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Background: The renal biopsy represents a cornerstone in the definition of monoclonal gammopathy of renal significance (MGRS), helping in identifying patients with sub-detectable neoplastic clones (MGUS) that would deserve aggressive chemotherapies. However, the rising complexity of this onco-nephrology field is significantly challenging the daily work of nephrologists and nephropathologists, leading to the formation of ultra-specialized international centers with dedicated personnel/instrumentation and stressing the need for a better understanding of the underlying molecular landscape of these entities. Summary: In this setting, the application of proteomic techniques, some with in situ capabilities (e.g., MALDI-MS imaging), for the investigation of the most challenging MGRS is progressively shedding light on the pathobiology of these diseases, providing new insights in the diagnosis and prognosis of these cases. This transformation is further enhanced by the application of next-generation digital pathology platforms, leading to a significant improvement of the cultural background for physicians thanks to second opinions, database and atlas creation, enhancement of diagnostic reports, with obvious repercussions for patients both in terms of turnaround time and appropriateness. Key Messages: The present review is aimed at bridging the gap between clinical questions (i.e., a better characterization of MGRS) and the molecular landscape of onco-nephrology entities.

Chronical kidney disease (CKD) is a worldwide health problem with increasing incidence, where the major part accounts for chronic glomerulonephrites (GN). It is a group of diseases of various aetiology and multiform clinical course, having various prognosis which is often hardly predictable as well as existing prognostic markers are not always certain. There is an urging need of new reliable and specific prognostic and therapeutic markers research. A modern proteomic technology - Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) has been employed in many GN studies showing promising results. We aimed to study several forms of primary and secondary glomerulonephrites (membranous nephropathy, IgA nephropathy, diabetic nephropathy) to enlighten possible molecular alterations significant of diseases’ progression. The studies were performed on renal tissue biopsies, analysing them with high spatial resolution MALDI-MSI to get better visualisation of signals’ co-localisation on tissue. We performed a comparison of molecular profiles of various forms of GN. As a result, we were able to generate and distinguish specific tryptic peptides profiles of different cell regions (tubules, glomeruli, interstitium, connective tissue) and detect proteins with an altered intensity, implicated in inflammatory and healing pathways. MALDI-MSI, being able to define renal structures, could provide additional diagnostic and prognostic information. Generation of collective diagnostic panels may fulfil our pathogenesis understanding and assist clinical prognostic assessment., Chronical kidney disease (CKD) is a worldwide health problem with increasing incidence, where the major part accounts for chronic glomerulonephrites (GN). It is a group of diseases of various aetiology and multiform clinical course, having various prognosis which is often hardly predictable as well as existing prognostic markers are not always certain. There is an urging need of new reliable and specific prognostic and therapeutic markers research. A modern proteomic technology - Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) has been employed in many GN studies showing promising results. We aimed to study several forms of primary and secondary glomerulonephrites (membranous nephropathy, IgA nephropathy, diabetic nephropathy) to enlighten possible molecular alterations significant of diseases’ progression. The studies were performed on renal tissue biopsies, analysing them with high spatial resolution MALDI-MSI to get better visualisation of signals’ co-localisation on tissue. We performed a comparison of molecular profiles of various forms of GN. As a result, we were able to generate and distinguish specific tryptic peptides profiles of different cell regions (tubules, glomeruli, interstitium, connective tissue) and detect proteins with an altered intensity, implicated in inflammatory and healing pathways. MALDI-MSI, being able to define renal structures, could provide additional diagnostic and prognostic information. Generation of collective diagnostic panels may fulfil our pathogenesis understanding and assist clinical prognostic assessment.

