Apheresis and Cell Therapy Unit

Pavia, Italy

Apheresis and Cell Therapy Unit

Pavia, Italy

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Del Fante C.,Apheresis and Cell Therapy Unit | Scudeller L.,Clinical Epidemiology and Biostatistics Unit | Martinasso A.,Apheresis and Cell Therapy Unit | Viarengo G.,Apheresis and Cell Therapy Unit | Perotti C.,Apheresis and Cell Therapy Unit
Transfusion | Year: 2016

BACKGROUND: Extracorporeal photopheresis (ECP) is an effective cell therapy employed in several diseases, including graft versus host disease (GVHD) and organ rejection. When ECP is performed using an off-line technique, mononuclear cell (MNC) collection by leukapheresis is necessary for further manipulation (addition of 8-methoxypsoralen and ultraviolet A irradiation before reinfusion to the patient). We report the results of the first crossover equivalence study on yield and purity of MNCs collected from patients undergoing ECP with two different automated systems: MNC and CMNC (working with intermittent and continuous-flow collection, respectively), released by Terumo BCT. STUDY DESIGN AND METHODS: Fifty-one patients (15 males and 36 females) with GVHD or chronic lung allograft dysfunction were consecutively enrolled and randomly assigned to MNC collection alternatively by the CMNC or MNC system within each ECP cycle (two procedures) in two consecutive cycles. ECP procedures were performed using the off-line technique, according to our internal protocol, processing 1.5 blood volumes. RESULTS: A total of 204 ECP procedures were evaluated. The MNC system showed a higher MNC concentration capacity than the CMNC (mean difference, −13.46; 95% confidence interval, −21.05 to −5.47; p < 0.001). Collection efficiency was higher with the CMNC system as well as total MNC bag content, while MNC bag purity was equivalent for both systems. Platelet loss was higher with the CMNC. Equivalence was shown for MNC purity and anticoagulant infused to the patient. CONCLUSION: CMNC and MNC systems offer different advantages in different clinical conditions, and both are safe and efficient in collecting MNCs for ECP. © 2016 AABB


PubMed | Clinical Epidemiology and Biostatistics Unit and Apheresis and Cell Therapy Unit
Type: Journal Article | Journal: Transfusion | Year: 2016

Extracorporeal photopheresis (ECP) is an effective cell therapy employed in several diseases, including graft versus host disease (GVHD) and organ rejection. When ECP is performed using an off-line technique, mononuclear cell (MNC) collection by leukapheresis is necessary for further manipulation (addition of 8-methoxypsoralen and ultraviolet A irradiation before reinfusion to the patient). We report the results of the first crossover equivalence study on yield and purity of MNCs collected from patients undergoing ECP with two different automated systems: MNC and CMNC (working with intermittent and continuous-flow collection, respectively), released by Terumo BCT.Fifty-one patients (15 males and 36 females) with GVHD or chronic lung allograft dysfunction were consecutively enrolled and randomly assigned to MNC collection alternatively by the CMNC or MNC system within each ECP cycle (two procedures) in two consecutive cycles. ECP procedures were performed using the off-line technique, according to our internal protocol, processing 1.5 blood volumes.A total of 204 ECP procedures were evaluated. The MNC system showed a higher MNC concentration capacity than the CMNC (mean difference, -13.46; 95% confidence interval, -21.05 to -5.47; p<0.001). Collection efficiency was higher with the CMNC system as well as total MNC bag content, while MNC bag purity was equivalent for both systems. Platelet loss was higher with the CMNC. Equivalence was shown for MNC purity and anticoagulant infused to the patient.CMNC and MNC systems offer different advantages in different clinical conditions, and both are safe and efficient in collecting MNCs for ECP.


