Role of HLA matching and donor specific antibody development in long-term survival, acute rejection and cardiac allograft vasculopathy.

Although there are different data supporting benefits of HLA matching in kidney transplantation, its role in heart transplantation is still unclear. HLA mismatch (MM) between donor and recipient can lead to the development of donor-specific antibodies (DSA) which produces negative events on the outcome of heart transplantation. Moreover, DSAs are involved in the development of antibody-mediated rejection (AMR) and are associated with an increase in cardiac allograft vasculopathy (CAV). In this study it is analyzed retrospectively the influence of HLA matching and anti-HLA antibodies on overall survival, AMR and CAV in heart transplantation. For this retrospective study are recruited heart transplanted patients at the Cardiac Transplantation Centre of Naples between 2000 and 2019. Among the 155 heart transplant patients, the mean number of HLA-A, B, -DR MM (0 to 6) between donor and recipient was 4.5 ± 1.1. The results show a negative association between MM HLA-DR and survival (p = 0.01). Comparison of patients with 0-1 MM at each locus to all others with 2 MM, for both HLA class I and class II, has not showed significant differences in the development of CAV. Our analysis detected DSA in 38.1% of patients. The production of de novo DSA reveals that there is not an influence on survival (p = 0.72) and/or AMR (p = 0.39). Instead, there is an association between the production of DSA class II and the probability of CAV development (p = 0.03). Mean fluorescence intensity (MFI) values were significantly higher in CAV-positive patients that CAV-negative patients (p = 0.02). Prospective studies are needed to evaluate HLA class II matching as an additional parameter for heart allocation, especially considering the increment of waiting list time.

Epigenetic-based therapy in allogenic hematopoietic stem cell transplantation: Novel opportunities for personalized treatment.

Current management of patients undergoing allogeneic hematopoietic stem cell transplantation (allo-HSCT) lacks immunosuppressant drugs able to block the host immune response toward the graft antigens. Novel treatments may include epigenetic compounds (epidrugs) some of which have been yet approved by the Food and Drugs Administration for the treatment of specific blood malignancies. The most investigated in clinical trials for allo-HSCT are DNA demethylating agents (DNMTi), such as azacitidine (Vidaza) and decitabine (Dacogen) as well as histone deacetylases inhibitors (HDACi), such as vorinostat (Zolinza) and panobinostat (Farydak). Indeed, azacitidine monotherapy before allo-HSCT may reduce the conventional chemotherapy-related complications, whereas it may reduce relapse risk and death after allo-HSCT. Besides, a decitabine-containing conditioning regimen could protect against graft versus host disease (GVHD) and respiratory infections after allo-HSCT. Regarding HDACi, the addition of vorinostat and panobinostat to the conditioning regimen after allo-HSCT seems to reduce the incidence of acute GVHD. Furthermore, panobinostat alone or in combination with low-dose decitabine may reduce the relapse rate in high-risk patients with acute myeloid leukemia patients after allo-HSCT. We discuss the phase 1 and 2 clinical trials evaluating the possible beneficial effects of repurposing specific epidrugs which may guide personalized therapy in the setting of allo-HSCT.

The epigenetic promise to improve prognosis of heart failure and heart transplantation.

Heart transplantation is still the only possible life-saving treatment for end-stage heart failure, the critical epilogue of several cardiac diseases. Epigenetic mechanisms are being intensively investigated because they could contribute to establishing innovative diagnostic and predictive biomarkers, as well as ground-breaking therapies both for heart failure and heart transplantation rejection. DNA methylation and histone modifications can modulate the innate and adaptive immune response by acting on the expression of immune-related genes that, in turn, are crucial determinants of transplantation outcome. Epigenetic drugs acting on methylation and histone-modification pathways may modulate Treg activity by acting as immunosuppressive agents. Moreover, the identification of non-invasive and reliable epigenetic biomarkers for the prediction of allograft rejection and for monitoring immunosuppressive therapies represents an attractive perspective in the management of transplanted patients. MiRNAs seem to fit particularly well to this purpose because they are differently expressed in patients at high and low risk of rejection and are detectable in biological fluids besides biopsies. Although increasing evidence supports the involvement of epigenetic tags in heart failure and transplantation, further short and long-term clinical studies are needed to translate the possible available findings into clinical setting.