Purpose The use of indoor living plants for enhancement of indoor relative humidity and the general environment of a large, modern, open plan office building; are studied using a mixed-methods paradigm. Design/methodology/approach The quantitative element involved designated experimental and control zones within the building, selected using orientation, user density, and users' work roles criteria. For a period of six months, relative humidity was monitored using data loggers at 30-minute intervals and volatile organic compounds (VOCs) were measured using air sampling. Qualitative ‘perception data’ of the building’s users, were collected via a structured questionnaire survey among both experimental and control zones. Findings Study findings include that living plants did not achieve the positive effect on relative humidity predicted by (a-priori) theoretical calculations; and that building users’ perceived improvements to indoor relative humidity, temperature, and background noise levels, were minimal. The strongest perceived improvement was for work environment aesthetics. Findings demonstrate the potential of indoor plants to reduce carbon emissions of the [as] built environment, through elimination or reduction of energy use and capital-intensive humidification air-conditioning systems. Originality/value The study’s practical value lies in its unique application of (mainly laboratory-derived) existing theory in a real-life work environment.

European railways have been shaped by multiple reforms since the mid-1990s, covering industry structure, market opening and economic regulation. However, the literature has given little attention to the latter; namely the evolution and impacts of regulatory reforms amongst Europe's railways. This paper fills this gap by providing an up-to-date, bottom-up assessment of current rail regulatory practice in Europe. We develop a survey of economic regulators across Europe, thus complementing top-down studies of the impact of economic regulation by enabling a richer insight into regulatory activity and its impacts. The questionnaire is based on a review of the literature on ideal regulatory characteristics across multiple industries. Our results show that European rail regulators, in general, exhibit many of the features of ideal regulation; in particular around key features such as independence, resourcing, longevity and expertise, transparency and in turn stability and predictability. However, we find that rail regulatory bodies could take a more proactive role in shaping track access charges, given their importance in respect of efficient use of the network and maintaining non-discriminatory access. Importantly, there is also scope for regulators to play a greater role in regulating the efficiency and quality of infrastructure managers, and potentially becoming more involved in the designing stages of passenger market opening as it emerges; and these changes could deliver substantial beneficial impacts for rail users and funders across Europe.

Le glomerulonefriti idiopatiche (GN), tra cui le nefropatie membranose (MN), le glomerulosclerosi segmentarie e focali (FSGS) e la nefropatia da IgA (IgAN), rappresentano le più frequenti patologie glomerulari primarie e sono spesso causa insufficienza renale terminale (ESRD). Senza una corretta diagnosi e la consegnuente scelta di un opportuno trattamento, la patologia presenta elevate probabilità di progredire verso uno stadio terminale, obbligando il paziente a sottoporsi a cicli di dialisi o al trapianto di rene. Ad oggi, la biopsia renale rimane la tecnica di elezione per la diagnosi delle GN idiopatiche, ma, considerando i problemi legati all’invasività di questa tecnica e la sua non perfetta capacità di garantire sempre una corretta diagnosi, biomarcatori diagnostici e prognostici, utilizzabili con tecniche meno invasive, stanno sempre di più diventando indispensabili. La spettrometria di massa MALDI-MS Imaging è una tecnologia avanzata che permette un’integrazione tra le informazioni molecolari ottenute da studi di proteomica tissutale e le caratteristiche morfologiche. La possibilità di correlare la distribuzione spaziale di una vasta gamma di analiti direttamente in situ con le informazioni istologiche rende questa tecnica uno strumento ottimale per la ricerca di nuovi biomarcatori, non solo a livello di alterazioni molecolari ma anche di variazioni nella posizione nel tessuto. Inoltre, per quanto riguarda il campo della nefrologia, il MALDI-MS Imaging permette potenzialmente di ottenere la firma proteomica dei glomeruli e dei tubuli, sia dei pazienti affetti da patologia che dei pazienti controllo. Questa tesi si focalizza sull’utilizzo del MALDI-MS Imaging per l’analisi di biopsie renali, sia di tessuti freschi/congelati che tessuti fissati in formalina ed inclusi in paraffina (FFPE), con lo scopo di identificare marker diagnostici e prognostici associabili alle più comuni forme di glomerulonefriti. In particolare, il MALDI-MS Imaging è stato ap, Idiopathic glomerulonephritis (GN), such as membranous nephropathy (MN), focal segmental glomerulosclerosis (FSGS) and IgA nephropathy (IgAN) represent the most frequent primary Glomerular Kidney Diseases (GKDs) worldwide and are a common cause of end-stage renal disease (ESRD). Without the correct diagnosis and, ultimately, the correct selection of therapeutic treatment, the patient has a higher probability of progressing to ESRD and requiring dialysis or transplantation. Although the renal biopsy currently remains the gold standard for the routine diagnosis of idiopathic GN, the invasiveness and difficulty of diagnoses related with this procedure means that there is a strong need for the detection of diagnostic and prognostic biomarkers that can be translated into less invasive diagnostic tools. Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) is a unique proteomic technology that explores the spatial distribution of biomolecules directly in situ, thus integrating molecular and morphological information. The possibility to correlate distribution maps of multiple analyses with histological features makes it an ideal research tool to discover new diagnostic and prognostic markers. This modern proteomic technique could represent an invaluable tool in the field of nephropathology, most specifically in the study of primary glomerulonephritis (GNs), due to its potential to obtain proteomic signatures from pathological glomeruli and tubuli. The body of work enclosed in this thesis follows the application of MALDI-MSI to fresh-frozen and formalin-fixed paraffin-embedded (FFPE) renal biopsies in order to detect diagnostic and prognostic molecular markers associated with the most common forms of glomerulonephritis. More specifically, MALDI-MSI was applied to fresh frozen bioptic renal tissue from patients with a histological diagnosis of FSGS (n=6), IgAN, (n=6) and MN (n=7), and from controls (n=4) in order to detect specific molecular sig

