Comprehensive Diagnosis, Treatment, and Outcome of Taenia crassiceps Cysticercosis in a Ring-Tailed Lemur (Lemur catta) from a Croatian Zoo: No Longer Unusual?

Taenia crassiceps is a zoonotic tapeworm of the genus Taenia that is distributed throughout the Northern Hemisphere. Wild and domestic carnivores are final hosts, while rodents and rabbits are primarily intermediate hosts, although many other mammals may harbour the larval stage, Cysticercus longicollis. This case report aims to describe C. longicollis infection in a lemur and molecularly characterise the isolated parasite. The excised lesion was subjected to morphological and histopathological examination, which revealed cysticerci of the tapeworm. Formalin-fixed and paraffin-embedded block (FFPEB), as well as the cysticerci fixed with formalin stored for one year, were subjected to molecular analysis, which aimed at detecting the partial mitochondrial cytochrome c oxidase subunit 1 (cox1) gene of Taenia sp. Based on the morphological characteristics, the parasite was identified as a metacestode of T. crassiceps. The presence of the cox1 gene was detected using polymerase chain reaction (PCR) in all samples. A randomly selected PCR product was sequenced and compared with other sequences from the GenBank database, confirming that the detected parasite was T. crassiceps. This article reports the first case of T. crassiceps cysticercosis in a lemur (Lemur catta) in Croatia and emphasises the potential risk of transmission from wild carnivores.

In vitro aging alters the gene expression and secretome composition of canine adipose-derived mesenchymal stem cells.

Introduction: Canine adipose-derived mesenchymal stem cells (cAD-MSCs) hold therapeutic promise due to their regenerative potential, particularly within their secretome. However, concerns arise regarding the impact of in vitro cultivation necessitated for storing therapeutic doses, prompting this study to comprehensively explore the impact of in vitro aging on gene expression and secretome composition. Methods: The study involved collecting abdominal adipose tissue samples from nine healthy female dogs, from which cAD-MSCs were extracted and cultured. Stem cells were validated through trilineage differentiation assays and flow cytometry immunophenotyping. Gene expression profiling using RT-qPCR array, and cAD-MSCs secretome LC-MS/MS analysis, were conducted at passages 3 and 6 to reveal gene expression and protein composition alterations during in vitro culture. Results and Discussion: The results demonstrate that the gene expression and secretome composition of cAD-MSCs were impacted by in vitro aging. Among many alterations in gene expression between two passages, two significant downregulations were noted in the MSC-associated PTPRC and IL10 genes. While the majority of proteins and their functional characteristics were shared between passages, the influence of cell aging on secretome composition is highlighted by 10% of proteins being distinctively expressed in each passage, along with 21 significant up- and downregulations. The functional attributes of proteins detected in passage 3 demonstrated a greater inclination towards supporting the regenerative capacity of cAD-MSCs. Moreover, proteins in passage 6 exhibited a noteworthy correlation with the blood coagulation pathway, suggesting an elevated likelihood of coagulation events. To the best of our knowledge, this study presents the first original perspective on the changes in secretome composition that occur when cAD-MSCs age in vitro. Furthermore, it contributes to broadening the currently restricted knowledge base concerning the secretome of cAD-MSCs. In conclusion, our findings show that the regenerative potential of cAD-MSCs, as well as their secretome, may be compromised by in vitro aging. Therefore, our study suggests a preference for earlier passages when considering these cells for therapeutic applications.

An Outstanding Role of Adipose Tissue in Canine Stem Cell Therapy.

Adipose tissue, previously known as connective tissue with a role in energy storage, is currently changing the course of treatments in veterinary medicine. Recent studies have revealed one particularly impressive function among all the newly discovered functions of adipose tissue. The interactive cells hosted by adipose tissue, the stromal vascular fraction (SVF), and their role in treating numerous diseases have provided a prospective course of research with positive outcomes in regenerative veterinary medicine (RVM). This review describes the main features of adipose tissue, emphasizing an eclectic combination of cells within the SVF and its thus far researched therapeutic possibilities in canine RVM. An afterwards focus is on a highly researched component of the SVF, adipose-derived mesenchymal stem cells (ASCs), which were shown to have an extraordinary impact relying on several proposed mechanisms of action on mitigating pathologies in canines. Furthermore, ASC therapy showed the most significant results in the orthopaedics field and in neurology, dermatology, ophthalmology, gastroenterology, and hepatology, which elevates the possibilities of ASC therapy to a whole new level. Therefore, this review article aims to raise awareness of the importance of research on cellular components, within abundant and easily accessible adipose tissue, in the direction of regenerative therapy in canines, considering the positive outcomes so far. Although the focus is on the positive aspects of cellular therapy in canines, the researchers should not forget the importance of identifying the potential negative aspects within published and upcoming research. Safe and standardized treatment represents a fundamental prerequisite for positively impacting the lives of canine patients.

Morphological and Molecular Evaluation of the Tissue Repair following Nasal Septum Biopsy in a Sheep Model.