Seroprevalence of Bartonella henselae in patients awaiting heart transplant in Southern Italy.

BACKGROUND: Bartonella henselae is the etiologic agent of cat-scratch disease. B. henselae infections are responsible for a widening spectrum of human diseases, although often symptomless, ranging from self-limited to life-threatening and show different courses and organ involvement due to the balance between host and pathogen. The role of the host immune response to B. henselae is critical in preventing progression to systemic disease. Indeed in immunocompromised patients, such as solid organ transplant patients, B. henselae results in severe disseminated disease and pathologic vasoproliferation. The purpose of this study was to determine the seroprevalence of B. henselae in patients awaiting heart transplant compared to healthy individuals enrolled in the Regional Reference Laboratory of Transplant Immunology of Second University of Naples. METHODS: Serum samples of 38 patients awaiting heart transplant in comparison to 50 healthy donors were examined using immunfluorescence assay. RESULTS: We found a B. henselae significant antibody positivity rate of 21% in patients awaiting heart transplant (p = 0.002). There was a positive rate of 8% (p > 0.05) for immunoglobulin (Ig)M and a significant value of 13% (p = 0.02) for IgG, whereas controls were negative both for IgM and IgG antibodies against B. henselae. The differences in comorbidity between cases and controls were statistically different (1.41 ± 0.96 vs 0.42 ± 0.32; p = 0.001). CONCLUSIONS: Although this study was conducted in a small number of patients, we suggest that the identification of these bacteria should be included as a routine screening analysis in pretransplant patients.

From HLA typing to anti-HLA antibody detection and beyond: The road ahead.

The complex polymorphism of the HLA genes and the need of a proper identification of anti-HLA antibodies have led to continuously develop novel practical and feasible technologies in the field of organ and tissue transplantation. Technologies to identify HLA molecules have evolved from the serological to the molecular methods and a true innovation in the DNA sequencing has taken place with the development of next generation sequencing. An interesting field to explore is how the information resulting from the HLA-DNA sequencing can be applied in the clinical setting by including the alloimmunization assessment. Indeed, a good characterization of anti-HLA antibody at epitope level can reduce the risk of immunization. Many anti-HLA antibodies are specific for epitopes rather than for HLA antigens and the knowledge of unacceptable epitopes allows to reduce the number of mismatched antigens. Furthermore, high resolution HLA allele typing could help to understand the epitopes against which antibodies are developed. However, the improvements should not only concern the diagnostic tools in the pre-transplantation phase, but also the monitoring of post transplantation outcome. There is a growing interest in developing new non-invasive biomarkers to monitor the rejection. Currently, increasing evidence has focused on miRNAs, epigenetic markers emerged as regulators of molecular events that are differently expressed in biopsies and blood as well as in urinary samples of transplanted recipients. The implementation of next generation sequencing and genome-wide expression analysis together with functional assays may provide useful tools to evaluate the epigenetic modulation in transplantation biology but many efforts are requested for translating in the clinical arena the results obtained in experimental models.

Severe Type 2 Diabetes Induces Reversible Modifications of Endothelial Progenitor Cells Which are Ameliorate by Glycemic Control.