Fine needle aspiration (FNA) biopsies are the current gold-standard for the preoperative evaluation of thyroid nodules. However, a significant number of them (15–30%) are unable to be affirmatively diagnosed and are given an “indeterminate for malignancy” final report, meaning that the malignant nature of the thyroid nodule remains unknown and the recommended therapeutic approach is total thyroidectomy. Furthermore, cytomorphological evaluation of biopsies taken post-surgery indicates that approximately 80% of nodules within this group of patients are in fact benign, and the total thyroidectomy unwarranted. Therefore, the identification of new possible diagnostic targets that can assist in the preoperative diagnosis of thyroid tumors and reduce the number of unnecessary thyroidectomies is imperative. Matrix-Assisted Laser Desorption/Ionization (MALDI)—Mass Spectrometry Imaging (MSI) has the ability to provide very precise and localized information regarding protein expression in cytological specimens. This enables the detection of cell subpopulations based on their different protein profiles, even within regions that are indistinguishable at the microscopic level, and the feasibility of this approach to investigate FNA specimens has already been highlighted in a number of studies. Here, an overview about the sample preparation procedure for the MALDI-MSI analysis of ex vivo FNA biopsies is provided, highlighting how molecular imaging can be combined with traditional histology to generate protein signatures of the different thyroid lesions, and, ultimately, build classification models that can be potentially used to classify benign and malignant thyroid nodules from a molecular standpoint.

Glomerulonephritis (GNs) are one of the most frequent causes of chronic kidney disease (CKD), a renal condition that often leads to end-stage renal failure, and a careful assessment of these diseases is essential for prognostic and therapeutic purposes. The application of MALDI-MSI directly on bioptic renal tissue represents a new stimulating perspective and facilitates the detection of specific proteomic indicators that are directly correlated with the pathological alterations that occur within the glomeruli during the development of glomerulonephritis. Here, we describe the standard workflow for the MALDI-MSI analysis of clinical fresh-frozen and FFPE renal biopsies and highlight how the obtained molecular information, when combined with histology, can be used to detect specific protein markers of GNs.

