"Mimics" of Injuries from Child Abuse: Case Series and Review of the Literature.

The phenomenon of child abuse/maltreatment is underestimated and often represents a difficult challenge for healthcare professionals and forensic pathologists who must proceed with the differential diagnosis with accidental or self-induced events, or with lesions due to pathologies that overlap with that of mistreatment, defined as "Mimics". This study presents a case series with the aim of discussing lesions that may mimic signs of physical abuse in children but are due to a different etiology to raise awareness and train healthcare professionals and forensic pathologists on possible confounding factors in order to avoid diagnostic errors. Six cases of "Mimics" out of 418 cases of suspected mistreatment (1.43% of cases) were identified, presenting skin lesions initially classified as injuries of abuse due to their location and type and, in particular, sexual abuse for three cases. Then, the lesions and the subjects, in particular the anamnestic history, were examined by a multidisciplinary team and the diagnosis of genital lichen sclerosus et atrophicus in three cases, and the results of popular healing techniques (i.e., "cupping") in the other three cases were ascertained. These situations require specific skills and a forensic background from healthcare professionals to conduct a correct differential diagnosis and the intervention of a multidisciplinary team to investigate every possible pathology or alternative therapeutic practice that could simulate child abuse. In particular, when "mimics" are due to alternative medicine, it should not strictly be considered child abuse, but professionals must be aware of the hypothesis of mistreatment in case of non-medical indication or potential personal injuries from other crimes, such as illegal practice of the medicine. This awareness is also crucial to direct the child toward appropriate medical care, and it is essential to recognize that these conditions can coexist within the same clinical presentation.

Comparison of CryoMACS Freezing Bags with Maco Biotech Freezing-Ethinyl Vinyl Acetate Bags for Hematopoietic Progenitor Cells Cryopreservation Using a CD34+-Enriched Product.

Background: Hematopoietic progenitor cells (HPCs) cryopreservation have applications, especially in the autologous setting, allowing therapeutic use several years after collection. Cryopreservation aims to preserve the therapeutic properties of HPCs, and successful cryopreservation depends on several factors such as preservation procedures, biopreservation media, freezing rates, and thawing procedures. In this context, the choice of the freezing bag is critical as it provides mechanical protection during the freezing process. Since Maco Biotech Freezing-ethinyl vinyl acetate (EVA) Bags® are no longer available in our country, a comparative study was developed to verify bioequivalence with the Miltenyi CryoMACS® freezing bag. Methods: In this study, a CD34+-enriched product was used to better reproduce HPC apheresis. Freezing bags were filled with the same volume, cryopreserved with controlled rate freezing, and stored in the vapor phase of liquid nitrogen for at least 6 months. After thawing, all bags were tested for integrity and sterility using a microbial challenge. In addition, a comparison was developed by evaluating recovery of white blood cells, mononuclear cells, lymphocytes, and CD34+ cells. Results: No significant differences between the two manufacturers' bags have been observed in terms of the evaluated parameters. Data were confirmed, even comparing bags according to filling volume. Data presented in this study support the conclusion that CryoMACS freezing bags are bioequivalent to Maco Biotech Freezing-EVA Bags for HPC cryopreservation.

An alternative procedure to leukapheresis for peripheral hematopoietic progenitor cell collection in very-low-weight children: A single pediatric center experience.

BACKGROUND: PBSC collection using a blood cell separator in very low weight patients can be frequently complicated by severe adverse effects and technical difficulties. MATERIAL AND METHODS: From March 2013 to January 2017, 14 PBSC collections were performed in 12 children weighing less than 10 kg, affected by different solid tumours. PBSC collection was performed with a "homemade" aseptically assembled circuit. The circuit is composed by a 150 mL collection bag connected with a 4 stopcock ramp, perfused with ACD. This circuit allows collection of a specific total blood amount from CVC, depending on CD34+ /kg target. RESULTS: Mean CD34+ cell performance per collection was 9.3 × 106 /kg. Tolerance to the procedure was very good as none of the patients experienced complications, with the exception of a patient who showed mild cyanosis and pallor after collection. Moreover, no bleeding or thrombotic complications have been observed. To date, 16 PBSC reinfusions have been performed in 7 children with a mean CD34+ cells viability of 98.1% ± 2.7 and mean WBC viability of 57% ± 10. Cell recovery after thawing was 87% ± 10.8. A rapid graft intake for both neutrophils and platelets, between day 7 and 20 after reinfusion was observed. DISCUSSION: The procedure of total blood collection without the use of a cell separator is feasible and allows a good PBSC collection without significant side effects in very low-weight children. Moreover, this method could represent a valid and safe alternative to leukapheresis in patients where classic procedure could be difficult to apply.

Human platelet lysate in mesenchymal stromal cell expansion according to a GMP grade protocol: a cell factory experience.