BACKGROUND: Circulating endothelial progenitors cells (EPCs) play a critical role in neovascularization and endothelial repair. There is a growing evidence that hyperglycemia related to Diabetes Mellitus (DM) decreases EPC number and function so promoting vascular complications. AIM OF THE STUDY: This study investigated whether an intensive glycemic control regimen in Type 2 DM can increase the number of EPCs and restores their function. METHODS: Sixty-two patients with Type 2 DM were studied. Patients were tested at baseline and after 3 months of an intensive regimen of glycemic control. The Type 2 DM group was compared to control group of subjects without diabetes. Patients with Type 2 DM (mean age 58.2±5.4 years, 25.6% women, disease duration of 15.4±6.3 years) had a baseline HgA1c of 8.7±0.5% and lower EPC levels (CD34+/KDR+) in comparison to healthy controls (p<0.01). RESULTS: The intensive glycemic control regimen (HgA1c decreased to 6.2±0.3%) was coupled with a significant increase of EPC levels (mean of 18%, p<0.04 vs. baseline) and number of EPCs CFUs (p<0.05 vs. baseline). CONCLUSION: This study confirms that number and bioactivity of EPCs are reduced in patients with Type 2 DM and, most importantly, that the intensive glycemic control in Type 2 DM promotes EPC improvement both in their number and in bioactivity.

Comprehensive assessment of sensitizing events and anti-HLA antibody development in women awaiting kidney transplantation.

BACKGROUND: Alloimmunization remains a critical factor which affects the success of kidney transplantation. Patients awaiting solid organ transplantation may develop anti-HLA antibodies after pregnancies, transfusions and previous events of transplantations. AIM: We evaluated the effects of different sensitizing events on the anti-HLA antibody production and the potential role of patient HLA alleles in the context of antibody development in both the overall and pregnancy sensitized groups. MATERIAL AND METHODS: We retrospectively stratified 411 women on waiting list for kidney transplantation by route of sensitization. The presence of anti-HLA antibodies was evaluated by Solid Phase Assay and HLA typing was performed by serological and molecular methods. RESULTS: In our study population, 54% of women had anti-HLA antibodies. We found that the 51.6% of women with pregnancy only, 44% of women with transfusion only and 100% of women with a history of transplantation only developed anti-HLA antibodies. Pregnancy only resulted significantly associated with all anti-HLA antibody development such as anti-A, -B, -C, -DR, -DP as well as anti-DQB and -DQA antibodies. We investigated the influence of patient HLA alleles on the antibody development in the overall study population. Patients expressing HLA A*32 (p=0.024, OR=0.42), B*14 (p=0.035, OR=0.44), HLA-B*44 (p=0.026, OR=0.51) and DRB1*01 (p=0.029, OR=0.55) alleles produced anti-HLA antibodies less frequently compared to subjects with other alleles. In the pregnancy only group, B*14 (p=0.010, OR=0.12) and B*51 (p=0.005, OR=0.24) alleles were associated with a low risk of anti-HLA antibody development, while A*11 (p=0.033, OR=3.56) and DRB1*04 (p=0.022, OR=3.03) alleles seem to represent a higher risk. CONCLUSIONS: Pregnancy still remains a strong sensitizing event in women awaiting kidney transplantation. The anti-HLA antibody development in pregnancy appears to be associated with the expression of particular HLA alleles.

Innate and adaptive immune response in stroke: Focus on epigenetic regulation.

Inflammation and immune response play a pivotal role in the pathophysiology of ischemic stroke giving their contribution to tissue damage and repair. Emerging evidence supports the involvement of epigenetic mechanisms such as methylation, histone modification and miRNAs in the pathogenesis of stroke. Interestingly, epigenetics can influence the molecular events involved in ischemic injury by controlling the switch from pro- to anti-inflammatory response, however, this is still a field to be fully explored. The knowledge of epigenetic processes could to allow for the discovery of more sensitive and specific biomarkers for risk, onset, and progression of disease as well as further novel tools to be used in both primary prevention and therapy of stroke. Indeed, studies performed in vitro and in small animal models seem to suggest a neuroprotective role of HDAC inhibitors (e.g. valproic acid) and antagomir (e.g. anti-miR-181a) in ischemic condition by modulation of both immune and inflammatory pathways. Thus, the clinical implications of altered epigenetic mechanisms for the prevention of stroke are very promising but clinical prospective studies and translational approaches are still warranted.

