A simplified regimen of targeted antifungal prophylaxis in liver transplant recipients: A single-center experience.

BACKGROUND: Invasive fungal infection (IFI) is a severe complication of liver transplantation burdened by high mortality. Guidelines recommend targeted rather than universal antifungal prophylaxis based on tiers of risk. METHODS: We aimed to evaluate IFI incidence, risk factors, and outcome after implementation of a simplified two-tiered targeted prophylaxis regimen based on a single broad-spectrum antifungal drug (amphotericin B). Patients presenting 1 or more risk factors according to literature were administered prophylaxis. Prospectively collected data on all adult patients transplanted in Turin from January 2011 to December 2015 were reviewed. RESULTS: Patients re-transplanted before postoperative day 7 were considered once, yielding a study cohort of 581 cases. Prophylaxis was administered to 299 (51.4%) patients; adherence to protocol was 94.1%. Sixteen patients developed 18 IFIs for an overall rate of 2.8%. All IFI cases were in targeted prophylaxis group; none of the non-prophylaxis group developed IFI. Most cases (81.3%) presented within 30 days after transplantation during prophylaxis; predominant pathogens were molds (94.4%). Only 1 case of candidemia was observed. One-year mortality in IFI patients was 33.3% vs 6.4% in patients without IFI (P = .001); IFI attributable mortality was 6.3%. At multivariate analysis, significant risk factors for IFI were renal replacement therapy (OR = 8.1) and re-operation (OR = 5.2). CONCLUSIONS: The implementation of a simplified targeted prophylaxis regimen appeared to be safe and applicable and was associated with low IFI incidence and mortality. Association of IFI with re-operation and renal replacement therapy calls for further studies to identify optimal prophylaxis in this subset of patients.

Liver transplantation in defects of cholesterol biosynthesis: the case of lathosterolosis.

We report the outcome of liver transplantation (LT) in the only surviving patient with lathosterolosis, a defect of cholesterol biosynthesis characterized by high lathosterol levels associated with progressive cholestasis, multiple congenital anomalies and mental retardation. From her diagnosis at age 2 she had shown autistic behavior, was unable to walk unaided and her sight was impaired by cataracts. By age 7 she developed end-stage liver disease. After a soul-searching discussion within the transplantation team, she was treated with LT as this represented her only lifesaving option. At 1-year follow-up, her lathosterol levels had returned to normal (0.61 mg/dL from 13.04 ± 2.65) and her nutrition improved. She began exploring her environment and walking by holding onto an adult's hand and then independently. Her brain magnetic resonance imaging (MRI) had shown a normal picture at age 1, whereas a volume reduction of white matter with ex vacuo ventricular dilatation and defective myelinization were observed before transplant. At 5-year follow-up, a complete biochemical recovery, an arrest of mental deterioration and a stable MRI picture were achieved, with a return to her every day life albeit with limitations. Timely liver transplant in defects of cholesterol biosynthesis might arrest the progression of neurological damage.

2-Aminothiophene-3-carboxylic acid ester derivatives as novel highly selective cytostatic agents.

Cytostatic agents often do not discriminate in their cytostatic potential between different tumor cell types in vitro. In this study, several 2-aminothiophene-3-carboxylic acid ester derivatives were discovered that show an unusual cytostatic selectivity for several T-cell (but not B-cell) lymphoma, prostate cancer, kidney carcinoma and hepatoma cell lines. Their 50 % cytostatic concentrations were generally in the higher nanomolar range and were approximately 20- to 50-fold lower for these tumor cell types than for any other tumor cell line or non-tumorigenic cells. The tumor-selective compounds caused a more preferential suppression of protein synthesis than DNA or RNA synthesis and the prototype compound 3 resulted in an accumulation of prostate cancer cells in the G1 phase of their cell cycle. Compound 3 was also shown to induce apoptosis in prostate cancer cells. The 2-aminothiophene-3-carboxylic acid ester derivatives represent novel candidate cytostatic agents to be further explored for their tumor-selective potential.

Bisphenol A alters ß-hCG and MIF release by human placenta: an in vitro study to understand the role of endometrial cells.