Perotti C.,Apheresis and Cell Therapy Unit | Del Fante C.,Apheresis and Cell Therapy Unit | Tinelli C.,Apheresis and Cell Therapy Unit | Viarengo G.,Apheresis and Cell Therapy Unit | And 4 more authors.
Transfusion | Year: 2010

BACKGROUND: Extracorporeal photochemotherapy (ECP) is a valid therapeutic option in the treatment of acute and chronic graft-versus-host disease (aGVHD and cGVHD, respectively). No standard clinical and laboratory criteria of response to ECP treatment are available at the moment. STUDY DESIGN AND METHODS: Clinical and laboratory variables on 73 pediatric patients with aGVHD (n = 50) and cGVHD (n = 23) were correlated with response to ECP and survival. RESULTS: An overall response (OR) was obtained in 34 of 50 (68%) aGVHD and in 16 of 23 (69.5%) cGVHD patients. Steroid tapering within 30 days of 1.3 mg/kg in OR (p = 0.004) was the sole highly significant correlation with response found in aGVHD while no correlation emerged for cGVHD (p = 0.28). Among aGVHD patients, response to ECP was inversely associated with death: among OR, deaths were 13 of 34 (38.2%), while among nonresponders, deaths were 15 of 16 (93.8%; p < 0.001). On the other hand, decrease of steroid dose at 30 days was associated with survival: for each 1 mg/kg reduction, the hazard ratio was 2.2, and the 95% confidence interval was 1.5 to 3.2 (p < 0.001). No other clinical or laboratory variables statistically associated with survival were found. CONCLUSIONS: Our results demonstrate that steroid tapering within the first 30 days of ECP treatment in aGVHD and response to ECP in acute and chronic GVHD are the only variables influencing response and survival, respectively. © 2010 American Association of Blood Banks.


Sandri G.,University of Pavia | Bonferoni M.C.,University of Pavia | Rossi S.,University of Pavia | Ferrari F.,University of Pavia | And 8 more authors.
Expert Opinion on Drug Delivery | Year: 2015

Introduction: The work presents the development of acellular scaffolds extemporaneously embedded with platelet lysate (PL), as an innovative approach in the field of tissue regeneration/reparation. PL embedded scaffolds should have a tridimensional architecture to support cell migration and growth, in order to restore skin integrity. For this reason, chondroitin sulfate (CS) was associated with sodium alginate (SA) to prepare highly porous systems.Methods: The developed scaffolds were characterized for chemical stability to γ-radiation, morphology, hydration and mechanical properties. Moreover, the capability of fibroblasts and endothelial cells to populate the scaffold was evaluated by means of proliferation test 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and confocal laser scanning microscopy study. The scaffolds, not altered by sterilization, were characterized by limited swelling and high flexibility, by foam-like structure with bubbles that formed a high surface area and irregular texture suitable for cell adhesion.Results: Cell growth and scaffold population were evident on the bubble surface, where the cells appeared anchored to the scaffold structure.Conclusion: Scaffold network based on CS and SA demonstrated to be an effective support to enhance and to allow fibroblasts and endothelial cells (human umbilical vein endothelial cells, HUVEC) adhesion and proliferation. In particular, it could be hypothesized that cell adhesion was facilitated by the synergic effect of PL and CS. Although further in vivo evaluation is needed, on the basis of in vitro results, PL embedded scaffolds seem promising systems for skin wound healing. © 2014 Informa UK, Ltd.


Del Fante C.,Apheresis and Cell Therapy Unit | Scudeller L.,Apheresis and Cell Therapy Unit | Viarengo G.,Apheresis and Cell Therapy Unit | Bernasconi P.,Apheresis and Cell Therapy Unit | Perotti C.,Apheresis and Cell Therapy Unit
Transfusion | Year: 2012

BACKGROUND: Chronic graft-versus-host disease (cGVHD) classification has recently been improved by the National Institutes of Health (NIH); patients' stratification with those new criteria has implications for patients' prognosis. STUDY DESIGN AND METHODS: To assess whether the NIH consensus classification (NCC) better predicts survival and response to extracorporeal photochemotherapy (ECP), and to identify variables associated with response and survival, we retrospectively analyzed 102 patients with cGVHD reclassified according to NCC treated with ECP (1997-2010) at our center. Cox regression was used in univariate and multivariate models. RESULTS: Of the 102 patients, 64 (62.7%) had classic cGVHD, 24 (23.5%) had overlap cGVHD, and seven (6.9%) patients each had late and persistent acute GVHD. The cumulative ECP-specific follow-up was 2333.3 person-years. Response was complete in 16 (15.7%), partial in 38 (37.3%), minimal in 28 (27.5%), and absent in 20 (19.6%). Of the 22 deaths, 15 (68.2%) occurred among patients with minimal or no response (p = 0.031). The only variables associated with response were nonmyeloablative transplant (hazard ratio, 3.5; 95% confidence interval [CI], 1.36-9.08; p = 0.009), donor lymphocyte infusion (hazard ratio, 2.58; 95% CI, 1.2-5.56; p = 0.015), and lung involvement (hazard ratio, 0.34; 95% CI, 0.12-0.94; p = 0.038). CONCLUSION: ECP is a safe and effective treatment for cGVHD and response to ECP is the only variable that influences survival. We found no correlation between response and NCC clinical subtype, number, or degree of organ involvement, except for lung, or the variables mentioned above. Prospective studies are needed to identify subsets of patients with higher probability of response. © 2012 American Association of Blood Banks.