Infections and cardiovascular disease: is Bartonella henselae contributing to this matter?

Cardiovascular disease is still the major cause of death worldwide despite the remarkable progress in its prevention and treatment. Endothelial progenitor cells (EPCs) have recently emerged as key players of vascular repair and regenerative medicine applied to cardiovascular disease. A large amount of effort has been put into discovering the factors that could aid or impair the number and function of EPCs, and also into characterizing these cells at the molecular level in order to facilitate their therapeutic applications in vascular disease. Interestingly, the major cardiovascular risk factors have been associated with reduced number and function of EPCs. The bacterial contribution to cardiovascular disease represents a long-standing controversy. The discovery that Bartonella henselae can infect and damage EPCs revitalizes the enduring debate about the microbiological contribution to atherosclerosis, thus allowing the hypothesis that this infection could impair the cardiovascular regenerative potential and increase the risk for cardiovascular disease. In this review, we summarize the rationale suggesting that Bartonella henselae could favour atherogenesis by infecting and damaging EPCs, thus reducing their vascular repair potential. These mechanisms suggest a novel link between communicable and non-communicable human diseases, and put forward the possibility that Bartonella henselae could enhance the susceptibility and worsen the prognosis in cardiovascular disease.

Epigenetic-related therapeutic challenges in cardiovascular disease.

Progress in human genetic and genomic research has led to the identification of genetic variants associated with specific cardiovascular diseases (CVDs), but the pathogenic mechanisms remain unclear. Recent studies have analyzed the involvement of epigenetic mechanisms such as DNA methylation and histone modifications in the development and progression of CVD. Preliminary work has investigated the correlations between DNA methylation, histone modifications, and RNA-based mechanisms with CVDs including atherosclerosis, heart failure (HF), myocardial infarction (MI), and cardiac hypertrophy. Remarkably, both in utero programming and postnatal hypercholesterolemia may affect the epigenetic signature in the human cardiovascular system, thereby providing novel early epigenetic-related pharmacological insights. Interestingly, some dietary compounds, including polyphenols, cocoa, and folic acid, can modulate DNA methylation status, whereas statins may promote epigenetic-based control in CVD prevention through histone modifications. We review recent findings on the epigenetic control of cardiovascular system and new challenges for therapeutic strategies in CVDs.

Epigenetic control of autoimmune diseases: from bench to bedside.

Genome-wide association studies have revealed several genes predisposing to autoimmunity, however, concordance rates in monozygotic twins are significantly below 50% for several autoimmune diseases. The limited presence of a strong genetic association only in some patients supports that other non-genetic mechanisms are active in these pathologies. Epigenetic modifications such as DNA methylation, histone modification, and microRNA signaling regulate gene expression and are sensitive to external stimuli and they might be as bridging between genetic and environmental factors. Some evidence has highlighted the involvement of epigenetic alterations in the pathogenesis of various autoimmune diseases giving rise to great expectations among clinicians and researchers. The direct role of these alterations in the initiation/progression of autoimmune diseases is still unclear. The knowledge in depth of these pathogenic and epigenetic mechanisms will increase the possibility of the control and/or prevention of autoimmune diseases through the use of drugs that target epigenetic pathways. Moreover, we could use epigenetic-related biomarkers to follow this complicated framework (for example H3K4me3 and miRNA-155 are among those proposed biomarkers). This article reviews current understanding of the epigenetic involvement in the field of autoimmune diseases especially in systemic lupus erythematosus, rheumatoid arthritis, sclerosis multiple and type 1 diabetes.

Epigenetic reprogramming in atherosclerosis.

