Stefano Bonardelli

The Challenge of Diagnosing Atheroembolic Renal Disease: Clinical Features and Prognostic Factors

Background— Atheroembolic renal disease (AERD) is caused by showers of cholesterol crystals released by eroded atherosclerotic plaques. Embolization may occur spontaneously or after angiographic/surgical procedures. We sought to determine clinical features and prognostic factors of AERD. Methods and Results— Incident cases of AERD were enrolled at multiple sites and followed up from diagnosis until dialysis and death. Diagnosis was based on clinical suspicion, confirmed by histology or ophthalmoscopy for all spontaneous forms and for most iatrogenic cases. Cox regression was used to model time to dialysis and death as a function of baseline characteristics, AERD presentation (acute/subacute versus chronic renal function decline), and extrarenal manifestations. Three hundred fifty-four subjects were followed up for an average of 2 years. They tended to be male (83%) and elderly (60% >70 years) and to have cardiovascular diseases (90%) and abnormal renal function at baseline (83%). AERD occurred spontaneously in 23.5% of the cases. During the study, 116 patients required dialysis, and 102 died. Baseline comorbidities, ie, reduced renal function, presence of diabetes, history of heart failure, acute/subacute presentation, and gastrointestinal tract involvement, were significant predictors of event occurrence. The risk of dialysis and death was 50% lower among those receiving statins. Conclusions— Clinical features of AERD are identifiable. These make diagnosis possible in most cases. Prognosis is influenced by disease type and severity.

Isolation and characterization of lymphatic microvascular endothelial cells from human tonsils

Human lymphatic endothelial cells (LECs) have isolated prevalently from human derma and tumors. As specialized lymphatic organs within the oropharynx, palatine tonsils are easily obtained and rich in lymphatic venules. Using a two‐step purification method based on the sorting of endothelial cells with Ulex Europaeus Agglutinin 1 (UEA‐1)‐coated beads, followed by purification with monoclonal antibody D2–40, we successfully purified LECs from human palatine tonsils. The LECs were expanded on flasks coated with collagen type 1 and fibronectin for up to 8–10 passages and then analyzed for phenotypic and functional properties. Cultured cells retained the phenotypic pattern of the lymphatic endothelium of palatine tonsils and expressed functional VEGFR‐3 molecules. In fact, stimulation with VEGFR‐3 ligand, the vascular endothelium grow factor C, induced a marked increase in cell proliferation. Similarly to blood endothelial cells (BECs), LECs were able to form tube‐like structure when seeded in Cultrex basement membrane extract. Comparative studies performed on LECs derived from palatine tonsils and iliac lymphatic vessels (ILVs), obtained with the same procedures, showed substantial discrepancies in the expression of various lymphatic markers. This points to the existence of micro‐ and macrovessel‐derived LECs with different phenotypes, possibly involving different biological activities and functions. Palatine tonsil‐ and ILV‐derived LECs may, therefore, represent new models for investigating function and biochemical properties of these lymphatic endothelia. J. Cell. Physiol. 207: 107–113, 2006.

Phenotypic and functional characteristics of tumour-derived microvascular endothelial cells.

We recently developed a method for the isolation and purification of tumour-derived endothelium. In this study the phenotypic and functional properties of human tumour-derived microvascular endothelial cells (TdMEC) were examined. Endothelium obtained from human adrenal gland specimens (HAMEC) was used as a reference microvascular endothelial cell population. TdMEC formed a confluent monolayer with the typical morphological appearance of endothelium and were positive for endothelial markers such as Ulex-1 lectin, CD31 antigen, von Willebrand Factor and VE-cadherin. The addition of acidic Fibroblast Growth Factor (aFGF), basic FGF (bFGF) or Vascular Endothelial Growth Factor (VEGF) substantially improved proliferation of TdMEC; and kidney carcinoma derived endothelial cells were more responsive to FGFs, whereas glioblastoma derived endothelial cells greatly responded to VEGF. TdMEC expressed high levels of the VEGF receptors, KDR/flk-1 and Flt-1, as shown by northern blot analysis. TdMEC expressed the adhesion molecules ICAM-1, VCAM-1 and E-selectin that could be further increased by exposing TdMEC culture to interleukin-1. All the TdMEC expressed interleukin-8 mRNA. These findings show that TdMEC in vitro maintain several of the features described for microvasculature. Thus, TdMEC represent a useful tool to study markers for tumor vasculature.

