Cellular xenotransplantation of animal cells into people: benefits and risk

The main benefit of xenotransplantation is its potential to overcome the worldwide organ shortage experienced in allotransplantation. Allogeneic transplantation is the only successful therapy for several life-threatening diseases, with cell, tissue or organ donation only partially meeting the demand and many patients dying while waiting for treatment. With supply falling short of demand, it is foreseen that the use of porcine material may at some stage overcome the existing gap between organ availability and clinical need. Recently, pig islet cells have been utilised in clinical trials, with safety being demonstrated. Indeed, pig-derived cells present several advantages: i) porcine cells have a stable function and differentiation pattern and are not tumorigenic; ii) pig cells have been shown to meet the physiological needs in large animal models; iii) the source of pig cells can be scaled up to meet demands in a highly standardised manner, and with respect to animal welfare regulations; iv) ‘designated-pathogen-free’ (DPF) pig lines can be produced, which could result in a higher safety profile than allotransplantation itself; v) the risk of zoonosis, which was raised years ago as the major hurdle, has been recently circumvented and is actually viewed as a controlled risk; and vi) immune risks are being circumvented via the use of genetically modified donor animals and encapsulation of porcine cells, particularly for the treatment of diabetes. Overall, the benefit appears to outweigh potential risks with respect to cellular xenotransplantation and this is discussed further in this review

Deceased donor-initiated Chains: first report of a successful deliberate case and its ethical implications

Background: The utilization of deceased donor kidneys to initiate chains of living donor kidney paired donation (KPD) has been proposed, although the potential gain of this practice needs to be quantified and the ethical implications must be addressed before starting its application. Methods: The gain of implementing deceased donor-initiated chains has been measured through a mathematical algorithm, using retrospective data on the pool of donor/recipient incompatible pairs at a single Center. Allocation rules of chain ending kidneys and characteristics/quality of the chain initiating kidney (CIK) are described. Results: the quantification of benefit analysis showed that with a pool of 69 kidneys from deceased donors and 16 pairs enrolled in the KPD program, over a period of 3 years it is possible to transplant 8/16 recipients (50%). Following the approval of the Bioethical Committee of the Veneto Region and the revision of the allocation policies by the Italian National Transplant Center, the first successful case has been performed. The waiting time of the recipient (male, 53 yo) after entering the program for the CIK with a kidney donor risk index (KDRI) equal to 0.61 and a kidney donor profile index (KDPI) of 3%, was 4 days. His willing donor (female, 53 yo) with a living kidney donor profile index (LKDPI) of 2, donated 2 days later to a chain ending recipient (male, 47 yo,) who had been on dialysis for 5 years. Conclusions: This is the first report of a deliberate deceased donor-initiated chain, which has been successfully performed. This has been made possible thanks to an extensive phase of evaluation of the ethical issues and allocation policy impact. This paper includes a preliminary efficacy assessment and the development a dedicated algorithm

Light-Induced Access to Carbazole-1,3-dicarbonitrile: A Thermally Activated Delayed Fluorescent (TADF) Photocatalyst for Cobalt-Mediated Allylations

The stability of a photocatalyst under irradiation is important in photoredox applications. In this work, we investigated the stability of a thermally activated delayed fluorescence (TADF) photocatalyst {3DPAFIPN [2,4,6-tris(diphenylamino)-5-fluoroisophthalonitrile]}, recently employed in photoredox-mediated processes, discovering that in the absence of quenchers the chromophore is unstable and is efficiently converted by irradiation with visible light into another species based on the carbazole-1,3-dicarbonitrile moiety. The new species obtained is itself a TADF emitter and finds useful applications in photoredox transformations. At the excited state, it is a strong reductant and was efficiently applied to cobalt-mediated allylation of aldehydes, whereas other TADFs (4CzIPN and 3DPAFIPN) failed to promote efficient photocatalytic cycles

Extended criteria donor lung reconditioning with the organ care system lung: a single institution experience

Lung transplantation is a life-saving procedure limited by donor's availability. Lung reconditioning by ex vivo lung perfusion represents a tool to expand the donor pool. In this study, we describe our experience with the OCS\u2122 Lung to assess and recondition extended criteria lungs. From January 2014 to October 2016, of 86 on-site donors evaluated, eight lungs have been identified as potentially treatable with OCS\u2122 Lung. We analyzed data from these donors and the recipient outcomes after transplantation. All donor lungs improved during OCS perfusion in particular regarding the PaO2/FiO2 ratio (from 340 mmHg in donor to 537 mmHg in OCS) leading to lung transplantation in all cases. Concerning postoperative results, primary graft dysfunction score 3 at 72 h was observed in one patient, while median mechanical ventilation time, ICU, and hospital stay were 60 h, 14 and 36 days respectively. One in-hospital death was recorded (12.5%), while other two patients died during follow-up leading to 1-year survival of 62.5%. The remaining five patients are alive and in good conditions. This case series demonstrates the feasibility and value of lung reconditioning with the OCS\u2122 Lung; a prospective trial is underway to validate its role to safely increase the number of donor lungs. \ua9 2018 Steunstichting ESO

