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BackgroundWe previously demonstrated the therapeutic benefits of pentosan polysulfate (PPS) in a rat model of mucopolysaccharidosis (MPS) type VI. Reduction of inflammation, reduction of glycosaminoglycan (GAG) storage, and improvement in the skeletal phenotype were shown. Herein, we evaluate the long-term safety and therapeutic effects of PPS in a large animal model of a different MPS type, MPS I dogs. We focused on the arterial phenotype since this is one of the most consistent and clinically significant features of the model.Methodology/Principal FindingsMPS I dogs were treated with daily oral or biweekly subcutaneous (subQ) PPS at a human equivalent dose of 1.6 mg/kg for 17 and 12 months, respectively. Safety parameters were assessed at 6 months and at the end of the study. Following treatment, cytokine and GAG levels were determined in fluids and tissues. Assessments of the aorta and carotid arteries also were performed. No drug-related increases in liver enzymes, coagulation factors, or other adverse effects were observed. Significantly reduced IL-8 and TNF-alpha were found in urine and cerebrospinal fluid (CSF). GAG reduction was observed in urine and tissues. Increases in the luminal openings and reduction of the intimal media thickening occurred in the carotids and aortas of PPS-treated animals, along with a reduction of storage vacuoles. These results were correlated with a reduction of GAG storage, reduction of clusterin 1 staining, and improved elastin integrity. No significant changes in the spines of the treated animals were observed.ConclusionsPPS treatment led to reductions of pro-inflammatory cytokines and GAG storage in urine and tissues of MPS I dogs, which were most evident after subQ administration. SubQ administration also led to significant cytokine reductions in the CSF. Both treatment groups exhibited markedly reduced carotid and aortic inflammation, increased vessel integrity, and improved histopathology. We conclude that PPS may be a safe and useful therapy for MPS I, either as an adjunct or as a stand-alone treatment that reduces inflammation and GAG storage.

Objective Hyaline cartilage degenerative pathologies induce morphologic and biomechanical changes resulting in cartilage tissue damage. In pursuit of therapeutic options, electrical and mechanical stimulation have been proposed for improving tissue engineering approaches for cartilage repair. The purpose of this review was to highlight the effect of electrical stimulation and mechanical stimuli in chondrocyte behavior. Design Different information sources and the MEDLINE database were systematically revised to summarize the different contributions for the past 40 years. Results It has been shown that electric stimulation may increase cell proliferation and stimulate the synthesis of molecules associated with the extracellular matrix of the articular cartilage, such as collagen type II, aggrecan and glycosaminoglycans, while mechanical loads trigger anabolic and catabolic responses in chondrocytes. Conclusion The biophysical stimuli can increase cell proliferation and stimulate molecules associated with hyaline cartilage extracellular matrix maintenance.

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Cell membrane is a lipid bilayer that allows the flow of ions through their ionic pumping proteins. The ionic flow can be stimulated with external stimuli to activate specific signaling pathways intracellularly. Although studies have applied electric and magnetic stimuli to modify the cell function, the parameters to stimulate the cell membrane are unknown. Accordingly, a computational model to simulate the effect of electric and magnetic fields on the cell membrane was developed. Cells were stimulated with electric fields from 45 × 103 V/m to 12.6 × 105 V/m and magnetic fields of 2 mT, at frequencies of 60 kHz, 10 MHz, and 1 GHz. Results showed that the electric fields applied to the cell membrane tend to increase according to the frequency used, while magnetic fields do not have any effect on it. It was observed that electric fields generate a high voltage concentrator in the cell membrane of ellipsoidal cells when a frequency window from 1 kHz to 1 GHz was applied. These findings demonstrate that depending on the intensity of the field and frequency, it was possible to stimulate different cell membrane zones. This model is a promising tool to establish the adequate parameters to stimulate cells, and accurately predict if the stimulation modifies the cell membrane potential.

sistema funcional para generación de puntaje semi-automático para diferenciación condrogénica. Corroboramos estos resultados mediante análisis bioquímico con resultados comparables. En nuestro saber este es el primer reporte en evaluar esta metodología, la cual puede ser útil para otras aplicaciones en el campo biológico o médico.Palabras clave: células mesenquimales, condrogénesis in vitro, glicosaminoglicanos, ImageJ. ResumoEstabelecimento de um sistema de pontuação semi-automático para ensaios histoquímicos e imuno-histoquímicos para avaliar diferenciação condrogênica in vitro. Durante o desenvolvimento embrionário os membros emergem a partir da condensação de células mesenquimais e sua diferenciação em condrócitos em um processo chamado condrogênese. Estes condrócitos sintetizam glicosaminoglicanos, desempenhando um papel importante neste processo. Existe um sistema de condrogénese in vitro utilizando células mesenquimatosas, geralmente avaliado por histoquímica. Objetivo. Estabelecer um sistema de pontuação semi-automático para ensaios histoquímicos e imuno-histoquímicos. Materiais e métodos. Na condrogênese as células foram cultivadas com meio indutor em agregados, durante três semanas. Os glicosaminoglicanos totais foram determinados pelo azul de dimetileno. Para a análise histológica os agregados foram corados com Azul Alciano e imuno-histoquímica para detecção de agrecan. A pontuação semi-automática foi obtida utilizando o programa ImageJ. Resultados. As células mesenquimais cultivadas em meio de diferenciação condrogênica tiveram uma concentração de proteína comparável durante as três semanas de cultura, o que sugere uma celularidade similar. A concentração de glicosaminoglicanos foi maior para os agregados cultivados em meio condrogênico. A mesma tendência foi observada para a coloração com Azul Alciano segundo as pontuações do observador cego e a análise com ImageJ. Finalmente, os resultados de imuno-histoquímica de pontuações dados pelo observador e aqueles dados pela análise ImageJ revelaram uma tendência decrescente ao longo do tempo para os agregados em meio condrogênico. Conclusão. Nós realizamos um sistema funcional para gerar pontuação semi-automática para diferenciação condrogênica. Nós corroboramos esses resultados por análise bioquímica com resultados comparáveis. Segundo nosso conhecimento, este é o primeiro estudo a avaliar esta metodologia, que pode ser útil para outras aplicações no campo biológico ou médico.Palavras-chave: células mesenquimais, condrogênese in vitro, glicosaminoglicanos, ImageJ.

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The growth plate is a cartilaginous structure located in the metaphysis of long bones, characterized histologically by its stratification and columnar arrangement. It is responsible for assuring longitudinal growth. Evaluation of growth plate histological characteristics has been traditionally performed using qualitative observation; however, some quantitative approaches have been reported using complex techniques. Here, we propose a simple quantitative images based analysis in order to evaluate objectively columnar arrangement within growth plate. For this, we defined six descriptors that were condensated in a geometric tensor. This tensor could be used as a single parameter to evaluate the growth plate organization. Validation of the tensor was performed with growth plate microphotographs of three healthy species (rat, pig and rabbit) and an abnormal one (Csf1tl/Csf1tl rat) found in specialized literature. According to our results, the descriptors and the tensor give a complete picture of the organization of the growth plate, reflecting the expected stratification and columnar arrangement of the cells within the tissue. This methodology could be a reliable tool for evaluation of growth plate structure for research and diagnostic purposes, taking into account that it can be easily implemented.