Search

Background: Clinical use of porcine cell-based bioartificial liver (BAL) support in acute liver failure as bridging therapy for liver transplantation exposes the patient to the risk of transmission of porcine endogenous retroviruses (PERVs) to human. This risk may be enhanced when patients receive liver transplant and are subsequently immunosuppressed. As further follow-up of previously reported patients (Di Nicuolo et al. 2005), an assessment of PERV infection was made in the same patient population pharmacologically immunosuppressed for several years after BAL treatment and in healthcare workers (HCWs) involved in the clinical trial at that time., Methods: Plasma and peripheral blood mononuclear cells (PBMCs) from eight patients treated with the Academic Medical Center-BAL (AMC-BAL), who survived to transplant, and 13 HCWs, who were involved in the trial, were assessed to detect PERV infection. A novel quantitative real-time polymerase chain reaction assay has been used., Results: Eight patients who received a liver transplant after AMC-BAL treatment are still alive under long-term pharmacological immunosuppression. The current clinical follow-up ranges from 5.6 to 8.7 yr after BAL treatment. A new q-real-time PCR assay has been developed and validated to detect PERV infection. The limit of quantification of PERV DNA was ≥ 5 copies per 1 × 10(5) PBMCs. The linear dynamic range was from 5 × 10(0) to 5 × 10(6) copies. In both patients and HCWs, neither PERV DNA in PBMCs nor PERV RNA in plasma and PBMC samples have been found., Conclusion: Up to 8.7 yr after exposure to treatment with porcine liver cell-based BAL, no PERV infection has been found in long-term immunosuppressed patients and in HCWs by a new highly sensitive and specific q-real-time PCR assay., (© 2010 John Wiley & Sons A/S.)

General anesthetics and contrast media can cause anaphylactic as well as anaphylactoid reactions. These events are of great concern to radiologists and anesthesiologists because of their relatively high prevalence, possible threat to life, and medical-legal consequences. Points discussed in this review are the critical evaluation of risk factors affecting prevention strategies, the need to be aware of pathogenic mechanisms relevant to prevention strategies, the use of alternative products if a culprit agent is known, the recognition of early signs of a reaction, the need to keep records of reactions on a patient's medical chart, the planning of prophylactic therapy, recommended actions after a reaction to an anesthetic or contrast medium, and the suggested establishment of allergy-anesthesiology centers to improve cooperation, and medical-legal issues. As any drug or contrast medium administered during general anesthesia or a diagnostic procedure can induce a potentially life-threatening or fatal event even in the absence of any evident risk factor in the patient's medical history or clinical status, we usually premedicate susceptible individuals at least to attenuate the severity of an unpredictable reaction, although we cannot rely on the efficacy of premedication to completely prevent a severe event. These recommendations, which are based on the literature and on the experience of our working group, aim to provide useful information for physicians and other specialists who operate in the absence of an allergy consultant.

Background: Currently a number of bioartificial livers (BAL) based on porcine liver cells have been developed as a treatment to bridge acute liver failure patients to orthotopic liver transplantation or liver regeneration. These xenotransplantation related treatments hold the risk of infection of treated patients by porcine endogenous retrovirus (PERV) released from the porcine cells, as in vitro infection experiments and transplantations in immunocompromised mice have shown that PERV is able to infect human cells. The Academic Medical Center (AMC)-BAL, unlike other BALs, is characterized by direct contact between porcine liver cells and human plasma, and might therefore be permissive for PERV transfer., Methods: Prior to a clinical phase I trial, human plasma perfused through the AMC-BAL was investigated for PERV DNA and RNA. Moreover productive infectivity was analyzed by exposing the plasma to HEK-293 cells that were subsequently tested for PERV DNA, PERV RNA and reverse transcriptase activity., Results: Although PERV DNA was detected in the perfused plasma, no productive infectivity was detected. Consequently fourteen patients were treated with the AMC-BAL and monitored for PERV transmission. Immediately after treatment the plasma of the patients was positive for PERV DNA, most probably due to porcine liver cell lysis. The PERV DNA was cleared within 2 weeks post-treatment and no PERV RNA was detected. No productive infectivity in human embryonic kidney (HEK)-293 cells exposed to plasma of treated patients was detectable., Conclusion: To conclude, no release of infective PERV particles from the AMC-BAL was observed. Therefore we consider the AMC-BAL as safe, however careful surveillance of patients will be continued.

Ischemia-reperfusion injury, a clinical problem during cardiac surgery, involves worsened adenosine trisphosphate (ATP) generation and damage to the heart. We studied carbon monoxide (CO) pretreatment, proven valuable in rodents but not previously tested in large animals, for its effects on pig hearts subjected to cardiopulmonary bypass with cardioplegic arrest. Hearts of CO-treated pigs showed significantly higher ATP and phosphocreatine levels, less interstitial edema, and apoptosis of cardiomyocytes and required fewer defibrillations after bypass. We conclude that treatment with CO improves the energy status, prevents edema formation and apoptosis, and facilitates recovery in a clinically relevant model of cardiopulmonary bypass surgery.

The furA-katG region of Mycobacterium tuberculosis, encoding a Fur-like protein and the catalase-peroxidase, is highly conserved among mycobacteria. Both genes are induced upon oxidative stress. In this work we analyzed the M. tuberculosis furA promoter region. DNA fragments were cloned upstream of the luciferase reporter gene, and promoter activity in Mycobacterium smegmatis was measured in both the presence and absence of oxidative stress. The shortest fragment containing an inducible promoter extends 45 bp upstream of furA. In this region, -35 and -10 promoter consensus sequences can be identified, as well as a 23-bp AT-rich sequence that is conserved in the nonpathogenic but closely related M. smegmatis. M. tuberculosis FurA was purified and found to bind upstream of furA by gel shift analysis. A ca. 30-bp DNA sequence, centered on the AT-rich region, was essential for FurA binding and protected by FurA in footprinting analysis. Peroxide treatment of FurA abolished DNA binding. Three different AT-rich sequences mutagenized by site-directed mutagenesis were constructed. In each mutant, both M. tuberculosis FurA binding in vitro and pfurA regulation upon oxidative-stress in M. smegmatis were abolished. Thus, pfurA is an oxidative stress-responsive promoter controlled by the FurA protein.

Recently a phase I clinical trial has been started in Italy to bridge patients with acute liver failure (ALF) to orthotopic liver transplantation (OLT) by the AMC-bioartificial liver (AMC-BAL). The AMC-BAL is charged with 10 × 109 viable primary porcine hepatocytes isolated from a specified pathogen-free (SPF) pig. Here we report a patient with ALF due to acute HBV infection. This patient was treated for 35 h by two AMC-BAL treatments and was bridged to OLT. There was improvement of biochemical and clinical parameters during the treatment. No severe adverse events were observed during treatment and follow-up of 15 months after hospital discharge. Possible porcine endogenous retrovirus (PERV) activity could not be detected in the patient's blood or blood cells up to 12 months after treatment.