Innovative Photonic Sensors for Safety and Security, Part II: Aerospace and Submarine Applications.

The employability of photonics technology in the modern era's highly demanding and sophisticated domain of aerospace and submarines has been an appealing challenge for the scientific communities. In this paper, we review our main results achieved so far on the use of optical fiber sensors for safety and security in innovative aerospace and submarine applications. In particular, recent results of in-field applications of optical fiber sensors in aircraft monitoring, from a weight and balance analysis to vehicle Structural Health Monitoring (SHM) and Landing Gear (LG) monitoring, are presented and discussed. Moreover, underwater fiber-optic hydrophones are presented from the design to marine application.

Innovative Photonic Sensors for Safety and Security, Part I: Fundamentals, Infrastructural and Ground Transportations.

Our group, involving researchers from different universities in Campania, Italy, has been working for the last twenty years in the field of photonic sensors for safety and security in healthcare, industrial and environment applications. This is the first in a series of three companion papers. In this paper, we introduce the main concepts of the technologies employed for the realization of our photonic sensors. Then, we review our main results concerning the innovative applications for infrastructural and transportation monitoring.

Innovative Photonic Sensors for Safety and Security, Part III: Environment, Agriculture and Soil Monitoring.

In order to complete this set of three companion papers, in this last, we focus our attention on environmental monitoring by taking advantage of photonic technologies. After reporting on some configurations useful for high precision agriculture, we explore the problems connected with soil water content measurement and landslide early warning. Then, we concentrate on a new generation of seismic sensors useful in both terrestrial and under water contests. Finally, we discuss a number of optical fiber sensors for use in radiation environments.

Cytokine-Induced Killer (CIK) Cells, In Vitro Expanded under Good Manufacturing Process (GMP) Conditions, Remain Stable over Time after Cryopreservation.

Cytokine-induced killer (CIK) cells are advanced therapy medicinal products, so their production and freezing process has to be validated before their clinical use, to verify their stability as a drug formulation according to the good manufacturing practice (GMP) guidelines. We designed a stability program for our GMP-manufactured CIK cells, evaluating the viability, identity and potency of cryopreserved CIK cells at varying time periods from freezing, and compared them with fresh CIK cells. We evaluated the effects of the cryopreservation method, transportation, and the length of time of different process phases (pre-freezing, freezing and post-thawing) on the stability of CIK cells. This included a worst case for each stage. The expanded CIK cells were viable for up to 30 min from the addition of the freezing solution, when transported on dry ice within 48 h once frozen, within 60 min from thawing and from 12 months of freezing while preserving their cytotoxic effects. The reference samples, cryopreserved simultaneously in tubes and following the same method, were considered representative of the batch and useful in the case of further analysis. Data obtained from this drug stability program can inform the accurate use of CIK cells in clinical settings.

Correction to: Cytokines induced killer cells produced in good manufacturing practices conditions: identification of the most advantageous and safest expansion method in terms of viability, cellular growth and identity.

Following publication of the original article [1], the authors reported that all of the authors' names were processed incorrectly so that their given and family names were interchanged. In this Correction the correct author names are shown. The original publication of this article has been corrected.

Analysis of Mesenchymal Stromal Cell Engraftment After Allogeneic HSCT in Pediatric Patients: A Large Multicenter Study.

The mesenchymal stem cell (MSC) role after allogeneic hematopoietic stem cell transplantation (HSCT) is still a matter of debate; in particular, MSC engraftment in recipient bone marrow (BM) is unclear. A total of 46 patients were analyzed for MSC and hemopoietic stem cell engraftment after HSCT. The majority of patients had the BM as the stem cell source, and acute leukemia was the main indication for HSCT. Mesenchymal and hematopoietic stem cell chimerism analysis was carried out through specific polymorphic tandemly repeated regions. All patients reached complete donor engraftment; no evidence of donor-derived MSC engraftment was noted. Our data indicate that MSCs after HSCT remain of recipient origin despite the following: (i) myeloablative conditioning; (ii) the stem cell source; (iii) the interval from HSCT to BM analysis; (iv) the underlying disease before HSCT; and (v) the patients' or the donors' age at HSCT.

