Low-intensity pulsed ultrasound (LIPUS) treatment of cultured chondrocytes stimulates production of CCN family protein 2 (CCN2), a protein involved in the regeneration of articular cartilage: mechanism underlying this stimulation.

OBJECTIVE: CCN family protein 2/connective tissue growth factor (CCN2/CTGF) promotes cartilage regeneration in experimental osteoarthritis (OA) models. However, CCN2 production is very low in articular cartilage. The aim of this study was to investigate whether or not CCN2 was promoted by cultured chondrocytes treated with low-intensity pulsed ultrasound (LIPUS) and to clarify its mechanism. METHODS: Human chondrocytic cell line (HCS)-2/8, rat primary epiphyseal and articular cartilage cells, and Ccn2-deficient chondrocytes that impaired chondrocyte differentiation, were treated with LIPUS for 20 min at 3.0 MHz frequency and 60 mW/cm2 power. Expressions of chondrocyte differentiation marker mRNAs were examined by real-time PCR (RT-PCR) analysis from HCS-2/8 cells and Ccn2-deficient chondrocytes at 30 min and 1 h after LIPUS treatment, respectively. CCN2 production was examined by Western blotting after 5 h of LIPUS treatment. Moreover, Ca2+ influx was measured by using a Fluo-4 probe. RESULTS: The gene expression of chondrocyte differentiation markers and CCN2 production were increased in cultured chondrocytes treated with LIPUS. In addition, Ca2+ influx and phosphorylation of p38 mitogen-activated protein kinase (MAPK) and extracellular signal-regulated kinase (ERK)1/2 were increased by LIPUS treatment, and the stability of TRPV4 and BKca channel mRNAs was decreased by siRNA against CCN2. Consistent with those findings, the LIPUS-induced the gene expressions of type II collagen (COL2a1) and Aggrecan (ACAN) observed in wild-type cells were not observed in the Ccn2-deficient chondrocytes. CONCLUSION: These data indicate that chondrocyte differentiation represented by CCN2 production was mediated via MAPK pathways activated by LIPUS-stimulated Ca2+ influx, which in turn was supported by the induced CCN2 molecules in articular chondrocytes.

Óleo de melaleuca (Melaleuca alternifolia Maiden & Betche, Cheel) no controle de cercosporiose em beterraba / Tea tree oil (Melaleuca alternifolia Maiden& Betche, Cheel) in control of Cercospora beticola Sacc. on Beet

RESUMO O óleo volátil da melaleuca (Melaleuca alternifolia Maiden & Betche, Cheel) possui atividade antimicrobiana podendo causar efeitos sobre as plantas. Avaliou-se a inibição do óleo em Cercospora beticolaSacc., e seu efeito no aumento da produção e qualidade de raízes de beterraba. As doses foram de 0,13; 0,67; 0,80 e 1,00% do óleo, além das testemunhas composta pelo meio de cultura Batata Dextrose Ágar (BDA) no experimento in vitro, e água no experimento in vivo. As plantas foram pulverizadas duas vezes por semana. O delineamento foi inteiramente casualizado, com 4 repetições, e as médias foram comparadas pelo teste Tukey a 5% de probabilidade. O índice de infecção das folhas foi determinado por escala diagramática além do peso e diâmetro das raízes. Os resultados de inibição do crescimento micelial para as doses do óleo foram 0; 56; 87; 83 e 99%, e os índices de infecção: 77,08; 35,62; 21,04; 19,37 e 20,00%, respectivamente, para a testemunha e as doses 0,13; 0,67; 0,80 e 1,00% do óleo. Somente na concentração de 0,80% o óleo proporcionou relação positiva entre o ganho de peso e o diâmetro das raízes. O óleo de Melaleuca foi eficaz no controle de C. beticola e, como consequência, houve produção de raízes de beterraba com melhor desenvolvimento.