A proper fetomaternal immune-endocrine cross-talk in pregnancy is fundamental for reproductive success. This might be unbalanced by exposure to environmental chemicals, such as bisphenol A (BPA). As fetoplacental contamination with BPA originates from the maternal compartment, this study investigated the role of the endometrium in BPA effects on the placenta. To this end, in vitro decidualized stromal cells were exposed to BPA 1 nM, and their conditioned medium (diluted 1 : 2) was used on chorionic villous explants from human placenta. Parallel cultures of placental explants were directly exposed to 0.5 nM BPA while, control cultures were exposed to the vehicle (EtOH 0.1%). After 24-48 h, culture medium from BPA-treated and control cultures was assayed for concentration of hormone human Chorionic Gonadotropin ( ß -hCG) and cytokine Macrophage Migration Inhibitory Factor (MIF). The results showed that direct exposure to BPA stimulated the release of both MIF and ß -hCG. These effects were abolished/diminished in placental cultures exposed to endometrial cell-conditioned medium. GM-MS analysis revealed that endometrial cells retain BPA, thus reducing the availability of this chemical for the placenta. The data obtained highlight the importance of in vitro models including the maternal component in reproducing the effects of environmental chemicals on human fetus/placenta.

Posthepatectomy liver failure after simultaneous versus staged resection of colorectal cancer and synchronous hepatic metastases.

BACKGROUND: Posthepatectomy liver failure (PHLF) is the third most frequent complication and the major cause of postoperative mortality after resection of colorectal cancer liver metastases (CRLM). In case of synchronous resectable CRLM, it is still unclear if surgical strategy (simultaneous versus staged resection of colorectal cancer and hepatic metastases) influences the incidence and severity of PHLF. The aim of this study was to evaluate the impact of surgical strategy on PHLF and on the early and long-term outcome. PATIENTS AND METHODS: Retrospective study on 106 consecutive patients undergoing hepatectomy for synchronous CRLM between 1997 and 2012. RESULTS: Of 106 patients, 46 underwent simultaneous resection and 60 had staged hepatectomy. The rate of PHLF was similar between groups (16.7% vs 15.2%; p=1) and subgroup analysis restricted to patients undergoing major hepatectomy confirmed this observation (31.8% vs 23.8%; p=0.56). Propensity-score analysis showed that preoperative total bilirubin level and the amount of intra-operative blood transfusion were independently associated with an increased risk of PHLF. Nevertheless, the risk of severe PHLF (grade B - C) was increased in patients who underwent simultaneous resection and major hepatectomy (OR: 4.82; p=0.035). No significant differences were observed in severe (Dindo - Clavien 3 - 4) postoperative morbidity (23.9% vs 20.0%; p=0.64) and survival (3 and 5-year survival: 55% and 34% vs 56% and 33%; p=0.83). CONCLUSIONS: The risk of PHLF is not associated with surgical strategy in the treatment of synchronous CRLM. Nevertheless, the risk of severe PHLF is increased in patients undergoing simultaneous resection and major hepatectomy.

How useful is the machine perfusion in liver transplantation? An answer from a national survey.

Machine perfusion (MP) has been shown worldwide to offer many advantages in liver transplantation, but it still has some gray areas. The purpose of the study is to evaluate the donor risk factors of grafts, perfused with any MP, that might predict an ineffective MP setting and those would trigger post-transplant early allograft dysfunction (EAD). Data from donors of all MP-perfused grafts at six liver transplant centers have been analyzed, whether implanted or discarded after perfusion. The first endpoint was the negative events after perfusion (NegE), which is the number of grafts discarded plus those that were implanted but lost after the transplant. A risk factor analysis for NegE was performed and marginal grafts for MP were identified. Finally, the risk of EAD was analyzed, considering only implanted grafts. From 2015 to September 2019, 158 grafts were perfused with MP: 151 grafts were implanted and 7 were discarded after the MP phase because they did not reach viability criteria. Of 151, 15 grafts were lost after transplant, so the NegE group consisted of 22 donors. In univariate analysis, the donor risk index >1.7, the presence of hypertension in the medical history, static cold ischemia time, and the moderate or severe macrovesicular steatosis were the significant factors for NegE. Multivariate analysis confirmed that macrosteatosis >30% was an independent risk factor for NegE (odd ratio 5.643, p = 0.023, 95% confidence interval, 1.27-24.98). Of 151 transplanted patients, 34% experienced EAD and had worse 1- and 3-year-survival, compared with those who did not face EAD (NoEAD), 96% and 96% for EAD vs. 89% and 71% for NoEAD, respectively (p = 0.03). None of the donor/graft characteristics was associated with EAD even if the graft was moderately steatotic or fibrotic or from an aged donor. For the first time, this study shows that macrovesicular steatosis >30% might be a warning factor involved in the risk of graft loss or a cause of graft discard after the MP treatment. On the other hand, the MP seems to be useful in reducing the donor and graft weight in the development of EAD.