Cytokines induced killer cells produced in good manufacturing practices conditions: identification of the most advantageous and safest expansion method in terms of viability, cellular growth and identity.

BACKGROUND: Cytokine-induced killer (CIK) cells are a very promising cell population raising growing interest in the field of cellular antitumor therapy. The aim of our study was to validate the most advantageous expansion method for this advanced therapy medicinal product (ATMP) and to translate it from preclinical field to good manufacturing practices (GMP). GMP ensures that ATMP are consistently produced and controlled to the quality standards required to their intended use. For this reason, the use of the xenogenic sera tended to be minimized by GMP for their high variability and the associated risk of transmitting infectious agents. RESULTS: We decided to replace Fetal Bovine Serum (FBS), largely used as medium supplement for CIKs expansion, with other culture media. Firstly, Human Serum (HS) and Human Pool Plasma (HPP) were tested as medium supplements giving not compliant results to acceptance criteria, established for CIKs, probably for the great batch to batch variability. Consequently, we decided to test three different serum free expansion media: X-VIVO 15, (largely used by other groups) and Tex Macs and Cell Genix GMP SCGM: two GMP manufactured media. We performed a validation consisting in three run-sand even if the small number of experiments didn't permit us to obtained statistical results we demonstrated that both X-VIVO 15 and Tex Macs fulfilled the quality standards in terms of cellular growth, viability and identity while Cell Genix GMP SCGM resulted not compliant as it caused some technical problems such as high mortality. CONCLUSION: In conclusion, these preclinical validation data lay the bases for a GMP-compliant process to improve the CIKs expansion method.

Fiber Optic Thermo-Hygrometers for Soil Moisture Monitoring.

This work deals with the fabrication, prototyping, and experimental validation of a fiber optic thermo-hygrometer-based soil moisture sensor, useful for rainfall-induced landslide prevention applications. In particular, we recently proposed a new generation of fiber Bragg grating (FBGs)-based soil moisture sensors for irrigation purposes. This device was realized by integrating, inside a customized aluminum protection package, a FBG thermo-hygrometer with a polymer micro-porous membrane. Here, we first verify the limitations, in terms of the volumetric water content (VWC) measuring range, of this first version of the soil moisture sensor for its exploitation in landslide prevention applications. Successively, we present the development, prototyping, and experimental validation of a novel, optimized version of a soil VWC sensor, still based on a FBG thermo-hygrometer, but able to reliably monitor, continuously and in real-time, VWC values up to 37% when buried in the soil.

A comprehensive characterization of mitochondrial DNA mutations in glioblastoma multiforme.

Glioblastoma multiforme (GBM) is the most malignant brain cancer in adults, with a poor prognosis, whose molecular stratification still represents a challenge in pathology and clinics. On the other hand, mitochondrial DNA (mtDNA) mutations have been found in most tumors as modifiers of the bioenergetics state, albeit in GBM a characterization of the mtDNA status is lacking to date. Here, a characterization of the burden of mtDNA mutations in GBM samples was performed. First, investigation of tumor-specific vs. non tumor-specific mutations was carried out with the MToolBox bioinformatics pipeline by analyzing 45 matched tumor/blood samples, from whole genome or whole exome sequencing datasets obtained from The Cancer Genome Atlas (TCGA) consortium. Additionally, the entire mtDNA sequence was obtained in a dataset of 104 fresh-frozen GBM samples. Mitochondrial mutations with potential pathogenic interest were prioritized based on heteroplasmic fraction, nucleotide variability, and in silico prediction of pathogenicity. A preliminary biochemical analysis of the activity of mitochondrial respiratory complexes was also performed on fresh-frozen GBM samples. Although a high number of mutations was detected, we report that the large majority of them does not pass the prioritization filters. Therefore, a relatively limited burden of pathogenic mutations is indeed carried by GBM, which did not appear to determine a general impairment of the respiratory chain. This article is part of a Directed Issue entitled: Energy Metabolism Disorders and Therapies.

[Ejaculatory system cysts: a case report]. / Raro caso di cisti del sistema eiaculatore.

