Exploring Canine Mammary Cancer through Liquid Biopsy: Proteomic Profiling of Small Extracellular Vesicles.

(Background). Canine mammary tumors (CMTs) have emerged as an important model for understanding pathophysiological aspects of human disease. Liquid biopsy (LB), which relies on blood-borne biomarkers and offers minimal invasiveness, holds promise for reflecting the disease status of patients. Small extracellular vesicles (SEVs) and their protein cargo have recently gained attention as potential tools for disease screening and monitoring. (Objectives). This study aimed to isolate SEVs from canine patients and analyze their proteomic profile to assess their diagnostic and prognostic potential. (Methods). Plasma samples were collected from female dogs grouped into CMT (malignant and benign), healthy controls, relapse, and remission groups. SEVs were isolated and characterized using ultracentrifugation (UC), nanoparticle tracking analysis (NTA) and transmission electron microscopy (TEM). Proteomic analysis of circulating SEVs was conducted using liquid chromatography-mass spectrometry (LC-MS). (Results). While no significant differences were observed in the concentration and size of exosomes among the studied groups, proteomic profiling revealed important variations. Mass spectrometry identified exclusive proteins that could serve as potential biomarkers for mammary cancer. These included Inter-alpha-trypsin inhibitor heavy chain (ITIH2 and ITI4), phosphopyruvate hydratase or alpha enolase (ENO1), eukaryotic translation elongation factor 2 (eEF2), actin (ACTB), transthyretin (TTR), beta-2-glycoprotein 1 (APOH) and gelsolin (GSN) found in female dogs with malignant tumors. Additionally, vitamin D-binding protein (VDBP), also known as group-specific component (GC), was identified as a protein present during remission. (Conclusions). The results underscore the potential of proteins found in SEVs as valuable biomarkers in CMTs. Despite the lack of differences in vesicle concentration and size between the groups, the analysis of protein content revealed promising markers with potential applications in CMT diagnosis and monitoring. These findings suggest a novel approach in the development of more precise and effective diagnostic tools for this challenging clinical condition.

MicroRNA Profiling Using a PCR-Based Method.

MicroRNAs (miRNAs) are small non-coding RNA molecules involved in the post-transcriptional regulation of specific mRNA targets, thus possibly controlling many biological processes. The miRNA profiling analysis can contribute to understanding several signaling pathways, as biomarkers for molecular diagnostic, as well as potential to be used as therapeutic targets. The miRNAs expression can be analyzed by quantitative reverse transcription PCR (RT-qPCR), microarrays, and RNA sequencing. The RT-qPCR method is sensitive and specific and has a lower cost when compared to other techniques as microarrays and RNA sequencing. Therefore, the protocol presented in this chapter describes step by step all the details to perform miRNA analysis using primer-based RT-qPCR.

Acute responses to barefoot 5 km treadmill running involve changes in landing kinematics and delayed onset muscle soreness.

BACKGROUND: Barefoot running has gained popularity among physical activity practitioners, but there is a lack of information regarding the acute adaptations to this running technique without supervision. Information about acute adaptations can help to define the best way to insert barefoot running in the routine of runners willing to, and also provide orientation for those people who want to experience this technique. RESEARCH QUESTION: What acute adaptations can be observed among recreational runners exposed to barefoot running? METHODS: Sagittal 2D kinematics, plantar pressure, foot sensitivity and delayed onset muscle soreness were compared between conditions of shod and barefoot running in 13 recreational runners who performed two trials of 5 km treadmill running. RESULTS: We found an acute effect of barefoot running on foot landing that changes from a rearfoot strike to a forefoot strike pattern. This change most likely contributed to the increase in neuromuscular recruitment of calf muscles (i.e. gastrocnemius and soleus) resulting in higher perception of delayed onset muscle soreness. Barefoot running also elicited higher stride cadence. Plantar pressure before and after running revealed higher pressure in the different foot regions after barefoot running. Foot sensitivity increased after running regardless of the footwear condition. CONCLUSION: Barefoot running has acute effects on running technique including higher perception of delayed onset muscle soreness in the 48 h following the exercise. SIGNIFICANCE: Our results highlight the importance of following participants for days after a first session of barefoot running in order to properly manage the acute adaptation periods as well provide precise advices for those trying the barefoot technique.