Balancing donor and recipient risk factors in liver transplantation: the value of D-MELD with particular reference to HCV recipients.

Donor-recipient match is a matter of debate in liver transplantation. D-MELD (donor age × recipient biochemical model for end-stage liver disease [MELD]) and other factors were analyzed on a national Italian database recording 5946 liver transplants. Primary endpoint was to determine factors predictive of 3-year patient survival. D-MELD cutoff predictive of 5-year patient survival <50% (5yrsPS<50%) was investigated. A prognosis calculator was implemented (http://www.D-MELD.com). Differences among D-MELD deciles allowed their regrouping into three D-MELD classes (A < 338, B 338-1628, C >1628). At 3 years, the odds ratio (OR) for death was 2.03 (95% confidence interval [CI], 1.44-2.85) in D-MELD class C versus B. The OR was 0.40 (95% CI, 0.24-0.66) in class A versus class B. Other predictors were hepatitis C virus (HCV; OR = 1.42; 95% CI, 1.11-1.81), hepatitis B virus (HBV; OR = 0.69; 95% CI, 0.51-0.93), retransplant (OR = 1.82; 95% CI, 1.16-2.87) and low-volume center (OR = 1.48; 95% CI, 1.11-1.99). Cox regressions up to 90 months confirmed results. The hazard ratio was 1.97 (95% CI, 1.59-2.43) for D-MELD class C versus class B and 0.42 (95% CI, 0.29-0.60) for D-MELD class A versus class B. Recipient age, HCV, HBV and retransplant were also significant. The 5yrsPS<50% cutoff was identified only in HCV patients (D-MELD ≥ 1750). The innovative approach offered by D-MELD and covariates is helpful in predicting outcome after liver transplantation, especially in HCV recipients.

Ethosomes for the delivery of anti-HSV-1 molecules: preparation, characterization and in vitro activity.

This paper describes the production, characterization and in vitro activity of ethosomes containing two molecules with antiviral activity, such as acyclovir (ACY) and N1-beta-D-ribofuranosyl-pyrazole [3,4d]pyridazin-7(6p-chlorine-phenyl)-one nucleoside (N1CP). Ethosomes were prepared and morphologically characterized by Cryo-TEM. The encapsulation efficiency was 92.3 +/- 2.5% for ACY and 94.2 +/- 2.8% for N1CP. The release of the drug from vesicles, determined by a Franz cell method, indicated that both drugs were released in a controlled manner. In order to possibly guarantee the stability during long-term storage ethosome suspensions was freeze-dried. It was found that the freeze-dried ethosomes' cakes were compact, glassy characterized by low density and quick re-hydration. However, the storage time slightly influences the percentage of drug encapsulation within ethosomes showing a drug leakage after re-hydration around 10%. The antiviral activity against HSV-1 of both drugs was tested by plaque reduction assay in monolayer cultures of Vero cells. Data showed that ethosomes allowed a reduction of the ED50 of N1CP evidencing an increase of its antiviral activity. However, ACY remains more active than N1CP. No differences are appreciable between drug-containing ethosomes before and after freeze-drying. Taken together these results, ethosomal formulation could be possibly proposed as mean for topical administration of anti-herpetic molecules.

Graft factors as determinants of postoperative delirium after liver transplantation.