INTRODUCTION: Male pelvic floor cysts are a rare clinical entity that include: Wolffian duct remnants, Müllerian duct remnants, cysts of the seminal vesicles, prostate and ejaculatory duct/vas deferens cysts.
 CASE REPORT: We report the clinical case of a 21-year-old male patient with a history of previous surgery in childhood and more precisely: partial colectomy for congenital megacolon, removal of dysplastic right kidney and subsequent surgical adhesiolysis for bowel obstruction.
 At 17, the patient was submitted to MRI for groin pain with an incidental finding of a cystic mass at the level of the right seminal vesicle. Consequently, a TUR-ED was performed at another urology unit, for a suspected seminal vesicle ectasia, without resolution of pain symptoms. The patient was referred to us for persistent genitourinary infections, ejaculation disorder and episodes of gross hematuria. An additional MRI confirmed the presence of a cystic mass of 5,5 cm with a suspected opening into prostatic urethra. Urethrocystoscopy and urethrocystography retrograde confirmed this anatomical communication. For the persistence of the symptoms we performed retropubic surgical exeresis of the mass, with a histopathological finding of benign cyst of the vas deferens.
 Two major postoperative complications were reported: a pelvic hematoma that required surgical exploration and a urinary extravasation at the level of prostatic urethra, which resolved with prolonged urethral catheterization.
 CONCLUSIONS: Male pelvic floor cysts are a rare disease with a complex clinical and therapeutic management. A correct diagnosis is based on clinical signs and symptoms together with imaging studies of the pelvic region. The high risk of erectile dysfunction and ejaculatory disorders correlated to a surgical approach, recommend a treatment of these lesions only for symptomatic cases.

[History of laser in BPH therapy]. / Storia del Laser nella terapia dell'IPB.

Laser technology has been used in the treatment of BPH for more than 15-20 years in order to challenge transurethral resection of the prostate. The aim of this review article is to analyze the evolution of laser in BPH therapy, from early coagulative techniques - progressively abandoned for their elevated postoperative morbidity and unfavorable outcomes - to the newer techniques of vaporization, resection and enucleation of the prostate. A better comprehension of tissue-laser interactions, the improvement of laser technology and a growing clinical experience have lead to the development of different laser systems (Holmium, KTP, Thulium laser) that challenge TURP. Today, HoLEP and, secondarily, PVP are the laser techniques supported by more clinical evidences and represent valid alternatives to TURP.

Inactivated human platelet lysate with psoralen: a new perspective for mesenchymal stromal cell production in Good Manufacturing Practice conditions.

BACKGROUND AIMS: Mesenchymal stromal cells (MSC) are ideal candidates for regenerative and immunomodulatory therapies. The use of xenogeneic protein-free Good Manufacturing Practice-compliant growth media is a prerequisite for clinical MSC isolation and expansion. Human platelet lysate (HPL) has been efficiently implemented into MSC clinical manufacturing as a substitute for fetal bovine serum (FBS). Because the use of human-derived blood materials alleviates immunologic risks but not the transmission of blood-borne viruses, the aim of our study was to test an even safer alternative than HPL to FBS: HPL subjected to pathogen inactivation by psoralen (iHPL). METHODS: Bone marrow samples were plated and expanded in α-minimum essential medium with 10% of three culture supplements: HPL, iHPL and FBS, at the same time. MSC morphology, growth and immunophenotype were analyzed at each passage. Karyotype, tumorigenicity and sterility were analyzed at the third passage. Statistical analyses were performed. RESULTS: The MSCs cultivated in the three different culture conditions showed no significant differences in terms of fibroblast colony-forming unit number, immunophenotype or in their multipotent capacity. Conversely, the HPL/iHPL-MSCs were smaller, more numerous, had a higher proliferative potential and showed a higher Oct-3/4 and NANOG protein expression than did FBS-MSCs. Although HPL/iHPL-MSCs exhibit characteristics that may be attributable to a higher primitive stemness than FBS-MSCs, no tumorigenic mutations or karyotype modifications were observed. CONCLUSIONS: We demonstrated that iHPL is safer than HPL and represents a good, Good Manufacturing Practice-compliant alternative to FBS for MSC clinical production that is even more advantageous in terms of cellular growth and stemness.

Validation of analytical methods in compliance with good manufacturing practice: a practical approach.