Sandri G.,University of Pavia | Bonferoni M.C.,University of Pavia | D'Autilia F.,University of Pavia | Rossi S.,University of Pavia | And 7 more authors.
European Journal of Pharmaceutics and Biopharmaceutics | Year: 2013

The management of difficult to heal wounds can considerably reduce the time required for tissue repairing and promote the healing process, minimizing the risk of infection. Silver compounds, especially silver sulfadiazine (AgSD), are often used to prevent or to treat wound colonization, also in presence of antibiotic-resistant bacteria. However, AgSD has been shown to be cytotoxic in vitro toward fibroblasts and keratinocytes and consequently to retard wound healing in vivo. Recently, platelet lysate (PL) has been proposed in clinical practice for the healing of persistent lesions. The aim of the present work was the development of wound dressings based on AgSD loaded in solid lipid nanoparticles (SLNs), to be used in association with PL for the treatment for skin lesions. SLN were based on chondroitin sulfate and sodium hyaluronate, bioactive polymers characterized by well-known tissue repairing properties. The encapsulation of AgSD in SLN aimed at preventing the cytotoxic effect of the drug on normal human dermal fibroblasts (NHDFs) and at enabling the association of the drug with PL. SLN were loaded in wound dressings based on hydroxypropylmethyl cellulose (HPMC) or chitosan glutamate (CS glu). These polymers were chosen to obtain a sponge matrix with suitable elasticity and softness and, moreover, with good bioadhesive behavior on skin lesions. Dressings based on chitosan glutamate showed antimicrobial activity with and without PL. Even though further in vivo evaluation could be envisaged, chitosan based dressings demonstrated to be a suitable prototype for the treatment for skin lesions. © 2012 Elsevier B.V. All rights reserved.


Malara A.,University of Pavia | Malara A.,Biotechnology Research Laboratories | Currao M.,University of Pavia | Currao M.,Biotechnology Research Laboratories | And 9 more authors.
Stem Cells | Year: 2014

Megakaryocytes associate with the bone marrow vasculature where they convert their cytoplasm into proplatelets that protrude through the vascular endothelium into the lumen and release platelets. The extracellular matrix (ECM) microenvironment plays a critical role in regulating these processes. In this work we demonstrate that, among bone marrow ECM components, fibronectin, type IV collagen, and laminin are the most abundant around bone marrow sinusoids and constitute a pericellular matrix surrounding megakaryocytes. Most importantly, we report, for the first time, that megakaryocytes express components of the basement membrane and that these molecules contribute to the regulation of megakaryocyte development and bone marrow ECM homeostasis both in vitro and in vivo. In vitro, fibronectin induced a threefold increase in the proliferation rate of mouse hematopoietic stem cells leading to higher megakaryocyte output with respect to cells treated only with thrombopoietin or other matrices. However, megakaryocyte ploidy level in fibronectin-treated cultures was significantly reduced. Stimulation with type IV collagen resulted in a 1.4-fold increase in megakaryocyte output, while all tested matrices supported proplatelet formation to a similar extent in megakaryocytes derived from fetal liver progenitor cells. In vivo, megakaryocyte expression of fibronectin and basement membrane components was upregulated during bone marrow reconstitution upon 5-fluorouracil induced myelosuppression, while only type IV collagen resulted upregulated upon induced thrombocytopenia. In conclusion, this work demonstrates that ECM components impact megakaryocyte behavior differently during their differentiation and highlights a new role for megakaryocyte as ECM-producing cells for the establishment of cell niches during bone marrow regeneration. © AlphaMed Press 2013.