Post-operative delirium (POD) is a frequent complication after surgery, occurring in 15-20% of patients. POD is associated with a higher complications rate and mortality. Literature on POD after liver transplantation (LT) is limited, with the few available studies reporting an incidence of 10-47%. The aim of this study was analyzing pattern, risk factors and clinical impact of POD after LT. Data on donor and recipient characteristics, postoperative course and POD of consecutive adult LT recipients from March 2016 to May 2018 were prospectively collected and retrospectively analyzed. Risk factors for POD were analyzed using univariable logistic regression and Lasso regression. Kaplan-Meier method was used for survival analysis. 309 patients underwent LT during study period; 3 were excluded due to perioperative death. Incidence of POD was 13.4% (n = 41). The median day of onset was 5th (IQR [4-7]) with a median duration of 4 days (IQR [3-7]). Several risk factors, related to the severity of liver disease and graft characteristics, were identified. Graft macrovesicular steatosis was the only factor independently associated with POD at multivariable analysis (OR 1.27, CI 1.09-1.51, p = 0.003). POD was associated with a higher rate of severe postoperative complications and longer intensive care unit and hospital stay, but did not significantly impact on patient and graft survival. Incidence of POD after LT is comparable to that observed after general surgery and graft factors are strongly associated with its onset. These results help identifying a subset of patients to be considered for preventive interventions.

Ischemic preconditioning (IP) of the liver as a safe and protective technique against ischemia/reperfusion injury (IRI).

The aim of the study was to evaluate safety and efficacy of IP in LT, particularly in marginal grafts. From 2007 to 2008, 75 LT donors were randomized to receive IP (IP+) or not (IP-). Considering the graft quality, we divided the main groups in two subgroups (marg+/marg-). IP was performed by 10-min inflow occlusion (Pringle maneuver utilizing a toruniquet). Donor variables considered were gender, age, AST/ALT, ischemia time and steatosis. Recipient variables were gender, age, indication to LT and MELD/CHILD/UNOS score. AST/ALT levels, INR, bilirubin, lactic acid, bile output on postoperative days 1, 3 and 7 were evaluated. Histological analysis was performed evaluating necrosis/steatosis, hepatocyte swelling, PMN infiltration and councilman bodies. Thirty patients received IP+ liver. No differences were seen between groups considering recipient and donor variables. Liver function and AST/ALT levels showed no significant differences between the main two groups. Marginal IP+ showed lower AST levels on day1 compared with untreated marginal livers (936.35 vs. 1268.23; p = 0.026). IP+ livers showed a significant reduction of moderate-severe hepatocyte swelling (33.3% vs. 65.9%; p = 0.043). IP+ patients had a significant reduction of positive early microbiological investigations (36.7% vs. 57.1%; p = 0.042). In our experience IP was safe also in marginal donors, showing a protective role against IRI.

Possible donor-recipient bartonellosis transmission in a pediatric liver transplant.

Bartonella henselae is the causative agent of cat-scratch disease and other disorders, including hepatosplenic granulomatosis. This infection has only rarely been reported after solid organ transplantation, where it can mimic the more common post-transplant lymphoproliferative disease. Here we present a case of asymptomatic B. henselae hepatic and lymph nodal granulomatosis in a pediatric patient who had received orthotopic liver transplant 2 months before; we hypothesize that the causative agent was transmitted from the donor. This infection developed early in the post-transplant period; the disease involved only the graft liver and the regional lymph nodes, and the patient did not have a cat or any history of contact, scratches, or bites by a cat. In our patient this infection resolved successfully with a combination of 2 associated antibiotics and reduction of immunosuppressive therapy.

Evidence for a role of TGF-beta1 in the expression and regulation of alpha-SMA in fetal growth restricted placentae.

There is evidence that alpha-smooth muscle actin (alpha-SMA) is a protein that plays a pivotal role in the production of contractile forces and it is induced by transforming growth factor-beta1 (TGF-beta1). We have analysed the expression of alpha-SMA, TGF-beta1, its receptor RI and the activator phospho-Smad2 in (a) fetal growth restriction pre-eclamptic placentae characterised by early onset and absence of end diastolic velocities in the umbilical arteries (FGR-AED) and (b) control placentae accurately matched for gestational age. The study was performed by immunohistochemical, quantitative Western blotting, ELISA, RT-PCR and in vitro analyses. We found that TGF-beta1 stimulates alpha-SMA production in chorionic villi cultured in vitro. In addition, we observed that in vivo TGF-beta1 concentration is significantly higher in FGR-AED placental samples than in control placentae and that this growth factor could have a paracrine action on villous stroma myofibroblasts expressing TGF-beta1 receptors and phospho-Smad2. Indeed, we report that alpha-SMA undergoes a redistribution in FGR-AED placental villous tree, i.e. we show that alpha-SMA is enhanced in medium and small stem villi and significantly decreased in the peripheral villi. Our data allow us to consider TGF-beta1 and alpha-SMA as key molecules related to FGR-AED placental villous tree phenotypic changes responsible for increased impedance to blood flow observable in this pathology.