Human cytomegalovirus productively infects lymphatic endothelial cells and induces a secretome that promotes angiogenesis and lymphangiogenesis through interleukin-6 and granulocyte–macrophage colony-stimulating factor

Endothelial cells (ECs) are a site of human cytomegalovirus (HCMV) productive replication, haematogenous dissemination and persistence, and are assumed to play a critical role in the development of HCMV-associated vascular diseases. Although early reports have shown the presence of HCMV antigens and DNA in lymphoid tissues, the ability of HCMV to infect lymphatic ECs (LECs) has remained unaddressed due to the lack of a suitable in vitro system. This study provided evidence that a clinical isolate of HCMV (retaining its natural endotheliotropism) was able to productively infect purified lymph node-derived LECs and that it dysregulated the expression of several LEC genes involved in the inflammatory response to viral infection. Qualitative and quantitative analysis of virus-free supernatants from HCMV-infected LEC cultures revealed virus-induced secretion of several cytokines, chemokines and growth factors, many of which are involved in the regulation of EC physiological properties. Indeed, functional assays demonstrated that the secretome produced by HCMV-infected LECs stimulated angiogenesis in both LECs and blood ECs, and that neutralization of either interleukin (IL)-6 or granulocyte-macrophage colony-stimulating factor (GM-CSF) in the secretome caused the loss of its angiogenic properties. The involvement of IL-6 and GM-CSF in the HCMV-mediated angiogenesis was further supported by the finding that the recombinant cytokines reproduced the angiogenic effects of the HCMV secretome. These findings suggest that HCMV induces haemangiogenesis and lymphangiogenesis through an indirect mechanism that relies on the stimulation of IL-6 and GM-CSF secretion from infected cells.

Is mild vascular cognitive impairment reversible? Evidence from a study on the effect of carotid endarterectomy

Abstract Mild vascular cognitive impairment (mVCI) is a broader term that is intended to detect cognitive loss before the development of dementia. The identification of preventable risk factors as well as therapeutic strategies of intervention is still unclear. It has been suggested that carotid endarterectomy (CEA) improves cognitive functions, beyond the well-known preventive effect upon future stroke events. In the present study, we evaluated the beneficial effect of CEA in restoring mVCI. Among a large sample of subjects, who underwent CEA for severe carotid stenosis, two groups were identified according to the absence (CON) or the presence of cognitive impairment (mVCI). A multidimensional neuropsychological and behavioural assessment was performed in the week prior, and at a 3-month follow-up after CEA. The incidence of mVCI in this sample was 38%. Seventy-eight patients completed the follow-up (48 CON, 30 mVCI). Both groups showed a clinical improvement after CEA, although the effect was significantly higher in the mVCI group in regard to verbal memory (short story, p < 0.05), and attention (digit span, p < 0.05) scores. At follow-up, 60% of mVCI subjects were classified as having normal cognitive functions. Index of disease severity and peripheral arterial disease were found to be the predictors of improvement. These findings support that mVCI represents a heterogeneous, in some cases reversible condition. CEA might be considered a therapeutic option to treat and prevent cognitive decline in mVCI patients.

Predictors and outcomes of acute kidney injury after thoracic aortic endograft repair

BACKGROUND This study analyzed the incidence and the predictive factors of postoperative acute kidney injury (AKI) after thoracic endovascular aortic repair (TEVAR) and evaluated the effect of AKI on postoperative survival. METHODS Between November 2000 and April 2011, all consecutive patients undergoing TEVAR of the descending thoracic or thoracoabdominal aorta were enrolled at four teaching hospitals. Estimated glomerular filtration rate (eGFR) was evaluated during the entire hospitalization. AKI was defined by the RIFLE (Risk, Injury, Failure, Loss of function, End-stage renal disease) consensus criteria. RESULTS The study included 171 patients (80% men) who were a mean age of 69±14 years (range, 18-87 years). AKI occurred in 24 patients (14%). Independent predictors of postoperative AKI were preoperative depressed eGFR, thoracoabdominal extent, and postoperative transfusion. Patients with AKI experienced major postoperative complications (P=.001), longer hospitalization (P=.008), and higher hospital mortality (29% vs 4%; P<.001). Kaplan-Meier analysis showed a survival of 82%, 51%, and 51% at 1, 3, and 5 years for patients who developed AKI, which was significantly worse than the 99%, 89%, and 80% for patients who did not experience AKI (P=.001). CONCLUSIONS Preoperative poor renal function, blood transfusions, and the thoracoabdominal extent of the aortic disease were the most important predictors for AKI.