ABSTRACT The volatile oil from Melaleuca (Melaleuca alternifolia Maiden & Betche Cheel.) has antimicrobial properties and can promote several effects on plant cultivation. The aim of this study was to evaluate the inhibition of the oil in Cercospora beticola Sacc. and if it favors the growth and development of beet root. The doses were 0.13, 0.67, 0.8 and 1% of oil, besides the control PDA (potato-dextrose-agar) in vitro (laboratory condition) and with water as treatment control in vivo (field conditions). The plants were sprayed twice a week. The treatments were completely randomized and the averages were compared using the Tukey test at 5%. The infection rate of leaves was measured by diagrammatic scale besides the weight and diameter of tubers. The inhibition results of the radial growth by oil treatments were 0; 56, 87, 83 and 99%, while the infection rate showed: 77.08, 35.62, 21.04, 19.37 and 20% respectively to the control and to the oil concentration of 0,13; 0,67; 0,80 e 1,00%. Only at concentration of 0.8% the tea tree oil showed a positive relationship between tuber´s weight and tuber´s diameter gains. It can be concluded that tea tree oil is effective to controlling C. beticola, and also promotes an increase on development in beet tubers.

Impaired glycolytic metabolism causes chondrocyte hypertrophy-like changes via promotion of phospho-Smad1/5/8 translocation into nucleus.

OBJECTIVE: Hypertrophy-like changes are often observed in chondrocytes during the development of osteoarthritis (OA). These changes play a crucial part in the OA-associated cartilage degradation and osteophyte formation. However, the pathogenesis leading to such changes is still unknown. In this study, we investigated the mechanism by which these hypertrophy-like changes are induced from the viewpoint of impaired glycolytic metabolism. METHODS: The effect of sodium fluoride (NaF) on glycolytic metabolism of cultured chondrocytes was confirmed by measurement of intracellular adenosine triphosphate (ATP) production. Translocation of phosphorylated Smad1/5/8 to the nucleus was evaluated by subcellular fractionation and Western blotting. Chondrocyte hypertrophy-like changes were investigated by real-time RT-PCR and Western blot analysis of differentiation markers. RESULTS: ATP production was dose-dependently decreased by NaF in the human chondrocytic cell line HCS-2/8. In addition, both chondrocyte proliferation and differentiation were inhibited, whereas cell death was promoted by treatment with NaF. Interestingly, combinational treatment with NaF and lactate enhanced translocation of phospho-Smad1/5/8 to the nucleus, as well as gene expression of ALP, VEGF, COL10a1, and matrix metalloproteinase13 (MMP13), which were the markers of late mature and hypertrophic chondrocytes. Furthermore, the production of type X collagen and activation of MMP9 were also promoted under the same conditions. CONCLUSIONS: These findings suggest that decreased ATP production by NaF promotes hypertrophy-like changes via activation of phospho-Smad1/5/8 in the presence of lactate. Novel metabolic aspects of OA pathogenesis are indicated herein.

Prevention of fatigue cracks in ultrahigh molecular weight polyethylene joint components by the addition of vitamin E.

Flaking-type wear, so-called delamination, is often observed in polyethylene joint components. This is thought to occur partly due to crack formation and propagation at grain boundaries. This study examined the effect of vitamin E on the crack formation and/or propagation in UHMWPE by using 2-dimensional sliding fatigue testing and micro indenter testing. An in vitro sliding fatigue test was performed under two simplified articulating movements, and the cracks produced were observed by scanning acoustic tomography (SAT). Gamma-irradiated ultrahigh molecular weight polyethylene (UHMWPE) specimens demonstrated a smaller area of accumulated cracks as compared to virgin specimens, when the loading movement was reciprocated on a single linear locus. However, four out of five gamma-irradiated UHMWPE specimens exhibited severe flaking-like destruction under the complicated sliding condition, suggesting that gamma irradiation accelerated crack propagation under multidirectional loading. All the gamma-irradiated vitamin-E-containing specimens demonstrated no subsurface crack formation and no flaking-like destruction. Results using micro indenter testing showed that the dynamic hardness at grain boundary was higher than that in grain, and was increased by gamma irradiation. This hardening at grain boundary was reduced by adding vitamin E. It is possible that the presence of vitamin E prevents crack propagation partly due to reduced hardness at grain boundaries. The gamma-irradiated vitamin-E-containing UHMWPE is a promising material to prevent flaking-like destruction of polyethylene joint components.