Liposomes and micellar dispersions for delivery of benzoheterocyclic derivatives of distamycin A.

In this article we describe the production and characterization of specialized delivery systems for some distamycin derivatives (DD), namely liposomes and micellar dispersions. All the formulations were designed to increase the solubility of DD in an aqueous environment and to reduce the possible toxicity problems related to the administration of these drugs. For instance, liposomes were prepared by reverse phase evaporation technique followed by extrusion through polycarbonate filters, then characterized in terms of dimensions, morphology, and encapsulation efficacy. The analysis of their in vitro antiproliferative activity on cultured human and mouse leukemic cells demonstrated that liposomes and micellar dispersions containing DD exert quite different effects. These effects were compared with those shown by the free drug depending on type of drug and also cell line used.

First Evidence of Cardiac Stem Cells From the Left Ventricular Apical Tip in Patients With Left Ventricular Assist Device Implantation.

BACKGROUND: Recent studies have challenged the dogma that the adult heart is a postmitotic organ and raise the possibility of the existence of resident cardiac stem cells (CSCs). Our study aimed to explore if these CSCs are present in the "ventricular tip" obtained during left ventricular assist device (LVAD) implantation from patients with end-stage heart failure (HF) and the relationship with LV dysfunctional area extent. METHODS: Four consecutive patients with ischemic cardiomyopathy and end-stage HF submitted to LVAD implantation were studied. The explanted "ventricular tip" was used as a sample of apical myocardial tissue for the pathological examination. Patients underwent clinical and echocardiographic examination, both standard transthoracic echocardiography (TTE) and speckle tracking echocardiography (STE), before LVAD implantation. RESULTS: All patients presented severe apical dysfunction, with apical akinesis/diskinesis and very low levels of apical longitudinal strain (-3.5 ± 2.9%). Despite this, the presence of CSCs was demonstrated in pathological myocardial samples of "ventricular tip" in all 4 of the patients. It was found to be a mean of 6 c-kit cells in 10 fields magnification 40×. CONCLUSIONS: Cardiac stem cells can be identified in the LV apical segment of patients who have undergone LVAD implantation despite LV apical fibrosis.

Characterization of toad liver glutathione transferase.

The major form of glutathione transferase from the toad liver previously designed as Bufo bufo liver GST-7.6 (A. Aceto, B. Dragani, T. Bucciarelli, P. Sacchetta, F. Martini, S. Angelucci, F. Amicarelli, M. Miranda and C. Di Ilio, Biochem. J. 289 (1993) 417-422) has been characterized. According to its partial amino acid sequence, the toad enzyme may be included in the pi class GST and named bbGST P2-2. However, bbGST P2-2 appears to be immunologically, structurally and kinetically distinct from any other members of pi family, including bbGST P1-1, suggesting that it may constitute a subset of pi class GST. The data support the hypothesis that the transition from aquatic to terrestrial life causes a switch of the GST amphibian pattern promoting the expression of a GST form (bbGST P2-2) able to counteract, with higher efficiency, the toxic effects of reactive metabolites of oxidative metabolism and those of hydrophobic xenobiotics.

Simultaneous Liver-Kidney Transplantation in Adults: A Single-center Experience Comparing Results With Isolated Liver Transplantation.