Recent data support the involvement of epigenetic alterations in the pathogenesis of atherosclerosis. The most widely investigated epigenetic mechanism is DNA methylation although also histone code changes occur during the diverse steps of atherosclerosis, such as endothelial cell proliferation, vascular smooth muscle cell (SMC) differentiation, and inflammatory pathway activation. In this review, we focus on the main genes that are epigenetically modified during the atherogenic process, particularly nitric oxide synthase (NOS), estrogen receptors (ERs), collagen type XV alpha 1 (COL15A1), vascular endothelial growth factor receptor (VEGFR), and ten-eleven translocation (TET), which are involved in endothelial dysfunction; gamma interferon (IFN-γ), forkhead box p3 (FOXP3), and tumor necrosis factor-α (TNF-α), associated with atherosclerotic inflammatory process; and p66shc, lectin-like oxLDL receptor (LOX1), and apolipoprotein E (APOE) genes, which are regulated by high cholesterol and homocysteine (Hcy) levels. Furthermore, we also discuss the role of non-coding RNAs (ncRNA) in atherosclerosis. NcRNAs are involved in epigenetic regulation of endothelial function, SMC proliferation, cholesterol synthesis, lipid metabolism, and inflammatory response.

Human leukocyte antigens and alloimmunization in heart transplantation: an open debate.

Considerable advances in heart transplantation outcome have been achieved through the improvement of donor-recipient selection, better organ preservation, lower rates of perioperative mortality and the use of innovative immunosuppressive protocols. Nevertheless, long-term survival is still influenced by late complications. We support the introduction of HLA matching as an additional criterion in the heart allocation. Indeed, allosensitization is an important factor affecting heart transplantation and the presence of anti-HLA antibodies causes an increased risk of antibody-mediated rejection and graft failure. On the other hand, the rate of heart-immunized patients awaiting transplantation is steadily increasing due to the limited availability of organs and an increased use of ventricular assist devices. Significant benefits may result from virtual crossmatch approach that prevents transplantation in the presence of unacceptable donor antigens. A combination of both virtual crossmatch and a tailored desensitization therapy could be a good compromise for a favorable outcome in highly sensitized patients. Here, we discuss the unresolved issue on the clinical immunology of heart transplantation.

Lights and shadows of anti-HLA antibodies detected by solid-phase assay.

Recently, management of patients awaiting solid organ transplantation has taken advantages after the development of more sensitive and accurate solid phase assays which have supported the historic complement dependent cytotoxicity. This approach has allowed the detection of antibodies in patients previously considered negative. The use of the single antigen beads resulted in a more accurate anti-human leukocyte antigen (HLA) antibody characterization. The detection of anti-HLA antibodies specific for C, DQ and DP loci that were not so well characterized has been possible through the implementation of the single antigen assay. The assessment of HLA compatibility has been expanded through the introduction of "epitope matching" concept and the definition of the unacceptable antigens for a more adequate evaluation of donor-recipient compatibility. However, the clinical impact of pre-formed and de novo anti-HLA antibodies detected by solid phase assays is still controversial due to the drawback related to result interpretation. Until today, the unresolved issues concern if all antibodies affect the medium and long term clinical outcome. An open debate on the clinical relevance of anti-HLA antibodies detected by single-antigen beads highlights needing to further investigations. Here, we describe the novel applications and the improvements of the solid-phase assay use.

Intravenous human immunoglobulin treatment of serum from HLA-sensitized patients in kidney transplantation.