Monitoring of Somatosensory Evoked Potentials During Carotid Endarterectomy: Relationship with Different Haemodynamic Parameters and Clinical Outcome

The authors studied the changes of Somatosensory Evoked Potentials (SEPs) performed in 241 anaesthetised patients during 264 carotid endarterectomies (CEs). SEP responses were considered significantly modified when the central conduction time was greater than 1 ms and/or when the amplitude of the complex N20-P25 decreased by at least 50%. Both CCT and N20-P25 have been correlated with different parameters, including the presence or absence of preoperative neurological deficits, the type of general anaesthesia, the status of the contralateral and the ipsilateral carotid artery, stump pressure, the use of an intraluminal shunt and the perioperative results. After carotid cross-clamping SEP responses were within the normal range in 236 CEs (89%), and abnormal in 28 (11%). A shunt was inserted 23 times in 264 (9%) cases. None of the patients operated on in this series experienced a permanent neurological deficit; there were three (1.1%) transient deficits (two Rinds and one TIA) and two deaths from non-neurological causes. Only one of the transient deficits was present when the patient woke from the anaesthetic and this event was predicted by significant modification of the SEP which did not reverse after removal of the clamps (a shunt was not used). None of the patients in our series who did not present significant modifications of SEPs during the operation had any postoperative neurological deficit. The authors conclude that SEP recording is a highly reliable and objective method for continuous monitoring of brain function during CE.

A prospective randomized trial on azathioprine addition to cyclosporine versus cyclosporine monotherapy at steroid withdrawal, 6 months after renal transplantation

BACKGROUND Many attempts have been made to withdraw steroid therapy in renal transplant patients in order to avoid its many side effects. Results have been, so far, controversial. In this randomized prospective study, we compare the efficacy of azathioprine adjuncts to cyclosporine at the time of steroid withdrawal, 6 months after transplantation, versus Cyclosporine monotherapy, in preventing acute rejection. METHODS One hundred and sixteen kidney transplant patients with good and stable renal function (creatininemia <2 mg/dl) received, in the first 6 months, cyclosporine + steroid. They were then randomized into two groups (A and B), and steroid therapy was withdrawn over 2 months. Group A (58 patients) continued on cyclosporine monotherapy, whereas group B (58 patients) added azathioprine (1 mg/kg/day) at the beginning of randomization and continued on cyclosporine + azathioprine. In both groups, patients resumed steroid therapy at the first episode of acute rejection. Follow-up after randomization was 5.3+/-1.6 years. RESULTS After 5 years, the incidence of steroid resumption was 57% in group A and 29% in group B (P<0.02); of those, 68% and 88% of them were within 6 months from randomization. Anti-rejection therapy was always successful. Five-year patient and graft survival rates were 90% and 88% in group A and 100% and 91% in group B. Creatininemia did not differ, at follow-up. Side effects differed only for mild and reversible leukopenia caused by azathioprine in group B. CONCLUSION Cyclosporine plus azathioprine is more effective than cyclosporine monotherapy in reducing the incidence of acute rejection after steroid withdrawal. Graft loss as a result of chronic rejection, mild in both groups, did not differ. Steroid withdrawal is feasible and advantageous, and the addition of azathioprine allowed 71% of our selected patients to remain steroid-free.

Combined endovascular and surgical approach (hybrid treatment) for management of type IV thoracoabdominal aneurysm.

The purpose of this article is to report successful hybrid treatment of a sovraprosthetis type IV thoracoabdominal aneurysm. This technique was used in a 65-year-old man with chronic rupture of a type IV thoracoabdominal aneurysm not suitable for aortic cross-clamping because of a severe cardiopathy (left ventricular ejection fraction 20%); the patient underwent previous repairs of aortic arch and infrarenal abdominal aortic aneurysms. Perioperative complications were absent. Postoperative day 21 computed tomography and monthly duplex ultrasonography confirmed the complete exclusion of the aneurysm with proper perfusion of visceral vessels. At the seventh postoperative month, the patient died of a massive recurrence of myocardial infarction. Hybrid treatment for thoracoabdominal aneurysms may represent a valid solution for those patients with poor cardiac and respiratory reserve, reducing cardiac stress and the duration of visceral ischemia, which are still the main causes of morbidity and mortality for this type of intervention.

Complete In Vitro Prosthesis Endothelialization Induced by Artificial Extracellular Matrix

This report presents our research on the conditions necessary to substain optimal in vitro prosthetic endothelialization using human endothelium cultures. Human vein endothelial cells were seeded at a concentration of 3 x 10(5)/cm2 in a gelatinized Dacron patch graft coated with a commercial collagen film, using a solution of fibrin glue. Endothelium adhesion, proliferation, and survival were measured by [3H]thymidine incorporation, after 7 days of incubation. Finally, the morphology of prosthetic endothelialization was analyzed by scanning electron microscopy. We observed that the Dacron patch grafts coated with collagen film were able to promote endothelialization better than the prostheses coated with highly concentrated collagen solution or gelatin. We therefore concluded that the collagen film that supports endothelial cell adhesion and proliferation uniformly covers the entire synthetic endoluminal surface of the Dacron graft, thus preventing endothelial cell alterations induced by direct contact with the synthetic prosthetic surface.