BACKGROUND: After introduction of the Model for End-Stage Liver Disease (MELD) score in 2002, a worldwide increasing number of simultaneous liver-kidney transplantations (SLKTx) has been observed. However, organ shortage puts into question the allocation of 2 grafts to 1 recipient. This retrospective, single-center study compared SLKTx results with isolated liver transplantation (LTx). METHODS: Between 1995 and 2013, 37 SLKTx were performed in adult recipients. Every SLKTx was matched by donor age (±5 years) and transplantation date with 2 LTx (n = 74). Pretransplant, intraoperative, and post-transplant variables were collected; liver graft and patient survivals were calculated. RESULTS: As expected, donor age was similar in the 2 groups (median, 39.7 years), whereas serum creatinine level, glomerular filtration rate, and MELD and D-MELD (donor age*MELD) scores were significantly higher in the SLKTx group. SLKTx had longer waiting list time (P = .0034) as well as higher surgical difficulty, testified by more blood transfusions (P = .0083), increased use of classic caval reconstruction (P = .0024), and more frequent need of abdominal packing for bleeding control (P = .0003). In addition, duration of hospital stay (P < .0001), second-look surgery (P = .0082), post-transplant dialysis (P < .0001), and post-transplant infections (P = .04) were significantly greater in SLKTx group. Acute rejection episodes involving the liver were significantly less in SLKTx than in LTx (14% vs 41%; P = .0045). Liver graft and patient survival at 10 years after transplantation was similar in the 2 groups (liver graft: SLKTx, 80% vs LTx, 77% [P = .85]; patient: SLKTx, 86% vs LTx, 79% [P = .56]). CONCLUSIONS: Despite being technically challenging, SLKTx provided excellent long-term results and was shown to be an effective use of liver grafts.

Heterocyclic analogs of DNA minor groove alkylating agents.

It is generally accepted that neoplastic transformation is related to genes alteration or oncogene activation. In particular, DNA minor groove binding drugs have been extensively studied through the years in order to influence the regulation of gene expression by means of specific interactions with DNA bases moieties. Pyrrolo[2,1-c],[1,4].benzodiazepines (PBDs), CC-1065 and distamycins are three classes of minor groove binders which showed interesting cytotoxicity profiles, refined through already reviewed processes of SAR studies. Among the modifications to the three families of antitumor compounds, heterocyclic substitutions have been extensively applied by many groups in order to either modify the reactivity profile or introduce extra interactions within the minor groove, thus changing the binding site or modulating the binding sequence. The updated material related to these modifications has been rationalised and ordered to offer an overview of the argument.

Synthesis and antitumor activity of new benzoheterocyclic derivatives of distamycin A.

The design, synthesis, and in vivo and in vitro antileukemic activity of a novel series of compounds (13-22 and 34), in which different benzoheterocyclic rings, bearing a nitrogen mustard or a benzoyl nitrogen mustard or an alpha-bromoacryloyl group as alkylating moieties, are tethered to a distamycin frame, are reported, and structure-activity relationships are discussed. The new derivatives were prepared by coupling nitrogen mustard-substituted, benzoyl nitrogen mustard-substituted, or alpha-bromoacryloyl-substituted benzoheterocyclic carboxylic acids 23-32 with desformyldistamycin (33) or in one case with its two-pyrrole analogue 35. With very few exceptions, the activities of compounds bearing the same alkylating moiety are slightly affected by the kind of the heteroatom present on the benzoheterocyclic ring. All novel compounds, with one exception, showed in vitro activity against L1210 murine leukemia cell line comparable to or better than that of tallimustine. The compounds in which the nitrogen mustard and the alpha-bromoacryloyl moieties are directly linked to benzoheterocyclic ring showed potent cytotoxic activities (IC(50) ranging from 2 to 14 nM), while benzoyl nitrogen mustard derivatives of benzoheterocycles showed reduced cytotoxic activities, and one compound (16) of this cluster was the sole derivative devoid of significant activity. Compound 18, a 5-nitrogen mustard N-methylindole derivative of distamycin, showed the best antileukemic activity in vivo, with a very long survival time (%T/C = 457), significantly increased in comparison to tallimustine (%T/C = 133), and was selected for further extensive evaluation. Arrested polymerase chain reaction and direct DNA fragmentation assays were performed for compound 18 and the structurally related compounds 13-17 and 19. The results obtained have shown that both alkylating groups and oligopeptide frames play a crucial role in the sequence selectivity of these compounds.