OBJECTIVE: Nasal septal pathologies requiring surgical intervention are common in the population. Additionally, nasal chondrocytes are becoming an important cell source in cartilage tissue engineering strategies for the repair of articular cartilage lesions. These procedures damage the nasal septal cartilage whose healing potential is limited due to its avascular, aneural, and alymphatic nature. Despite the high incidence of various surgical interventions that affect septum cartilage, limited nasal cartilage repair characterizations have been performed to date. METHODS: To evaluate the healing of the nasal septum cartilage perforation, a septal biopsy was performed in 14 sheep. Two and 6 months later, the tissue formed on the place of perforation was explanted and compared with the native tissue. Tissue morphology, protein and gene expression of explanted tissue was determined using histological, immunohistochemical and real-time quantitative polymerase chain reaction analysis. RESULTS: Tissue formed on the defect site, 2 and 6 months after the biopsy was characterized as mostly connective tissue with the presence of fibroblastic cells. This newly formed tissue contained no glycosaminoglycans and collagen type II but was positively stained for collagen type I. Cartilage-specific genes COL2, AGG, and COMP were significantly decreased in 2- and 6-month samples compared with the native nasal cartilage. Levels of COL1, COL4, and CRABP1 genes specific for perichondrium and connective tissue were higher in both test group samples in comparison with native cartilage. CONCLUSIONS: Newly formed tissue was not cartilage but rather fibrous tissue suggesting the role of perichondrium and mucosa in tissue repair after nasal septum injury.

The Expression Pattern of Surface Markers in Canine Adipose-Derived Mesenchymal Stem Cells.

The influence of cultivation on the expression pattern of canine adipose-derived mesenchymal stem cells (cAD-MSCs) surface markers, contributing to, among others, the promotion of growth, proliferation, differentiation and immunomodulatory mechanisms of an excellent therapeutic, is still unknown. To fill the gap, we investigated CD90, CD44, CD73, CD29, CD271, CD105, CD45 and CD14 patterns of expression at the protein level with flow cytometry and mRNA level using a real-time polymerase chain reaction array. Gentle variations of expression occurred during cultivation, along with increased CD90, CD44 and CD29 expression, low and decreasing CD271 and CD73 expression and a decrease of initially high CD105. As expected, CD45 and CD14 were not expressed by cAD-MSCs. Interestingly, we discovered a significant decrease of CD73 expression, compared to early (P1-P3) to late (P4-P6) passages, although the CD73 gene expression was found to be stable. The percentage of positive cells was found to be higher for all positive markers up to P4. As CD73's one important feature is a modulation from a pro-inflammatory environment to an anti-inflammatory milieu, the expression of CD73 in our conditions indicate the need to consider the time cells spend in vitro before being transplanted into patients, since it could impact their favourable therapeutical properties.

Nasal Chondrocyte-Based Engineered Grafts for the Repair of Articular Cartilage "Kissing" Lesions: A Pilot Large-Animal Study.

BACKGROUND: Bipolar or "kissing" cartilage lesions formed on 2 opposite articular surfaces of the knee joint are commonly listed as exclusion criteria for advanced cartilage therapies. PURPOSE: To test, in a pilot large-animal study, whether autologous nasal chondrocyte (NC)-based tissue engineering, recently introduced for the treatment of focal cartilage injuries, could provide a solution for challenging kissing lesions. STUDY DESIGN: Controlled laboratory study. METHODS: Osteochondral kissing lesions were freshly introduced into the knee joints of 26 sheep and covered with NC-based grafts with a low or high hyaline-like extracellular matrix; a control group was treated with a cell-free scaffold collagen membrane (SCA). The cartilage repair site was assessed at 6 weeks and 6 months after implantation by histology, immunohistochemistry, and magnetic resonance imaging evaluation. RESULTS: NC-based grafts, independently of their composition, induced partial hyaline cartilage repair with stable integrity in surrounding healthy tissue at 6 months after treatment. The SCA repaired cartilage to a similar degree to that of NC-based grafts. CONCLUSION: Kissing lesion repair, as evidenced in this sheep study, demonstrated the feasibility of the treatment of complex cartilage injuries with advanced biological methods. However, the potential advantages of an NC-based approach over a cell-free approach warrant further investigations in a more relevant preclinical model. CLINICAL RELEVANCE: NC-based grafts currently undergoing phase II clinical trials have a high potential to replace existing cartilage therapies that show significant limitations in the quality and reproducibility of the repair method. We have brought this innovative concept to the next level by addressing a new clinical indication.

Bioreactor-manufactured cartilage grafts repair acute and chronic osteochondral defects in large animal studies.

OBJECTIVES: Bioreactor-based production systems have the potential to overcome limitations associated with conventional tissue engineering manufacturing methods, facilitating regulatory compliant and cost-effective production of engineered grafts for widespread clinical use. In this work, we established a bioreactor-based manufacturing system for the production of cartilage grafts. MATERIALS & METHODS: All bioprocesses, from cartilage biopsy digestion through the generation of engineered grafts, were performed in our bioreactor-based manufacturing system. All bioreactor technologies and cartilage tissue engineering bioprocesses were transferred to an independent GMP facility, where engineered grafts were manufactured for two large animal studies. RESULTS: The results of these studies demonstrate the safety and feasibility of the bioreactor-based manufacturing approach. Moreover, grafts produced in the manufacturing system were first shown to accelerate the repair of acute osteochondral defects, compared to cell-free scaffold implants. We then demonstrated that grafts produced in the system also facilitated faster repair in a more clinically relevant chronic defect model. Our data also suggested that bioreactor-manufactured grafts may result in a more robust repair in the longer term. CONCLUSION: By demonstrating the safety and efficacy of bioreactor-generated grafts in two large animal models, this work represents a pivotal step towards implementing the bioreactor-based manufacturing system for the production of human cartilage grafts for clinical applications. Read the Editorial for this article on doi:10.1111/cpr.12625.