ARNALDO JOSE HERNANDEZ
Projetos de Pesquisa
Unidades Organizacionais
Departamento de Ortopediae Traumatologia, Faculdade de Medicina - Docente
LIM/41 - Laboratório de Investigação Médica do Sistema Músculoesquelético, Hospital das Clínicas, Faculdade de Medicina
LIM/41 - Laboratório de Investigação Médica do Sistema Músculoesquelético, Hospital das Clínicas, Faculdade de Medicina
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conferenceObject Biomechanical Analysis Of A Cell Therapy And Tissue Engineering Strategy For Articular Cartilage Restoration - A Pre-clinical Study(2022) FARIA, R. R.; SANTANNA, J. P.; ASSAD, I. D.; PINHEIRO, C. C.; FERNANDES, T. L.; BUENO, D. F.; HERNANDEZ, A. J.- Human Synovial Mesenchymal Stem Cells Good Manufacturing Practices for Articular Cartilage Regeneration(2018) FERNANDES, Tiago Lazzaretti; KIMURA, Heitor Akio; PINHEIRO, Carla Cristina Gomes; SHIMOMURA, Kazunori; NAKAMURA, Norimasa; FERREIRA, Jose Ricardo; GOMOLL, Andreas H.; HERNANDEZ, Arnaldo Jose; BUENO, Daniela FrancoBackground: Cartilage restoration is a desperately needed bridge for patients with symptomatic cartilage lesions. Chondral lesion is a pathology with high prevalence, reaching as much as 63% of general population and 36% among athletes. Despite autologous chondrocyte implantation versatility, it still fails to fully reproduce hyaline articular cartilage characteristics. Mesenchymal stem cells (MSCs) may be isolated from various known tissues, including discarded fragments at arthroscopy such as synovial membrane. Choice of harvesting site is motivated by MSCs' abilities to modulate immunologic and inflammatory response through paracrine communication. Synovial MSCs have a greater proliferation and strong chondrogenic potential than bone and adipose MSCs and a less hypertrophic differentiation than bone MSCs. Good manufacturing practice (GMP) laboratory techniques for human clinical trials are still novel. To our knowledge, there are only two clinical trials in humans published since today. Purpose: Therefore, this work aimed to isolate and characterize synovial MSCs and evaluated their differentiation properties according to GMP standards. Materials and Methods: One-gram tissue sample from three patients of synovia was harvested at the beginning of arthroscopy surgery. MSCs were isolated, expanded, and characterized by flow cytometry. Results: It was possible to isolate and expand MSCs cultures from synovia, characterize MSCs by flow cytometry using proper monoclonal antibodies, and differentiate MSCs by coloring technique after chondrogenic, adipogenic, and osteogenic differentiations. Cartilage treatment may benefit from these tissue engineering protocols since arthroscopic procedures are routinely performed for different purposes in a previous stage and a favorable chondronegic differentiation cell lineage may be collected and stored in a less invasive way. Conclusion: Laboratory protocols established according to presented GMP were able to isolate and characterize MSCs obtained from synovia.
