Kisspeptin (Kp-10) inhibits in vitro osteogenic differentiation of multipotent mesenchymal stromal cells extracted from the bone marrow of adult rats.

Kisspeptin (Kp-10) is a neuropeptide that binds to GPR54 receptors, exerting several functions mainly in the nervous and reproductive systems of the body. However, its effects and mechanisms of action on the skeletal system remain poorly understood. This study evaluated the effects of different concentrations of Kp-10 on in vitro osteogenic differentiation of multipotent mesenchymal stromal cells (MSCs) extracted from the bone marrow (BM) of adult Wistar rats. Two-month-old female rats were euthanized to extract BM from long bones to obtain MSCs. Four experimental groups were established in vitro: a control and Kp-10 at concentrations of 0.01, 0.05 and, 0.1 µg/mL. After induction of osteogenic differentiation, cell viability was evaluated using the 3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyl tetrazolium bromide (MTT) assay, alkaline phosphatase activity, collagen synthesis, percentage of area covered by MSCs/field and mineralized nodules/field, and immunocytochemistry of the GPR54 receptor tests. Furthermore, evaluation of gene transcripts for type I collagen, Runx-2, Bmp-2, bone sialoprotein, osteocalcin and osteopontin was performed using real-time RT-qPCR. It was observed that MSCs expressed GPR54 receptor to which Kp-10 binds during osteogenic differentiation, promoting a negative effect on osteogenic differentiation. This effect was observed at all the Kp-10 concentrations in a concentration-dependent manner, characterized by a decrease in the activity of alkaline phosphatase, collagen synthesis, mineralized nodules, and decreased expression of gene transcripts for type I collagen, osteocalcin, osteopontin, and Runx-2. Thus, Kp-10 inhibits in vitro osteogenic differentiation of MSCs extracted from the BM of adult Wistar rats.

TNF-α mediated upregulation of NaV1.7 currents in rat dorsal root ganglion neurons is independent of CRMP2 SUMOylation.

Clinical and preclinical studies have shown that patients with Diabetic Neuropathy Pain (DNP) present with increased tumor necrosis factor alpha (TNF-α) serum concentration, whereas studies with diabetic animals have shown that TNF-α induces an increase in NaV1.7 sodium channel expression. This is expected to result in sensitization of nociceptor neuron terminals, and therefore the development of DNP. For further study of this mechanism, dissociated dorsal root ganglion (DRG) neurons were exposed to TNF-α for 6 h, at a concentration equivalent to that measured in STZ-induced diabetic rats that developed hyperalgesia. Tetrodotoxin sensitive (TTXs), resistant (TTXr) and total sodium current was studied in these DRG neurons. Total sodium current was also studied in DRG neurons expressing the collapsin response mediator protein 2 (CRMP2) SUMO-incompetent mutant protein (CRMP2-K374A), which causes a significant reduction in NaV1.7 membrane cell expression levels. Our results show that TNF-α exposure increased the density of the total, TTXs and TTXr sodium current in DRG neurons. Furthermore, TNF-α shifted the steady state activation and inactivation curves of the total and TTXs sodium current. DRG neurons expressing the CRMP2-K374A mutant also exhibited total sodium current increases after exposure to TNF-α, indicating that these effects were independent of SUMOylation of CRMP2. In conclusion, TNF-α sensitizes DRG neurons via augmentation of whole cell sodium current. This may underlie the pronociceptive effects of TNF-α and suggests a molecular mechanism responsible for pain hypersensitivity in diabetic neuropathy patients.

Propriedades mecânicas de meniscos frescos de coelhos e preservados em glicerina 98 por cento / Mechanical properties of the fresh rabbit menisci and of the menisci preserved in glycerin 98 percent

O presente estudo avaliou a resistência à compressão de meniscos mediais de coelhos da raça Nova Zelândia, por meio de teste mecânico de compressão. Trinta meniscos foram distribuídos em três grupos: grupo MF, composto por dez meniscos frescos; grupo MG, dez meniscos preservados em glicerina 98 por cento, por 30 dias, e grupo MR, dez meniscos preservados em glicerina 98 por cento, por 30 dias e reidratados em NaCl 0,9 por cento, por 12 horas. Os meniscos de cada grupo foram submetidos ao teste de compressão no sentido perpendicular ao seu plano anatômico regular e foram avaliados o limite de elasticidade, a deformação elástica, a tensão ao ponto de ruptura e ao limite de elasticidade e ainda, o índice de rigidez. Os meniscos dos grupos preservados, MG e MR, tiveram o limite elástico semelhante ao grupo de meniscos frescos (MF). O grupo de meniscos em glicerina (MG) apresentou menor capacidade de deformação elástica (P<0,05) que os grupos MF e MR, e maior capacidade de sofrer tensão ao limite elástico. Os meniscos do grupo (MG) apresentaram maior rigidez (P<0,05) que os meniscos dos grupos MF e MR. Conclui-se que o grupo de meniscos preservados em glicerina 98 por cento, seguido de reidratação em NaCl 0,9 por cento (MR), não apresentou alterações significativas na capacidade de resistência ao limite elástico dos meniscos.

The present study evaluated the compressive strength of medial menisci of New Zealand rabbits, through mechanical compression test. Thirty menisci were distributed in three groups: group MF, composed by ten fresh menisci; MG group, composed by ten menisci preserved in 98 percent glycerin for 30 days; and, group MR, ten menisci preserved in 98 percent glycerin for 30 days and rehydrated in NaCl 0.9 percent for 12 hours. The menisci in each group were submitted to compression test in the perpendicular direction to the anatomical plane and had the elasticity limit, the elastic deformity, the rupture stress point and the stiffness index evaluated. The menisci from the preserved groups MG and MR had the elastic limit similar to the fresh menisci group (MF). The group of menisci preserved in glycerin (MG) presented lower elastic deformity capacity (P<0.05) if compared to the other groups, MF and MR, and a higher tension capacity at elastic limit. The menisci from group (MG) presented higher stiffness (P<0.05) than the ones in the MF and MR groups. It can be concluded that the group menisci preserved in glycerin 98 percent followed by rehydration in Nacl 0,9 percent (MR), did not showed any significant alterations in the capacity of the menisci elastic limit.