Del Fante C.,Apheresis and Cell Therapy Unit | Scudeller L.,Apheresis and Cell Therapy Unit | Viarengo G.,Apheresis and Cell Therapy Unit | Cervio M.,Apheresis and Cell Therapy Unit | Perotti C.,Apheresis and Cell Therapy Unit
Transfusion | Year: 2013

Background Extracorporeal photochemotherapy (ECP) is an effective cell therapy employed in several diseases, including graft-versus-host disease (GVHD) and organ rejection after transplantation. When ECP is performed using the off-line technique, mononuclear cell (MNC) collection by leukapheresis is necessary for further manipulation. Semiautomated apheresis systems require experienced personnel to obtain a good MNC collection; an automated device, able to efficiently collect MNCs with high purity, is desirable. We compared the semiautomated COBE Spectra MNC and the new automated Spectra Optia v.5.0 MNC (Terumo BCT) devices in terms of efficacy and safety. Study Design and Methods Adult patients with GVHD or bronchiolitis obliterans syndrome (BOS) after lung transplant undergoing ECP at our center were alternatively assigned, within the same ECP cycle (composed by two procedures each), to MNC collection with either device. Patients' characteristics, procedure, and product-related variables were compared. Results Thirty-nine patients (24 with GVHD and 15 with BOS) underwent a total of 126 ECP procedures, with good compliance to both devices. Product volume and platelet (PLT) and red blood cell contamination were significantly lower with the Spectra Optia. MNC collection efficiency (CE), purity, and PLT loss were similar between the two devices, while white blood cells CE was in favor of the COBE Spectra. Conclusion The Spectra Optia device proved to be a good option for MNC collection in the difficult ECP setting, since it ensures high-quality MNC collection, while at the same time saving personnel's time, guaranteeing optimal monitoring and care to this frail patient population. © 2013 American Association of Blood Banks.


PubMed | Azienda Ospedaliera della Provincia di Lodi, University of Pavia and Apheresis and Cell Therapy Unit
Type: Journal Article | Journal: International journal of pharmaceutics | Year: 2016

Hemoderivative tear substitutes contain various ephiteliotrophic factors, such as growth factors (GF), involved in ocular surface homeostasis without immunogenic properties. The aim of the present work was the loading of platelet lysate into contact lenses to improve the precorneal permanence of platelet lysate growth factors on the ocular surface to enhance the treatment of corneal lesions. To this purpose, chondroitin sulfate, a sulfated glycosaminoglycan, which is normally present in the extracellular matrix, was associated with platelet lysate. In fact, chondroitin sulfate is capable of electrostatic interaction with positively charged growth factors, in particular, with bFGF, IGF, VEGF, PDGF and TGF-, resulting in their stabilization and reduced degradation in solution. In the present work, various types of commercially available contact lenses have been loaded with chondroitin sulfate or chondroitin sulfate in association with platelet lysate to achieve a release of growth factors directly onto the corneal surface lesions. One type of contact lenses (PureVision()) showed in vitro good proliferation properties towards corneal cells and were able to enhance cut closure in cornea constructs.


PubMed | Humanitas Clinical and Research Center, National Diagnostics, University of Pavia, The International Center for Genetic Engineering and Biotechnology and Apheresis and Cell Therapy Unit
Type: Journal Article | Journal: Stem cells (Dayton, Ohio) | Year: 2016

Fibronectin (FN) is a major extracellular matrix protein implicated in cell adhesion and differentiation in the bone marrow (BM) environment. Alternative splicing of FN gene results in the generation of protein variants containing an additional EIIIA domain that sustains cell proliferation or differentiation during physiological or pathological tissue remodeling. To date its expression and role in adult hematopoiesis has not been explored. In our research, we demonstrate that during physiological hematopoiesis a small fraction of BM derived FN contains the EIIIA domain and that mice constitutively including (EIIIA(+/+) ) or excluding (EIIIA(-/-) ) the EIIIA exon present comparable levels of hematopoietic stem cells, myeloid and lymphoid progenitors within BM. Moreover, only minor alterations were detected in blood parameters and in hematopoietic frequencies of BM granulocytes/monocytes and B cells. As opposed to other tissues, unique compensatory mechanisms, such as increased FN accumulation and variable expression of the EIIIA receptors, Toll like receptor-4 and alpha9 integrin subunit, characterized the BM of these mice. Our data demonstrate that FN is a fundamental component of the hematopoietic tissue and that the EIIIA exon may play a key role in modulating hematopiesis in conditions of BM stress or diseases. Stem Cells 2016;34:2263-2268.

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