- Tissue Engineering and Cell Therapy for Cartilage Repair: Preclinical Evaluation Methods(2022) SANTANNA, Joao P. C.; FARIA, Rafaella R.; ASSAD, Isabella P.; PINHEIRO, Carla C. G.; AIELLO, Vera D.; ALBUQUERQUE-NETO, Cyro; BORTOLUSSI, Roberto; CESTARI, Idagene A.; MAIZATO, Marina J. S.; HERNANDEZ, Arnaldo J.; BUENO, Daniela F.; FERNANDES, Tiago L.A chondral injury is a limiting disease that can affect the quality of life and be an economic burden due to the cost of immediate treatment and loss in work productivity. If left untreated, such an injury may progress to osteoarthritis, a degenerative and debilitating joint disease characterized by pain and functional impairment. Mesenchymal stromal cells (MSCs), which have immune-modulatory properties and the ability to differentiate into chondroblasts and osteoblasts, are a predictable source for the treatment of cartilage injuries. This article presents tools to evaluate cartilage restoration by tissue engineering and cell therapy treatment in a translational and preclinical large animal model. In this controlled experimental study with 14 miniature pigs, a scaffold-free tissue engineering construct (TEC) derived from dental pulp and synovial MSCs for cartilage therapy was tested. Total thickness cartilage defects were performed in both posterior knees. The defect was left empty in one of the knees, and the other received the TEC. The tissue repair was morphologically assessed by magnetic resonance imaging (MRI) using the three-dimensional double echo steady-state (3D-DESS) sequence, and compositional assessment was carried out based on the T2 mapping technique. The osteochondral specimens were fixed for histopathology, decalcified, subjected to standard histological processing, sectioned, and stained with hematoxylin and eosin. The sections stained for immunohistochemical detection of collagen types were digested with pepsin and chondroitinase and incubated with antibodies against them. The mechanical evaluation involved analysis of Young's modulus of the cartilage samples based on the indentation and maximum compression test. In addition, a finite element model was used to simulate and characterize properties of the osteochondral block. At 6 months after surgery, there were no complications with the animals and the MRI, histological, immunohistochemical, and biomechanical evaluations proved to be effective and qualified to differentiate good quality chondral repair from inadequate repair tissue. The proposed methods were feasible and capable to properly evaluate the defect filled with TEC containing stromal cells after 6 months of follow-up in a large animal model for articular cartilage restoration. Impact StatementArticular chondral injuries are prevalent and represent an economic burden due to the cost of treatment. The engineering of cartilage tissue can promote the repair of chondral injuries and is dependent on selecting appropriate cells and biocompatible frameworks. In this article, methods for evaluation of a scaffold-free cell delivery system made from mesenchymal stromal cells were present in a translational study that allows further clinical safety and efficacy trials.
conferenceObject Articular Cartilage Repair By Tissue Engineering And Cell Therapy: A Pre-clinical Trialimaging Evaluation(2022) SANTANNA, J. P.; FARIA, R. R.; PINHEIRO, C. C.; AIELLO, V. D.; FERNANDES, T. L.; BUENO, D. F.; HERNANDEZ, A. J.conferenceObject Articular Cartilage Repair By Tissue Engineering And Cell Therapy: A Pre-clinical Trialimaging Evaluation(2022) SANTANNA, J. P.; FARIA, R. R.; ASSAD, I. P.; PINHEIROS, C. C.; BUENO, D. F.; HERNANDEZ, A. J.; FERNANDES, T. L.- Development of a Novel Large Animal Model to Evaluate Human Dental Pulp Stem Cells for Articular Cartilage Treatment(2018) FERNANDES, Tiago Lazzaretti; SHIMOMURA, Kazunori; ASPERTI, Andre; PINHEIRO, Carla Cristina Gomes; CAETANO, Heloisa Vasconcellos Amaral; OLIVEIRA, Claudia Regina G. C. M.; NAKAMURA, Norimasa; HERNANDEZ, Arnaldo Jose; BUENO, Daniela FrancoChondral lesion is a pathology with high prevalence, reaching as much as 63% of general population and 36% among athletes. The ability of human Dental Pulp Stem Cells (DPSCs) to differentiate into chondroblasts in vitro suggests that this stem cell type may be useful for tissue bioengineering. However, we have yet to identify a study of large animal models in which DPSCs were used to repair articular cartilage. Therefore, this study aimed to describe a novel treatment for cartilage lesion with DPSCs on a large animal model. Mesenchymal stem cells (MSC) were obtained from deciduous teeth and characterized by flow cytometry. DPSCs were cultured and added to a collagen type I/III biomaterial composite scaffold. Brazilian miniature pig (BR-1) was used. A 6-mm diameter, full-thickness chondral defect was created in each posterior medial condyle. The defects were covered with scaffold alone or scaffold + DPSCs on the contralateral side. Animals were euthanized 6 weeks post-surgery. Cartilage defects were analyzed macroscopically and histology according to modified O'Driscoll scoring system. Flow cytometry confirmed characterization of DPSCs as MSCs. Macroscopic and histological findings suggested that this time period was reasonable for evaluating cartilage repair. To our knowledge, this study provides the first description of an animal model using DPSCs to study the differentiation of hyaline articular cartilage in vivo. The animals tolerated the procedure well and did not show clinical or histological rejection of the DPSCs, reinforcing the feasibility of this descriptive miniature pig model for pre-clinical studies.