Improved flow injection analysis (FIA) method for determining selenium in biological samples, and the effect of captopril administration on selenium levels and glutathione peroxidase activity in rat.

An improved flow injection analysis (FIA) method has been developed for the determination of trace selenium in biological samples, and this method has been applied to investigate the effect of captopril, an antihypertensive drug having a thiol group, on selenium concentrations in the rat blood, liver and urine. After oral administration of captopril, selenium levels in the blood decreased, while those in the liver increased significantly. However, no pronounced effect was observed on the urinary excretion rate. The glutathione peroxidase activities in the blood and the liver were comparable to the changes in the selenium levels.

[Rapid diagnosis of gram-negative bacteriuria with endotoxin-specific chromogenic test].

Limulus amebocyte lysate test is a simple, yet sensitive laboratory method for detecting endotoxin. Although the specificity of the test was questioned, we recently solved this problem by removing a (1----3)-beta-D-glucan-sensitive factor G from the lysate (Endospecy, Seikagaku Kogyo Co., Ltd, Tokyo). In this study, we have established a method for the determination of endotoxin urine in using Endospecy. An aliquot of 0.2 ml of Endospecy dissolved in 4.4 ml of 0.1 mol/l Tris-HCl buffer (pH 8.0) was added to 5 microliter of urine sample. The mixture was incubated at 37 degrees C for 30 min. After diazo-coupling, absorbance was measured at 545 nm. The standard curve using Escherichia coli 0111: B4 endotoxin showed a good linearity. The addition-recovery test of various endotoxins showed nearly 100% recovery. Normal urine contained less than 60 pg/ml of endotoxin. Urine from patients with Gram-negative bacteriuria showed definitely high values, whereas urines infected either with Gram-positive bacteria or with fungi all showed nearly normal values. This method will be useful as a rapid and reliable test for the diagnosis of urinary tract infection with Gram-negative bacteria.

Thiol exchange reactions involving selenotrisulfides.

The occurrence of in vitro thiol exchange reactions involving selenotrisulfides has been documented by HPLC analyses of reaction solutions. Asymmetric selenotrisulfide (RSSeSR') (R,R' = penicillamine, cysteine, glutathione) was formed by the reactions between (i) a mixture of thiols and selenite, (ii) thiol (R'SH) and symmetric selenotrisulfide (RSSeSR), and (iii) symmetric selenotrisulfides (RSSeSR and R'SSeSR'). Further reaction of an asymmetric selenotrisulfide with thiol (R'SH) produced another symmetric selenotrisulfide (R'SSeSR'). These thiol exchange reactions may offer significant information to elucidate intake and metabolism of selenium in vivo.

Tratamento cirurgico dos adenomas hiperfuncionais das paratiroides. / Surgical treatment of hyperfunctional adenomas of the parathyroid

Os autores apresentam dados clinicos, bioquimicos, radiologicos, cirurgicos, posoperatorios precoces e tardios de 12 casos de hiperparatiroidismo (HPT) (11 primarios e 1 secundario), operados na Secao de Cirurgia Endocrina do HSPE (11 casos) e no Hospital de Itaquera (1 caso). No diagnostico clinico, chamam a atencao para as lesoes osseas presentes em cerca de 83% dos doentes e as renais, em cerca de 66%. Dos exames laboratoriais, a calcemia total encontrava-se elevada em todos os doentes portadores de adenoma (11/11). Em todos os 11 casos de HPT primario, o adenoma localizava-se no pescoco, sendo, em 2, intratiroidianos. Em 7 casos, para facilitar a identificacao do adenoma, foi utilizado o azul de toluidina intravenoso; em 5 casos, o resultado foi excelente; num caso, nao se encontrou o adenoma na regiao cervical, nem no torax; o doente faleceu, sendo encontrado o adenoma dentro da tiroide. Em 10 dos casos de HPT primario, houve regressao do quadro clinico, laboratorial e radiologico apos a cirurgia. O tempo medio de seguimento pos-operatorio foi de 6,8 anos (1 a 13)