Prevalence of Chagas disease and strongyloidiasis among HIV-infected Latin American immigrants in Italy – The CHILI study

INTRODUCTION: Screening HIV-positive migrants for neglected tropical diseases having potential for life-threatening reactivation, such as Chagas disease and strongyloidiasis is not widely implemented. We evaluated the prevalence of these infections among a large cohort of HIV-infected migrants from Latin America living in Italy. METHOD: Cross-sectional study evaluating the prevalence of Trypanosoma cruzi and Strongyloides stercoralis infections in HIV-infected migrants from Latin America enrolled in the Italian Cohort of Antiretroviral-Naïve patients (ICONA) between 1997 and 2018, based on serology performed on sera stored in the ICONA Foundation biobank. Screening for Chagas disease was performed using two commercial ELISA complemented by commercial Immunoblot and CLIA if discordant. Strongyloidiasis was evaluated using a commercial ELISA. RESULTS: 389 patients were analysed. Fifteen (3.86%) had at least one positive Chagas ELISA test. Prevalence of Chagas disease was 0.5% or 1.29% depending on the confirmatory technique. Serology for strongyloidiasis was positive in 16 (4.11%) patients. Only Nadir CD4+ T cell count was associated with discordant serology for Chagas disease (p = 0.046). CONCLUSIONS: The accuracy of seroassays for Chagas disease and strongyloidiasis in HIV-positive patients is unclear. To avoid missing potentially life-threatening infections, we suggest implementing additional diagnostic strategies in at-risk patients with inconclusive serology results

Biosafety in surgical pathology in the era of SARS-Cov2 pandemia. A statement of the Italian Society of Surgical Pathology and Cytology

Surgical pathology units face chemical and biological risks. While chemical risks have been intensely evaluated since the formalin ban, less attention has been drown to biological risks. The actual epidemiologic situation due to the SARS-CoV-2 pandemia has raised a series of questions, which need to be addressed as soon as possible. We have to pursue two lines of action: on one hand we must immediately adopt urgent measures to reduce the risk of SARS-CoV-2 infection of laboratory personnel, and on the other hand, we must address crucial technical and organizational aspects of biological risk reduction, preserving as much as possible the quality of tissue and cell samples. The evaluation of biological risk is an analytical process which involves different steps: a) characterization of the hazard (also known as risk assessment) and b) definition of a risk reduction strategy (also known as risk mitigation) 1. Risk assessment implies a) the identification of the intrinsic biologic characteristics of the infectious agent, and b) the identification of the laboratory procedures related to the agent. The intrinsic biologic characteristics of infectious agents are classified in 4 risk groups (RG) by the laboratory biosafety manual of the WHO 2. The RG range from level 1 (RG1) which includes microorganisms that are unlikely to cause human or animal disease, to level 4 (RG4) which includes pathogens which cause serious diseases, that can be readily transmitted from one individual to another, and for which effective treatment and preventive measures are not usually available. Risk mitigation includes the definition of the appropriate a) level of biosafety of the laboratory, b) type of personal protection equipment (PPE), c) type of infrastructure and equipment, and d) education of involved personnel. Laboratory biosafety is graded in 4 levels (BSL-1 to BSL-4) as exhaustively described in the laboratory biosafety manual of the WHO 2, and these levels are usually also defined by law (in Italy by the D. Lgs. 81/2008). BSL are a series of protections, which include individual safeguards designed to protect laboratory personnel, as well as the surrounding environment and community. The biosafety level required in laboratories derives from the characterization of the risk, and is not automatically derived from the risk group to which the pathogenic agent belongs. It is obvious that the biosafety level for a laboratory which cultivates a RG3 agent, will be higher than the level needed for a laboratory which performs diagnostic tests on inactivated biomaterials on the same agent. Specific checklists, derived from the WHO laboratory biosafety manual, which in Italy are also defined by the National Institute of Labor Safety Insurance (Istituto Nazionale Assicurazione Infortuni sul Lavoro) in its 6th Fascicle published in 2010 3 are necessary to verify the compliance of a given laboratory with the required biosafety level

Generation of cattle knockout for galactose‐α1,3‐galactose and N‐glycolylneuraminic acid antigens

