Guidelines for minimal information on cellular senescence experimentation in vivo.

Cellular senescence is a cell fate triggered in response to stress and is characterized by stable cell-cycle arrest and a hypersecretory state. It has diverse biological roles, ranging from tissue repair to chronic disease. The development of new tools to study senescence in vivo has paved the way for uncovering its physiological and pathological roles and testing senescent cells as a therapeutic target. However, the lack of specific and broadly applicable markers makes it difficult to identify and characterize senescent cells in tissues and living organisms. To address this, we provide practical guidelines called "minimum information for cellular senescence experimentation in vivo" (MICSE). It presents an overview of senescence markers in rodent tissues, transgenic models, non-mammalian systems, human tissues, and tumors and their use in the identification and specification of senescent cells. These guidelines provide a uniform, state-of-the-art, and accessible toolset to improve our understanding of cellular senescence in vivo.

Potency assay to predict the anti-inflammatory capacity of a cell therapy product for macrophage-driven diseases: overcoming the challenges of assay development and validation.

BACKGROUND: Given the high level of product complexity and limited regulatory guidance, designing and implementing appropriate potency assays is often the most challenging part of establishing a quality control testing matrix for a cell-based medicinal product. Among the most elusive tasks are the selection of suitable read-out parameters, the development of assay designs that most closely model the pathophysiological conditions, and the validation of the methods. Here we describe these challenges and how they were addressed in developing an assay that measures the anti-inflammatory potency of mesenchymal stromal cells (MSCs) in an M1 macrophage-dominated inflammatory environment. METHODS: An in vitro inflammation model was established by coculturing skin-derived ABCB5+ MSCs with THP-1 monocyte-derived M1-polarized macrophages. Readout was the amount of interleukin 1 receptor antagonist (IL-1RA) secreted by the MSCs in the coculture, measured by an enzyme-linked immunosorbent assay. RESULTS: IL-1RA was quantified with guideline-concordant selectivity, accuracy and precision over a relevant concentration range. Consistent induction of the macrophage markers CD36 and CD80 indicated successful macrophage differentiation and M1 polarization of THP-1 cells, which was functionally confirmed by release of proinflammatory tumor necrosis factor α. Testing a wide range of MSC/macrophage ratios revealed the optimal ratio for near-maximal stimulation of MSCs to secrete IL-1RA, providing absolute maximum levels per individual MSC that can be used for future comparison with clinical efficacy. Batch release testing of 71 consecutively manufactured MSC batches showed a low overall failure rate and a high comparability between donors. CONCLUSIONS: We describe the systematic development and validation of a therapeutically relevant, straightforward, robust and reproducible potency assay to measure the immunomodulatory capacity of MSCs in M1 macrophage-driven inflammation. The insights into the challenges and how they were addressed may also be helpful to developers of potency assays related to other cellular functions and clinical indications.

The Usefulness of Ultrasonography for Supporting the Differentiation, Diagnosis, and Treatment of Atypical Fibroxanthoma and Pleomorphic Dermal Sarcoma.

Atypical fibroxanthoma (AFX) and pleomorphic dermal sarcoma (PDS) are rare histomorphological variants of a disease spectrum. After ruling out other tumor entities by immunohistochemistry, PDS can be differentiated from AFX by infiltration into the subcutis, while AFX remains confined to the dermis. The therapeutic approach is more aggressive in PDS as it can potentially metastasize. We assessed the usefulness of preoperative sonography in differentiating between the two tumor entities by identifying a potential subcutaneous infiltration. In our patients (n = 13), preoperative sonography identified and differentiated AFX and PDS with 100% accuracy and even changed the initial histological suspicion of AFX to PDS in 3 cases (23%), which was confirmed after tumor resection. Preoperative sonography of these tumors could strengthen the clinical diagnosis, avoid a delay in therapy initiation and improve patient counseling. While for AFX, micrographic-controlled surgery suffices, for PDS, resection with 2 cm safety margins and lymph node sonography to rule out lymphonodal involvement is necessary. Hence, ultrasonography can improve clinical practice by providing helpful information for dermatosurgeons, which cannot be obtained during clinical examination.

Pre-operative high-frequency ultrasound: a reliable management tool in auricular and nasal non-melanoma skin cancer.

BACKGROUND AND OBJECTIVES: The knowledge of depth infiltration in non-melanoma skin cancer (NMSC) using pre-operative ultrasound could enable clinicians to choose the most adequate therapeutic approach, avoiding unnecessary surgeries and expensive imaging methods, delaying diagnosis and treatment. Our single-center retrospective study determined the usefulness of high-frequency ultrasound (HFUS) for depth infiltration assessment in auricular and nasal NMSC and assessed the subsequent change in therapeutic approach. PATIENTS AND METHODS: In 60 NMSC cases, we assessed the accuracy of HFUS in cartilaginous/bone infiltration detection as well as the correlation of sonographic and histological parameters. RESULTS: In 16.6% of cases, a deep cartilaginous/bone involvement or locoregional disease was identified pre-operatively, resulting in a changed therapeutical scheme of radio-immunological treatment rather than surgery. In two cases, pre-operative HFUS identified local cartilage infiltration, reducing the number of surgical procedures. Forty-eight remaining lesions with no depth infiltration were excised; a correlation of > 99% between the histologic and sonographic tumor depth (p<0.001) was found. CONCLUSIONS: Pre-surgical HFUS influences the therapeutic management in NMSC by detecting subclinical involvement of deeper structures, avoiding more extensive diagnostics, reducing costs, and improving healthcare quality. High-frequency ultrasound should be implemented in dermatosurgery before tumor excision for optimized therapy and improved patient counseling.

Rationale for Using High-Frequency Ultrasound as a Routine Examination in Skin Cancer Surgery: A Practical Approach.

Ultrasound and high-frequency ultrasound assessment of melanoma and non-melanoma skin cancer in the pre-therapeutical setting is becoming increasingly popular in the field of dermatosurgery and dermatooncology, as it can provide clinicians with relevant, "in vivo" parameters regarding tumor lateral and depth extension as well as potential locoregional spread, cancelling the need of more extensive imaging methods and avoiding a delay in diagnosis. Furthermore, preoperative sonography and color Doppler can aid in orienting the clinical diagnosis, being able in numerous situations to differentiate between benign and malignant lesions, which require a different therapeutic approach. This preoperative knowledge is of paramount importance for planning an individualized treatment regimen. Using sonography at the time of diagnosis, important surgical complications, such as neurovascular damage, can be avoided by performing a preoperative neurovascular mapping. Furthermore, sonography can help reduce the number of surgical steps by identifying the lesions' extent prior to surgery, but it can also spare unnecessary surgical interventions in cases of locally advanced lesions, which infiltrate the bone or already present with locoregional metastases, which usually require modern radiooncological therapies in accordance to European guidelines. With this review, we intend to summarize the current indications of sonography in the field of skin cancer surgery, which can help us improve the therapeutic attitude toward our patients and enhance patient counseling. In the era of modern systemic radiooncological therapies, sonography can help better select patients who qualify for surgical procedures or require systemic treatments due to tumoral extension.