BACKGROUND: Due to the recent proposal of the non-invasive follicular thyroid neoplasm with papillary-like nuclear feature (NIFTP) category, the authors analyse the state of the art in the challenging diagnosis of follicular thyroid neoplasms in routine practice. METHODS AND RESULTS: A consecutive series of 200 histological diagnoses, with complete cytological correlation, was analysed following the introduction of the NIFTP definition. The study was conducted in a general hospital with a high prevalence of thyroid benign nodules that accounted for approximately 60% of surgically-treated nodules. The significant incidence of the new NIFTP category was 7%. Concurrently, a gradual decrease of the follicular variant of papillary thyroid carcinoma (fvPTC) was observed (3.5%). When evaluating the FNA biopsies within the NIFTP group, despite the systematic evaluation of nuclear crowding, enlargement, irregularities and clearing, the final cytological class was often indeterminate for malignancy (Thy3/III-IV, 71%). At histology, the application of the semiquantitative NIFTP score for the evaluation of the PTC-like nuclear features was able to discriminate benign lesions (score 0/1) from fvPTC (score 2/3). A certain degree of overlapping still persisted between NIFTP and fvPTC (score 2) or between NIFTP and benign lesions (score 1). CONCLUSIONS: In the routine evaluation of FNA biopsies, the presence of subtle and questionable PTC-like nuclear features still remains a controversial aspect of the diagnostic workflow. Given that the NIFTP category was introduced to stratify the low-risk group of thyroid tumours more precisely, pathologists should force themselves to apply the nuclear score rigorously and to classify cases assigned a score of 1 as benign proliferations

The current study proposes the successful use of a mass spectrometry-imaging technology that explores the composition of biomolecules and their spatial distribution directly on-tissue to differentially classify benign and malignant cases, as well as different histotypes. To identify new specific markers, we investigated with this technology a wide histological Tissue Microarray (TMA)-based thyroid lesion series. Results showed specific protein signatures for malignant and benign specimens and allowed to build clusters comprising several proteins with discriminant capabilities. Among them, FINC, ACTB1, LMNA, HSP7C and KAD1 were identified by LC-ESI-MS/MS and found up-expressed in malignant lesions. These findings represent the opening of further investigations for their translation into clinical practice, e.g. for setting up new immunohistochemical stainings, and for a better understanding of thyroid lesions

The purpose of this paper is to investigate the use of living plants in enhancing the indoor air quality (IAQ) and the general indoor environment within a large modern open-plan office building with a central atrium design and a building management system (BMS) in place. Poor indoor air quality was measured within the building, primarily due to the low relative humidity during the winter months. Previous literature suggests that the incorporation of plants in buildings helps to regulate relative humidity whilst also bringing perceptual benefits and potentially reducing short-term sickness absence. The investigation was developed through quantitative and qualitative data. The quantitative element involved the use of experimental and control zones within the building, selected on the basis of orientation, user density and users' work roles. Plants were selected based on the transpiration rates of various commercially-available species. Relative humidity was continuously monitored using data loggers with half-hourly logging intervals for a duration of six months. Carbon dioxide gas concentration was measured using a dedicated hand-held sensor. Qualitative user perception data was gathered through the use of a structured questionnaire distributed to staff members working in each of the experimental and control zones. Initial findings suggest that the plants have not instigated the positive effects on IAQ that were expected. The recorded data on relative humidity displayed only non-significant variations between the experimental and control zones. These findings are attributed, in part, to the atrium design, which results in a substantial volume of air within the building, leading to cross-contamination and excessive dilution of the introduced humidity as a result of plant transpiration. The study extends the previous, mainly laboratory-based, investigations to a real working environment. However, this introduces a range of other experimental factors, thus impacting the

Glomerulonephritis (GN) is one of the most common origins of chronic kidney disease and its careful evaluation is crucial for prognostic and therapeutic purposes, with the renal biopsy still playing a central role for the diagnosis. However, due to its invasiveness, it is not devoid of complications and many investigations have focused on identifying biomarkers for chronic kidney diseases using less-invasive and easy-to-collect samples, such as urine and blood. In this context, proteomics has played a crucial role in determining the molecular changes related to disease progression and early pathological glomerular modifications. Here, we report a review of selected literature for each GN, based on selected works published in the last 10 years, showing how these approaches have generated clinically relevant findings in the study of glomerulonephritis. We also describe several proteomic strategies, highlighting their technical advantages and limitations, future perspectives for proteomic applications in the study of GNs, and their possible application in routine practice.