Two well-characterized carbohydrate epitopes are absent in humans but present in other mammals. These are galactose-α1,3-galactose (αGal) and N-glycolylneuraminic acid (Neu5Gc) which are introduced by the activities of two enzymes including α(1,3) galactosyltransferase (encoded by the GGTA1 gene) and CMP-Neu5Gc hydroxylase (encoded by the CMAH gene) that are inactive in humans but present in cattle. Hence, bovine-derived products are antigenic in humans who receive bioprosthetic heart valves (BHVs) or those that suffer from red meat syndrome. Using programmable nucleases, we disrupted (knockout, KO) GGTA1 and CMAH genes encoding for the enzymes that catalyse the synthesis of αGal and Neu5Gc, respectively, in both male and female bovine fibroblasts. The KO in clonally selected fibroblasts was detected by polymerase chain reaction (PCR) and confirmed by Sanger sequencing. Selected fibroblasts colonies were used for somatic cell nuclear transfer (SCNT) to produce cloned embryos that were implanted in surrogate recipient heifers. Fifty-three embryos were implanted in 33 recipients heifers; 3 pregnancies were carried to term and delivered 3 live calves. Primary cell cultures were established from the 3 calves and following molecular analyses confirmed the genetic deletions. FACS analysis showed the double-KO phenotype for both antigens confirming the mutated genotypes. Availability of such cattle double-KO model lacking both αGal and Neu5Gc offers a unique opportunity to study the functionality of BHV manufactured with tissues of potentially lower immunogenicity, as well as a possible new clinical approaches to help patients with red meat allergy syndrome due to the presence of these xenoantigens in the diet

Stereotactic Ablative Radiotherapy (SABR) in inoperable oligometastatic disease from colorectal cancer: a safe and effective approach

Background: To assess the safety and efficacy of Stereotactic Ablative Radiotherapy (SABR) in oligometastatic patients from colorectal cancer. Methods: 82 patients with 1-3 inoperable metastases confined to one organ (liver or lung), were treated with SABR for a total of 112 lesions in an observational study. Prescription dose ranged between 48 and 75Gy in 3 or 4 consecutive fractions. Primary end-points were local control (LC), overall survival (OS) and progression-free survival (PFS). Secondary end-point was toxicity. Results: Median follow-up was 24 months (range 3-47). One, two and three years LC rate was 90%,80% and 75% (85%,75% and 70% for lung and 95%, 90% and 85% for liver metastases; no statistically significance was found). The difference in LC between the subgroup of lesions treated with >= 60 Gy (n = 58) and those irradiated with 3 cm (p 3 toxicity. Conclusions: SABR is a safe and feasible alternative treatment of oligometastatic colorectal liver and lung metastases in patients not amenable to surgery or other ablative treatments

Characterization of immunogenic Neu5Gc in bioprosthetic heart valves

Background: The two common sialic acids (Sias) in mammals are N-acetylneuraminic acid (Neu5Ac) and its hydroxylated form N-glycolylneuraminic acid (Neu5Gc). Unlike most mammals, humans cannot synthesize Neu5Gc that is considered foreign and recognized by circulating antibodies. Thus, Neu5Gc is a potential xenogenic carbohydrate antigen in bioprosthetic heart valves (BHV) that tend to deteriorate in time within human patients. Methods: We investigated Neu5Gc expression in non-engineered animal-derived cardiac tissues and in clinically used commercial BHV, and evaluated Neu5Gc immunogenicity on BHV through recognition by human anti-Neu5Gc IgG. Results: Neu5Gc was detected by immunohistochemistry in porcine aortic valves and in porcine and bovine pericardium. Qualitative analysis of Sia linkages revealed Siaa2-3> Siaa2-6 on porcine/bovine pericardium while the opposite in porcine aortic/pulmonary valve cusps. Similarly, six commercial BHV containing either porcine aortic valve or porcine/bovine/equine pericardium revealed Siaa2-3> Siaa2-6 expression. Quantitative analysis of Sia by HPLC showed porcine/bovine pericardium express 4-fold higher Neu5Gc levels compared to the porcine aortic/pulmonary valves, with Neu5Ac at 6-fold over Neu5Gc. Likewise, Neu5Gc was expressed on commercial BHV (186.3 +/- 16.9 pmol Sia/mu g protein), with Neu5Ac at 8-fold over Neu5Gc. Affinity-purified human anti-Neu5Gc IgG showing high specificity toward Neu5Gc-glycans (with no binding to Neu5Ac-glycans) on a glycan microarray, strongly bound to all tested commercial BHV, demonstrating Neu5Gc immune recognition in cardiac xenografts. Conclusions: We conclusively demonstrated Neu5Gc expression in native cardiac tissues, as well as in six commercial BHV. These Neu5Gc xeno-antigens were recognized by human anti-Neu5Gc IgG, supporting their immunogenicity. Altogether, these findings suggest BHV-Neu5Gc/anti-Neu5Gc may play a role in valve deterioration warranting further investigation