ABSTRACT Triple-negative breast cancer (TNBC) represents a particularly aggressive form of breast tumors. Mitochondrial dysfunction represses the proliferation of TNBC cells. Ubiquitin-specific proteases 34 (USP34) has been predicted to be abnormally overexpressed in TNBC. This research examined the role of USP34 in the mitochondrial function modulation of TNBC. Herein, cell proliferation was evaluated by the 5-ethynyl-2’-deoxyuridine assay. Mitochondrial membrane potential was detected employing the JC-1 assay. Mitochondrial superoxide was measured utilizing MitoSOX Red assay. Mito‑Tracker Red CMXRos staining was selected to monitor mitochondrial network structure. The relationship among USP34, eukaryotic translation initiation factor 3 m (eIF3m), and mitochondrial carrier homolog 2 (MTCH2) was validated by co-immunoprecipitation, GST-pull down, RNA immunoprecipitation and RNA-pull down analysis. We found that USP34 silencing inhibited cell proliferation by inducing mitochondrial dysfunction in TNBC cells. USP34 maintained the stability of the eIF3m protein through deubiquitination. Overexpression of eIF3m countered the mitochondrial dysfunction induced by USP34 silencing. Furthermore, eIF3m upregulated the MTCH2 level by directly binding to its 5’UTR region. MTCH2 overexpression reversed the damaging effect of eIF3m silencing on mitochondrial function. Collectively, USP34 maintained the stability of eIF3m protein through deubiquitination; the upregulated eIF3m bound to the 5’UTR of MTCH2 mRNA to promote MTCH2 expression, thereby maintaining mitochondrial function and promoting the malignant progression of TNBC.

ABSTRACT Due to the prevalence of hematoxylin and eosin (H&E) staining in routine histological preparations, understanding the factors that impact stain color characteristics is vital to attain consistently high-quality stains. In the last decade, increased use of digital pathology and image analysis (specifically by optical density [OD] measures) has provided new ways of assessing staining precision. This paper combines data from two studies that tracked H&E staining quality in both nuclear and cytoplasmic components of 12 tissues (11 human and 1 porcine) by OD after overuse of H&E staining reagents from 5 vendors. Both studies showed a decrease in eosin stain intensity by OD and visual inspection (by a histologist) with reagent overuse. This trend varied in degree by tissue type and reagent vendor. Nonetheless, staining quality for H&E staining from all vendors and for all organs remained acceptable (but not always optimal) for microscopic evaluation by the College of American Pathologists (CAP) and National Society for Histotechnology (NSH) staining criteria when stained sections were reviewed by a board-certified veterinary pathologist.

ABSTRACT The application of Clear Gorilla Glue® household adhesive to microscope slides has been found to enhance the mounting and retention of traditionally difficult tissue types, notably bone and tooth specimens. Improvement in end results were observed across H&E staining, immunohistochemistry, and in situ hybridization.

ABSTRACT The discovery of biomarkers, essential for successful drug development, is often hindered by the limited availability of tissue samples, typically obtained through core needle biopsies. Standard ‘omics platforms can consume significant amounts of tissue, forcing scientist to trade off spatial context for high-plex assays, such as genome-wide assays. While bulk gene expression approaches and standard single-cell transcriptomics have been valuable in defining various molecular and cellular mechanisms, they do not retain spatial context. As such, they have limited power in resolving tissue heterogeneity and cell–cell interactions. Current spatial transcriptomics platforms offer limited transcriptome coverage and have low throughput, restricting the number of samples that can be analyzed daily or even weekly. While the Digital Spatial Profiling (DSP) method does not provide single-cell resolution, it presents a significant advancement by enabling scalable whole transcriptome and ultrahigh-plex protein analysis from distinct tissue compartments and structures using a single tissue slide. These capabilities overcome significant constraints in biomarker analysis in solid tissue specimens. These advancements in tissue profiling play a crucial role in deepening our understanding of disease biology and in identifying potential therapeutic targets and biomarkers. To enhance the use of spatial biology tools in drug discovery and development, the DSP Scientific Consortium has created best practices guidelines. These guidelines, built on digital spatial profiling data and expertise, offer a practical framework for designing spatial studies and using current and future spatial biology platforms. The aim is to improve tissue analysis in all research areas supporting drug discovery and development.

ABSTRACT With rates growing quickly with age, osteoarthritis (OA) is the most common cause of chronic disability in aging persons. The discomfort and reduced motion associated with osteoarthritis have a significant impact on quality of life, and there is no known solution. Runt-related transcription factor 1(Runx1) has been shown to play a protective role in the development of osteoarthritis by promoting chondrogenesis. We had created models of ageing mice with osteoarthritis by anterior cruciate ligament transection (ACLT) and analyzed the effects of intra-articular injection of adeno-associated virus/Runx1 (AAV/Runx1) on the models. The results showed that the AAV/Runx1-group maintained better articular cartilage integrity and retained more proteoglycan than the OA group after injection of AAV-Runx1. The markers related to pathological changes in cartilage were downregulated, while the markers related to physiological changes in cartilage were upregulated. This suggests that Runx1 may impede OA progression on the knee joint of ageing mice, potentially playing a protective role in OA and becoming a probable treatment target for osteoarthritis among ageing patients in the future.

ABSTRACT Digital pathology (DP) is indisputably the future for histopathology laboratories. The process of digital implementation requires deep workflow reorganisation which involves an interdisciplinary team. This transformation may have the greatest impact on the Histotechnologist (HTL) profession. Our review of the literature has clearly revealed that the role of HTLs in the establishment of DP is being unnoticed and guidance is limited. This article aims to bring HTLs from behind-the-scenes into the spotlight. Our objective is to provide them guidance and practical recommendations to successfully contribute to the implementation of a new digital workflow. Furthermore, it also intends to contribute for improvement of study programs, ensuring the role of HTL in DP is addressed as part of graduate and post-graduate education. In our review, we report on the differences encountered between workflow schemes and the limitations observed in this process. The authors propose a digital workflow to achieve its limitless potential, focusing on the HTL’s role. This article explores the novel responsibilities of HTLs during specimen gross dissection, embedding, microtomy, staining, digital scanning, and whole slide image quality control. Furthermore, we highlight the benefits and challenges that DP implementation might bring the HTLs career. HTLs have an important role in the digital workflow: the responsibility of achieving the perfect glass slide.

ABSTRACT Collection, preservation, and shipment of histological specimens in low-resource settings is challenging. We present a novel method that achieved excellent preservation of placental specimens from rural Mali by using formalin fixation, ethanol dehydration, and long-term storage in a solar-powered freezer. Sample preservation success was 92%, permitting evaluation of current and past malaria infection, anemia, placental maturity, and inflammation. Using RNAscope® hybridization we were able to visualize cell-specific gene expression patterns in the formalin-fixed paraffin-embedded (FFPE) specimens. Additionally, our method entailed mirrored sampling from the two cut faces of a cotyledon, one for the FFPE workflows and the other for storage in RNAlater™ and RNA-seq.