- Systematic Review of Human Dental Pulp Stem Cells for Cartilage Regeneration(2020) FERNANDES, Tiago Lazzaretti; SANTANNA, Joao Paulo Cortez de; FRISENE, Igor; GAZARINI, Joao Paulo; PINHEIRO, Carla Cristina Gomes; GOMOLL, Andreas H.; LATTERMANN, Christian; HERNANDEZ, Arnaldo Jose; BUENO, Daniela FrancoBackground: Symptomatic cartilage lesions and early osteoarthritis produce significant clinical and economic burdens. Cartilage repair can improve the symptoms and delay arthroplasty. The complete healing of damaged cartilage with the consistent reproduction of normal hyaline cartilage has not yet been achieved. The choice of harvesting site might influence the cells' abilities to modulate immunologic and inflammatory responses. Recently, dental pulp has been shown to contain a stem cell niche consisting of dental pulp stem cells (DPSCs) that maintain their self-renewal capacity due to the active environment in the dental pulp of deciduous teeth. Objective: The aim of this study was to critically review the current literature on the potential and limitations of the use of dental pulp-derived mesenchymal stem cells in cell-based therapies for cartilage regeneration. Methods: An electronic, customized search of scientific articles was conducted using the PubMed/MEDLINE and EMBASE databases from their inception to December 2018. The inclusion criteria were applied, and the articles that described the use of DPSC in cartilage treatment were selected for complete evaluation. The articles were classified according to the scaffold used, experimental model, chondrogenic differentiation features, defect location, cartilage evaluation, and results. After the application of the eligibility criteria, a total of nine studies were selected and fully analyzed. Results: A variety of animal models were used, including mice, rats, rabbits, and miniature pigs, to evaluate the quality and safety of human DPSCs in the repair of cartilage defects. Among the articles, two studies focused on preclinical models of cartilage tissue engineering. Five studies implanted DPSCs in other animal sites. Conclusion: The use of DPSCs is a potential new stem cell therapy for articular cartilage repair. The preclinical evidence discussed in this article provides a solid foundation for future clinical trials. Impact statement Osteoarthritis presents an ever-increasing clinical and socioeconomic burden. While cartilage repair has the potential to improve symptoms and delay joint replacement, complete regeneration of hyaline cartilage has been an elusive goal. Dental pulp has been shown to contain a niche that protects dental pulp stem cells (DPSCs) from the cumulative effects of genetic and environmental factors and maintains their self-renewal capacity due to the active environment. Transplantation and preclinical trials have demonstrated the strong potential of regenerative tissue-engineering protocols using DPSCs.
conferenceObject Cartilage regeneration with human dental pulp stem cells - a systematic review(2020) SANTANNA, J.; FERNANDEZ, T. L.; BUENO, D. F.; PINHEIRO, C. C.; HERNANDEZ, A. J.conferenceObject Biomechanical Analysis Of A Cell Therapy And Tissue Engineering Strategy For Articular Cartilage Restoration - A Pre-clinical Study(2022) SANTANNA, J. P.; FARIA, R. R.; PINHEIRO, C. C.; TABET, C. G.; HERNANDEZ, A. J.; BUENO, D. F.; FERNANDES, T. L.