BACKGROUND: The use of platelet lysate (PL) for the ex-vivo expansion of mesenchymal stromal/stem cells (MSCs) was initially proposed by Doucet et al. in 2005, as an alternative to animal serum. Moreover, regulatory authorities discourage the use of fetal bovine serum (FBS) or other animal derivatives, to avoid risk of zoonoses and xenogeneic immune reactions. Even if many studies investigated PL composition, there still are some open issues related to its use in ex-vivo MSC expansion, especially according to good manufacturing practice (GMP) grade protocols. METHODS: As an authorized cell factory, we report our experience using standardized PL produced by Azienda Ospedaliero Universitaria Meyer Transfusion Service for MSC expansion according to a GMP grade clinical protocol. As suggested by other authors, we performed an in-vitro test on MSCs versus MSCs cultured with FBS that still represents the best way to test PL batches. We compared 12 MSC batches cultured with DMEM 5% PL with similar batches cultured with DMEM 10% FBS, focusing on the MSC proliferation rate, MSC surface marker expression, MSC immunomodulatory and differentiation potential, and finally MSC relative telomere length. RESULTS: Results confirmed the literature data as PL increases cell proliferation without affecting the MSC immunophenotype, immunomodulatory potential, differentiation potential and relative telomere length. CONCLUSIONS: PL can be considered a safe alternative to FBS for ex-vivo expansion of MSC according to a GMP grade protocol. Our experience confirms the literature data: a large number of MSCs for clinical applications can be obtained by expansion with PL, without affecting the MSC main features. Our experience underlines the benefits of a close collaboration between the PL producers (transfusion service) and the end users (cell factory) in a synergy of skills and experiences that can lead to standardized PL production.

Bone marrow-derived mesenchymal stem cells promote invasiveness and transendothelial migration of osteosarcoma cells via a mesenchymal to amoeboid transition.

There is growing evidence to suggest that bone marrow-derived mesenchymal stem cells (BM-MSCs) are key players in tumour stroma. Here, we investigated the cross-talk between BM-MSCs and osteosarcoma (OS) cells. We revealed a strong tropism of BM-MSCs towards these tumour cells and identified monocyte chemoattractant protein (MCP)-1, growth-regulated oncogene (GRO)-α and transforming growth factor (TGF)-ß1 as pivotal factors for BM-MSC chemotaxis. Once in contact with OS cells, BM-MSCs trans-differentiate into cancer-associated fibroblasts, further increasing MCP-1, GRO-α, interleukin (IL)-6 and IL-8 levels in the tumour microenvironment. These cytokines promote mesenchymal to amoeboid transition (MAT), driven by activation of the small GTPase RhoA, in OS cells, as illustrated by the in vitro assay and live imaging. The outcome is a significant increase of aggressiveness in OS cells in terms of motility, invasiveness and transendothelial migration. In keeping with their enhanced transendothelial migration abilities, OS cells stimulated by BM-MSCs also sustain migration, invasion and formation of the in vitro capillary network of endothelial cells. Thus, BM-MSC recruitment to the OS site and the consequent cytokine-induced MAT are crucial events in OS malignancy.

Mesenchymal Stem Cells are Recruited and Activated into Carcinoma-Associated Fibroblasts by Prostate Cancer Microenvironment-Derived TGF-ß1.

Tumor stromal cells can supply appropriate signals that may develop aggressive phenotypes of carcinoma cells and establish a complex scenario which culminates in metastasis. Recent works proposed that bone marrow-derived mesenchymal stem cells (MSC) are recruited to primary tumors. However, the exact functions of these cells in the tumor microenvironment are not well characterized, as it is reported that MSC can either promote or inhibit tumor progression. In the present study, we aim at investigating the signaling molecules which regulate the interplay between MSC, prostate carcinoma (PCa) cells and two important cellular types constituting the tumor-associated stroma, macrophages and fibroblasts, during their progression toward malignancy. We identified TGF-ß1 as a crucial molecule able to attract MSC recruitment both to PCa cells as well as to tumor stroma components. Moreover, PCa- and tumor stroma-secreted TGF-ß1 is important to induce MSC transdifferentiation into carcinoma-associated fibroblast (CAF)-like cells. Consequently, the CAF-like phenotype acquired by MSC is central to promote tumor progression related effects. Thus, tumor-educated MSC enhance PCa invasiveness compared to nonactivated MSC. Additionally, differing from normal MSC, CAF-like MSC perform vascular mimicry and recruit monocytes, which can be further polarized to M2 macrophages within the PCa environment. Our findings indicate a prominent role for TGF-ß1 in MSC mobilization and activation strengthened by the fact that the blockade of TGF-ß1 signaling impairs MSC promotion of PCa progression. Stem Cells 2016;34:2536-2547.

Tumor microenvironment: bone marrow-mesenchymal stem cells as key players.

Tumor progression is a multistep phenomenon in which tumor-associated stromal cells perform an intricate cross-talk with tumor cells, supplying appropriate signals that may promote tumor aggressiveness. Among several cell types that constitute the tumor stroma, the discovery that bone marrow-derived mesenchymal stem cells (BM-MSC) have a strong tropism for tumors has achieved notoriety in recent years. Not only are the BM-MSC recruited, but they can also engraft at tumor sites and transdifferentiate into cells such as activated fibroblasts, perivascular cells and macrophages, which will perform a key role in tumor progression. Whether the BM-MSC and their derived cells promote or suppress the tumor progression is a controversial issue. Recently, it has been proposed that proinflammatory stimuli can be decisive in driving BM-MSC polarization into cells with either tumor-supportive or tumor-repressive phenotypes (MSC1/MSC2). These considerations are extremely important both to an understanding of tumor biology and to the putative use of BM-MSC as "magic bullets" against tumors. In this review, we discuss the role of BM-MSC in many steps in tumor progression, focusing on the factors that attract BM-MSC to tumors, BM-MSC differentiation ability, the role of BM-MSC in tumor support or inhibition, the immunomodulation promoted by BM-MSC and metastatic niche formation by these cells.