Introduction: Renal cell carcinoma (RCC) is the most fatal of the common urologic cancers, with approximately 35% of patients dying within 5 years following diagnosis. Therefore, there is a need for non-invasive markers that are capable of detecting and determining the severity of small renal masses at an early stage in order to tailor treatment and follow-up. Proteomic studies have proved to be very useful in the study of tumors. Areas covered: In this review, we will detail the current knowledge obtained by the different proteomic approaches, focusing on MS-based strategies, used to investigate RCC biology in order to identify diagnostic, prognostic and predictive biomarkers on tissue, cultured cells and biological fluids. Expert commentary: Currently, no reliable biomarkers or targets for RCC have been translated into the clinical setting. Moreover, despite the efforts of proteomics and other-omics disciplines, only a small number of them have been observed as shared targets between the different analytical platforms and biological specimens. The difficulty to define a specific molecular pattern for RCC and its subtypes highlights a peculiar profile and a heterogeneity that must be taken into account in future studies.

Introduction: Despite the unquestionable advantages of Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry Imaging in visualizing the spatial distribution and the relative abundance of biomolecules directly on-tissue, the yielded data is complex and high dimensional. Therefore, analysis and interpretation of this huge amount of information is mathematically, statistically and computationally challenging. Areas covered: This article reviews some of the challenges in data elaboration with particular emphasis on machine learning techniques employed in clinical applications, and can be useful in general as an entry point for those who want to study the computational aspects. Several characteristics of data processing are described, enlightening advantages and disadvantages. Different approaches for data elaboration focused on clinical applications are also provided. Practical tutorial based upon Orange Canvas and Weka software is included, helping familiarization with the data processing. Expert commentary: Recently, MALDI-MSI has gained considerable attention and has been employed for research and diagnostic purposes, with successful results. Data dimensionality constitutes an important issue and statistical methods for information-preserving data reduction represent one of the most challenging aspects. The most common data reduction methods are characterized by collecting independent observations into a single table. However, the incorporation of relational information can improve the discriminatory capability of the data.

Objective: This paper analyzes a series of ultrasound (US)-guided orbital fine needle aspirations (FNAs) which provide diagnostic information that cytopathologists approaching orbital lesions for the first time can find useful and underlines the importance of teamwork. Study Design: The investigators retrospectively obtained data from 24 consecutive orbital FNAs. For all patients, a complete clinicoradiological database was created. FNAs were performed under US guidance with 25-gauge needles and an aspiration biopsy syringe gun, and sent to the Department of Pathology for examination and data management. Results: The mean age of the patients was 54 years. Imaging studies included US, magnetic resonance imaging and computed tomography scans; 9 lesions involved the right orbit and 15 the left orbit. The mean lesion size was 23.6 ± 7.2 mm. After microscopic examination, 7 smears were labeled as 'nondiagnostic', while in 17 cases a definitive diagnosis was proposed, which always proved to be correct (70.8%, specificity = 100%). Conclusions: The investigators believe that FNA biopsy of orbital masses is a necessary step; its weaknesses lie in the particularly delicate site of sampling and the extreme heterogeneity of lesions. Nevertheless, when orbital FNA is performed within a well-coordinated multidisciplinary team, it is a powerful tool that can be used to define the most appropriate management of these patients.

Background The glomerulocentric International Society of Nephrology/Renal Pathology Society (ISN/RPS) classification is the gold standard for the evaluation of lupus nephritis, while tubulointerstitial (TIN) parameters are often under-recognized in pathological reports. Methods Renal biopsies from 142 patients who underwent repeat biopsy (RB) were evaluated for the following histological parameters: (i) inflammatory interstitial infiltrates; (ii) interstitial fibrosis; (iii) tubulitis; and (iv) tubular atrophy. The inter-relationships between the four TIN variables were explored by multivariate analysis. A linear mixed model was used to investigate the potential impact of TIN variables on EGFR and proteinuria at the two biopsy occasions. Results The study showed that moderate-severe lesions were not so frequent at the reference biopsy, but more extensively represented upon RB. A strong association was found between the two inflammatory indices and between those related to chronic damage, while the relationship with the ISN/RPS classification was present at RB. If class IV-G was the most related with TIN (especially at RB), the existence of primary TIN in class II patients was also confirmed. Finally, our results support the hypothesis that tubulitis is an independent predictive factor for EGFR. Conclusions We recommend that the standard histological evaluation of SLE nephritis also includes TIN features.