BACKGROUND: The quality and safety of cell therapy products must be maintained throughout their production and quality control cycle, ensuring their final use in the patient. We validated the Lymulus Amebocyte Lysate (LAL) test and immunophenotype according to International Conference on Harmonization Q2 Guidelines and the EU Pharmacopoeia, considering accuracy, precision, repeatability, linearity and range. METHODS: For the endotoxin test we used a kinetic chromogenic LAL test. As this is a limit test for the control of impurities, in compliance with International Conference on Harmonization Q2 Guidelines and the EU Pharmacopoeia, we evaluated the specificity and detection limit.For the immunophenotype test, an identity test, we evaluated specificity through the Fluorescence Minus One method and we repeated all experiments thrice to verify precision. The immunophenotype validation required a performance qualification of the flow cytometer using two types of standard beads which have to be used daily to check cytometer reproducibly set up. The results were compared together.Collected data were statistically analyzed calculating mean, standard deviation and coefficient of variation percentage (CV%). RESULTS: The LAL test is repeatable and specific. The spike recovery value of each sample was between 0.25 EU/ml and 1 EU/ml with a CV% < 10%. The correlation coefficient (≥ 0.980) and CV% (< 10%) of the standard curve tested in duplicate showed the test's linearity and a minimum detectable concentration value of 0.005 EU/ml.The immunophenotype method performed thrice on our cell therapy products is specific and repeatable as showed by CV% inter -experiment < 10%. CONCLUSIONS: Our data demonstrated that validated analytical procedures are suitable as quality controls for the batch release of cell therapy products.Our paper could offer an important contribution for the scientific community in the field of CTPs, above all to small Cell Factories such as ours, where it is not always possible to have CFR21 compliant software.

Analysis of different medulloblastoma histotypes by two-dimensional gel and MALDI-TOF.

OBJECTIVE: The purpose of this study is to detect different protein profiles in medulloblastoma (MDB) that may be clinically relevant and to check the correspondence of histological classification of MDB with proteomic profiles. MATERIALS AND METHODS: Surgical specimens, snap frozen at the time of neurosurgery, entered the proteomic study. Eight samples from patients (age range, 4 months-26 years) with different MDB histotypes (five classic, one desmoplastic/nodular, one with extensive nodularity, and one anaplastic) were analyzed by two-dimensional gel electrophoresis. One sample for each histotype was further characterized by matrix-assisted laser desorption/ionization time of flight mass spectrometry analysis. RESULTS: Eighty-six unique proteins were identified and compared to histology, with the determination of proteins expressed by single histotypes and of a smaller number of proteins shared by two or three histotypes. The sharp difference of protein expression was found to be in agreement with WHO histological classification, with the identification of type-specific proteins with limited overlapping between histotypes. CONCLUSION: Proteomic analysis confirmed and strengthened the difference between histotypes as biologically relevant. Cluster analysis enhanced the distance of extensive nodularity MDB from other histotypes. Possible innovative approaches to therapy may rely upon a proteomic-based classification of MDB tightly correlated to histology. The utility of snap freezing tumoral samples must be stressed and should become a mandatory task for pathologists.

Nanosized corners for trapping and detecting magnetic nanoparticles.

We present a device concept based on controlled micromagnetic configurations in a corner-shaped permalloy nanostructure terminated with two circular disks, specifically designed for the capture and detection of a small number of magnetic beads in suspension. A transverse head-to-head domain wall (TDW) placed at the corner of the structure plays the role of an attracting pole for magnetic beads. The TDW is annihilated in the terminating disks by applying an appropriate magnetic field, whose value is affected by the presence of beads chemically bound to the surface. In the case where the beads are not chemically bound to the surface, the annihilation of the TDW causes their release into the suspension. The variation of the voltage drop across the corner, due to the anisotropic magnetoresistance (AMR) while sweeping the magnetic field, is used to detect the presence of a chemically bound bead. The device response has been characterized by using both synthetic antiferromagnetic nanoparticles (disks of 70 nm diameter and 20 nm height) and magnetic nanobeads, for different thicknesses of the protective capping layer. We demonstrate the detection down to a single nanoparticle, therefore the device holds the potential for the localization and detection of small numbers of molecules immobilized on the particle's functionalized surface.