic failure and preserve cardiac function in pressure-loaded hearts.

Five-year review of an international clinical research-training program.

The exponential increase in clinical research has profoundly changed medical sciences. Evidence that has accumulated in the past three decades from clinical trials has led to the proposal that clinical care should not be based solely on clinical expertise and patient values, and should integrate robust data from systematic research. As a consequence, clinical research has become more complex and methods have become more rigorous, and evidence is usually not easily translated into clinical practice. Therefore, the instruction of clinical research methods for scientists and clinicians must adapt to this new reality. To address this challenge, a global distance-learning clinical research-training program was developed, based on collaborative learning, the pedagogical goal of which was to develop critical thinking skills in clinical research. We describe and analyze the challenges and possible solutions of this course after 5 years of experience (2008-2012) with this program. Through evaluation by students and faculty, we identified and reviewed the following challenges of our program: 1) student engagement and motivation, 2) impact of heterogeneous audience on learning, 3) learning in large groups, 4) enhancing group learning, 5) enhancing social presence, 6) dropouts, 7) quality control, and 8) course management. We discuss these issues and potential alternatives with regard to our research and background.

Prolongation of alloskin graft survival by catalytic scavengers of reactive oxygen species.

We tested the effects of salen manganese (Salen-Mn) complexes, which are scavengers of reactive oxygen species exhibiting superoxide dismutase and catalase activities on the rejection of and alloresponse to fully allogeneic skin grafts in mice. We showed that pre-transplant treatment of C57Bl/6 donor skin or of BALB/c recipients with Salen-Mn complexes significantly delayed allograft rejection. ELISPOT analysis of alloimmune response of treated mice revealed a significant reduction of the frequency of type 1 cytokine (pro-inflammatory) producing T-cells, while the number of activated T-cells producing type 2 cytokines was elevated. In addition, anti-oxidative treatment of graft recipients resulted in a profound inhibition of their donor-specific cytotoxic T-cell response. Our results indicate that salen manganese complexes mediate their effect on graft rejection both by reducing the susceptibility of graft tissue to ROS-mediated injury and by exerting an anti-inflammatory effect in recipients.

Tolerance to noninherited maternal MHC antigens in mice.

The phenomenon of tolerance to noninherited maternal Ags (NIMA) is poorly understood. To analyze the NIMA effect C57BL/6 (H-2(b/b)) males were mated with B6D2F(1) (H-2(b/d)) females, whereby 50% of the offspring are H-2(b/b) mice that have been exposed to maternal H-2(d) alloantigens. Controls were H-2(b/b) offspring of C57BL/6 mothers, either inbred C57BL/6 mice or F(1) backcross mice from breedings with H-2(b/d) fathers. We found that 57% of the H-2(b/b) offspring of semiallogeneic (H-2(b/d)) mothers accepted fully allogeneic DBA/2 (H-2(d/d)) heart grafts for >180 days, while similar transplants were all rejected by day 11 in controls (p < 0.0004). Foster nursing studies showed that both oral and in utero exposure to NIMA are required for this tolerogenic effect. An effect of NIMA was also found to extend the survival of skin grafts from a semiallogeneic donor (p < 0.02). Pretransplant analysis of splenocytes showed a 40-90% reduction of IL-2-, IL-5-, and IFN-gamma-producing T cells responding to H-2(d)-expressing APC in NIMA(d)-exposed vs control mice. Injection of pregnant BALB/c-dm2 (H-2L(d)-negative) female mice i.v. with H-2L(d)(61-80) peptide profoundly suppressed the offspring's indirect pathway alloreactive CD4(+) T cell response to H-2L(d). These results suggest that the natural exposure of the fetus and newborn to maternal cells and/or soluble MHC Ags suppresses NIMA-allospecific T cells of the offspring, predisposing to organ transplant tolerance in adult mice.

T-cell response to cardiac myosin persists in the absence of an alloimmune response in recipients with chronic cardiac allograft rejection.

BACKGROUND: Immune-mediated injury to the graft has been implicated in the pathogenesis of chronic rejection. However, little is known regarding the nature of the antigen(s) involved in this immune process. We demonstrated that cardiac transplantation in mice induces an autoimmune T-cell response to a heart tissue-specific protein, cardiac myosin (CM). This response contributes to transplant rejection in that its modulation affects cardiac graft survival. This study investigates whether anti-CM T cells undergo activation and expansion in mice with chronic cardiac allograft rejection. METHODS: The frequency of CM- and donor major histocompatibility complex (MHC)-specific interferon (IFN)-gamma-producing T cells were assessed by ELISPOT in BALB/c mice, which were injected with anti-CD40L (MR1) mAb (chronic rejection group) or CTLA4Ig fusion protein (tolerant group) and transplanted with C57BL/6 cardiac allografts. RESULTS AND CONCLUSIONS: MR1-treated BALB/c recipients of C57BL/6 hearts with chronic rejection displayed a high frequency of activated CM-specific T cells, whereas the frequency of activated alloreactive T cells were similar to naïve, nontransplanted mice. In contrast, no activation of CM-reactive T cells was detected in tolerant recipients after CTLA4Ig treatment. Therefore, in the absence of alloimmunity, chronic rejection is associated with persistence of a T-cell response against CM. Our data indicate that anti-CM autoimmunity may be involved in the immune mechanisms of chronic rejection and suggest that tolerance strategies should target both allo- and autoimmune responses to prevent this process.

Modulation of tissue-specific immune response to cardiac myosin can prolong survival of allogeneic heart transplants.

The role of immune response to tissue-specific Ags in transplant rejection is poorly defined. We have previously reported that transplantation of cardiac allografts triggers a CD4(+) Th1 cell response to cardiac myosin (CM), a major contractile protein of the heart, and that pretransplant activation of proinflammatory CM-specific T cells accelerates rejection. In this study, we show that administration of CM together with IFA (CM/IFA) can prevent acute rejection of an allogeneic heart transplant. Prolongation of cardiac graft survival is associated with activation of CM- and allo-specific T cells secreting type 2 cytokines (IL-4, IL-5) and reduction of the frequency of proinflammatory IFN-gamma-secreting (type 1) alloreactive T cells. Blocking of IL-4 cytokine with Abs abrogates the prolongation. CM/IFA treatment prevents acute rejection of MHC class I-mismatched, but not fully mismatched grafts. However, if donor heart is devoid of MHC class II expression, CM-IFA administration delays rejection of fully allogeneic cardiac transplants. This finding suggests that the effect of CM modulation depends on the type (direct vs indirect) and strength of recipient's CD4(+) T cell alloresponse. Our results underscore the important role of host immunity to tissue-specific Ags in the rejection of an allograft. This study demonstrates that modulation of the immune response to a tissue-specific Ag can significantly prolong cardiac allograft survival, an observation that may have important implications for the development of novel selective immune therapies in transplantation.