OBJECTIVE: Intravenous immunoglobulin (IVIG) products are known to have beneficial immunomodulatory effects on several inflammatory and autoimmune disorders. These effects could be attributed to a different inhibitory action on complement factors, but other mechanisms could be implicated, e.g., immunocomplexes development and/or anti-idiotypic antibodies. Positive results on the reduction of anti-Human Leukocyte Antigens (HLA) antibodies in highly sensitized patients have also been found. The present study focuses on the effect of IVIG on the reduction of Panel Reactive Antibody level and crossmatch positivity in sensitized patients awaiting kidney transplantation. METHODS: The study was performed adapting an in vitro assay on sensitized patients' sera in waiting list for kidney transplantation. Sera of twelve highly sensitized patients were evaluated for the cytotoxicity inhibition after 10% IVIG treatment. RESULTS: A reduction of anti- HLA antibody levels was observed in 75% (9/12) of treated patients in vitro, while 25% (3/12) resulted unresponsiveness. Particularly, our data showed a significantly higher Panel Reactive Antibody reduction for T lymphocytes (p<0.010) than B lymphocytes (p<0.032). CONCLUSIONS: In this study, we have used an in vitro assay to investigate susceptibility to desensitization with IVIG treatment of sensitized patient sera. These findings reveal that the variable effect of IVIG on reducing Panel Reactive Antibody in our immunized patients could be attributed to a different inhibitory action on complement, likely due to the type and the titre of anti-HLA antibodies.

Long-term follow-up of kidney transplants in a region of Southern Italy.

OBJECTIVES: Several donor and recipient factors are known to be associated with graft loss in a kidney transplant. In this retrospective single-center study, we analyzed the effect of clinical and immunologic factors on kidney transplant outcomes in our region in Italy. MATERIALS AND METHODS: The study included 245 transplanted recipients from deceased donors at Federico II University of Naples, Kidney Transplant Centre, between the years 2000 and 2006. Age, cause of death, history of hypertension, hypotension or cardiac arrest, length of time spent in the intensive care unit, serum creatinine levels and human leukocyte antigen typing all were evaluated in the donors. Age, time spent on the wait list, human leukocyte antigen typing, antibody sensitization, and allocation were evaluated in the recipients. Age, donor/recipient matching, and human leukocyte antigen mismatches also were evaluated. RESULTS: Cox regression analysis showed that in recipients, time spent on the wait list increased the risk of restarting dialysis (OR 1.019, 95% CI: 1.000-1.038; P = .050) and dying (OR 1.017, 95% CI: 1.000-1.038; P = .032). Patients who received a kidney from a donor with a history of hypertension presented a major risk of death (OR 3.212, 95% CI: 1.190-8.668; P = .021), while human leukocyte antigen-A mismatch increased the risk of restarting dialysis (OR 3.137, 95% CI: 1.255-7.842; P = .014). CONCLUSIONS: In our study, in recipients, time spent on the wait list, and a history of hypertension were associated with a greater risk of death. Human leukocyte antigen-A mismatch is associated with a greater risk of restarting dialysis.

Association between human leukocyte antigen class I and II alleles and hepatitis C virus infection in high-risk hemodialysis patients awaiting kidney transplantation.

Recent evidences have shown that several host genetic factors influence susceptibility or protection to hepatitis C virus (HCV) infection. There are controversial data regarding the associations of human leukocyte antigens (HLA) and the clearance or progression of HCV. The aim of this study was to investigate whether particular HLA molecules were associated with HCV infection in recipients awaiting kidney transplantation considered at high-risk to infection due to protracted hemodialysis treatment. To this purpose, 301 kidney recipients with HCV infection and 1103 uninfected recipients were examined for HLA class I and II molecules. In our case-control study, HLA-A(*)26 is positively associated with HCV infection while HLA-A(*)29, -B(*)40 and -DRB1(*)01 are negatively associated with HCV infection. Multiple logistic regression analysis demonstrated that age (OR = 1.02; 95% CI = 1.01-1.04; p < 0.00), HLA-A(*)26, -A(*)29, -B(*)40 and -DRB1(*)01 [(OR = 1.54; 95% CI = 1.03-2.30; p = 0.03); (OR = 0.50; 95% CI = 0.26-0.99; p = 0.05); (OR = 0.42; 95% CI = 0.23-0, 7; p = 0.01); (OR = 0.62; 95% CI = 0.41-0, 94; p = 0.03); respectively] are independent predictors of HCV infection. Our results suggest that particular HLA molecules, as host genetic factors, may have a relationship with susceptibility or protection to HCV infection also in recipients awaiting kidney transplantation.