Background: Several promising biomarkers have been found for RCC, but none of them has been used in clinical practice for predicting tumour progression. The most widely used features for predicting tumour aggressiveness still remain the cancer stage, size and grade. Therefore, the aim of our study is to investigate the urinary peptidome to search and identify peptides whose concentrations in urine are linked to tumour growth measure and clinical data. Methods: A proteomic approach applied to ccRCC urinary peptidome (n = 117) based on prefractionation with activated magnetic beads followed by MALDI-TOF profiling was used. A systematic correlation study was performed on urinary peptide profiles obtained from MS analysis. Peptide identity was obtained by LC-ESI-MS/MS. Results: Fifteen, twenty-six and five peptides showed a statistically significant alteration of their urinary concentration according to tumour size, pT and grade, respectively. Furthermore, 15 and 9 signals were observed to have urinary levels statistically modified in patients at different pT or grade values, even at very early stages. Among them, C1RL, A1AGx, ZAG2G, PGBM, MMP23, GP162, ADA19, G3P, RSPH3, DREB, NOTC2 SAFB2 and CC168 were identified. Conclusions: We identified several peptides whose urinary abundance varied according to tumour size, stage and grade. Among them, several play a possible role in tumorigenesis, progression and aggressiveness. These results could be a useful starting point for future studies aimed at verifying their possible use in the managements of RCC patients.

Recent advancements in Matrix Assisted Laser Desorption/Ionisation (MALDI) Mass Spectrometry Imaging (MSI) technology have enabled the analysis of formalin-fixed paraffin-embedded (FFPE) tissue samples, unlocking a wealth of new proteomic information and facilitating the possibility of performing studies with higher statistical power as well as multi-centric collaborations within the field of proteomics research. However, current methods used to analyse these specimens are often time-consuming and they need to be modified when applied to human tissues of different origin. Here we present a reproducible and time-effective method that could address these aforementioned issues and widen the applicability of this technology to a number of challenging tissue types. Additionally, tissue molecular images show high spatial resolution and a strong correlation with the morphological features, enabling the identification of tissue morphology using statistically derived visualisation, without any prior knowledge.

Objective: Matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) is a unique proteomic technology that explores the spatial distribution of biomolecules directly in situ, thus integrating molecular and morphological information. The possibility of correlating distribution maps of multiple analyses with cytological features makes it an ideal research tool for discovering new diagnostic markers. A previous study showed that MALDI-IMS could help discrimination between different types of thyroid lesions, especially papillary thyroid carcinoma (PTC); the present feasibility study on ex vivo fine needle aspiration (FNA) smears describes its potential in detecting new proteomic targets of other thyroid lesions (follicular lesions, medullary carcinoma). Methods: MALDI-IMS was conducted on ex vivo FNAs obtained from surgical specimens and corresponding in vivo samples. Differences between proteomic profiles of different thyroid lesions were compared. Results: Comparing the protein profiles of hyperplastic nodules obtained from three different patients with each other, and with a new PTC, showed a high degree of concordance, indicating good reproducibility of the IMS technology on cytological samples, suggesting its potential as a tool for biomarker discovery. Furthermore, comparison of the average proteomic profiles of hyperplastic nodules with a Hürthle cell adenoma revealed significant differences, underlying the capability of MALDI-IMS to distinguish between different thyroid lesions. Finally, the proteomic profile of medullary thyroid carcinoma was also characterized. Conclusions: Our results confirmed the possible role of MALDI-IMS in the search for diagnostic targets of PTC and follicular lesions, which could be applied in larger trials aimed at the identification of proteins, convertible to cost-effective diagnostic tools such as immunohistochemistry. These tests could be used to analyse in vivo cytological smears, improving the preoperat