Human leukocyte antigen-DR mismatch is associated with increased in-hospital mortality after a heart transplant.

OBJECTIVES: Although previous studies have investigated the effect of human leukocyte antigen matching on long-term outcomes after heart transplants, its role in the prognosis after a heart transplant remains unclear, particularly with respect to short-term survival. MATERIALS AND METHODS: We evaluated the human leukocyte antigen mismatch on in-hospital mortality of 158 consecutive patients who had undergone a heart transplant between 2000 and 2008. Human leukocyte antigens-A, -B, and -DR were determined by means of serologic and molecular techniques. Univariate analysis and a multiple logistic regression models evaluated the effect of human leukocyte antigen variants on mortality, independent of clinical variables. RESULTS: In-hospital mortality was 11.4%. Higher prevalence of acute kidney injury (50.0% vs 12.9%), higher levels of troponins 48 hours after transplant (15.6 ± 12.0 ng/mL vs 9.7 ± 9.4 ng/mL), prolonged ischemia (188.2 ± 32.5 min vs 162.6 ± 40.7 min), higher frequency of reoperation (61.1% vs 17.9%), and higher human leukocyte antigen-DR mismatch (1.61 ± 0.5 vs 1.30 ± 0.6) were found in patients who died. By logistic regression analysis, humanleukocyte antigen-DR mismatch is associated with in-hospital mortality (OR=5.159, 95% CI=1.348-19.754), independent of the effect of covariates such as recipient age, mismatch sex, mismatch human leukocyte antigen-A, human leukocyte antigen-B, acute kidney injury, reoperation, ischemia duration, and levels of troponins. CONCLUSIONS: Human leukocyte antigen-DR mismatch is associated with in-hospital mortality in heart transplant.

Heart transplant with donor-specific antibody after immunoadsorption plus rituximab: a case report.

Different desensitization strategies are available for treating patients with preformed human leukocyte antigen (HLA) antibodies. A highly presensitized heart recipient received immunoadsorption and rituximab therapy. The patient, with end-stage heart failure, was positive only for antibodies of HLA class I (anti-A2, A10, B17), and Luminex platform (One Lambda kit) showed a panel-reactive antibody score of 64%. The patient's serum was tested repeatedly in both complement-dependent cytotoxicity and flow-cytometry crossmatches against cells from different potential organ donors. The results of these crossmatches were positive on flow cytometry when tested with HLA-A2, A10, and B17 but were still negative on cytotoxicity. The patient was treated with a desensitization regimen; this treatment immediately decreased antibody levels of 70% and the patient subsequently received a transplant with donor-specific HLA antibody (HLA-A2). After more than 2 years, graft function remains normal and the clinical status of the patient is stable.

Recent advances in proteomic technologies applied to cardiovascular disease.

In recent years, the diagnosis of cardiovascular disease (CVD) has increased its potential, also thanks to mass spectrometry (MS) proteomics. Modern MS proteomics tools permit analyzing a variety of biological samples, ranging from single cells to tissues and body fluids, like plasma and urine. This approach enhances the search for informative biomarkers in biological samples from apparently healthy individuals or patients, thus allowing an earlier and more precise diagnosis and a deeper comprehension of pathogenesis, development and outcome of CVD to further reduce the enormous burden of this disease on public health. In fact, many differences in protein expression between CVD-affected and healthy subjects have been detected, but only a few of them have been useful to establish clinical biomarkers because they did not pass the verification and validation tests. For a concrete clinical support of MS proteomics to CVD, it is, therefore, necessary to: ameliorate the resolution, sensitivity, specificity, throughput, precision, and accuracy of MS platform components; standardize procedures for sample collection, preparation, and analysis; lower the costs of the analyses; reduce the time of biomarker verification and validation. At the same time, it will be fundamental, for the future perspectives of proteomics in clinical trials, to define the normal protein maps and the global patterns of normal protein levels, as well as those specific for the different expressions of CVD.