Essential oils from the leaves and flowers of Leucophyllum frutescens (Scrophulariaceae): phytochemical analysis and inhibitory effects against elastase and collagenase in vitro.

Leucophyllum frutescens (Scrophulariaceae) is a medicinal plant of Mexican traditional medicine. The aim of this study was to analyse the volatile components from the leaves and flowers by GC/MS and to assess their anti-aging activity for the first time. A total of 30 compounds were identified where 1-octen-3-ol (73.56%) and D-limonene (11.12%) represented the major ingredients in the leaves, while n-heneicosane (32.30%) and dehydroepingaione (15.15%) were the major components in the flowers. In vitro anti-aging activity was measured via assessing collagenase and elastase inhibition. Essential oils from the leaves and flowers showed potential collagenase inhibitory activity with IC50 of 55.7 and 47.4 µg/mL. However, the oils from the leaves and flowers showed moderate anti-elastase activity with IC50 of 60.8 and 97.7 µg/mL. Therefore, the oil of Leucophyllum frutescens could afford a promising natural anti-aging drug.

Effect of low-level laser therapy in an experimental model of osteoarthritis in rats evaluated through Raman spectroscopy.

OBJECTIVE: This work aimed to investigate the biochemical changes associated with low-level laser therapy (LLLT) using 660 and 780 nm, on a well-established experimental model of osteoarthritis (OA) in the knees of rats with induced collagenase, using histomorphometry and Raman spectroscopy. MATERIALS AND METHODS: Thirty-six Wistar rats were divided into four groups: control (GCON, n=9), collagenase without treatment (GCOL, n=9), collagenase with LLLT 660 nm treatment (G660, n=8), and collagenase with LLLT 780 nm treatment (G780, n=10). LLLT protocol was: 30 mW power output, 10 sec irradiation time, 0.04 cm(2) spot size, 0.3 J energy, 0.75 W/cm(2) irradiance, and 7.5 J/cm(2) fluence per session per day, during 14 days. Then, knees were withdrawn and submitted to histomorphometry and Raman spectroscopy analysis. Principal components analysis (PCA) and Mahalanobis distance were employed to characterize the spectral findings. RESULTS: Histomorphometry revealed a significant increase in the amount of collagen III for the group irradiated with 660 nm. The Raman bands at 1247, 1273, and 1453 cm(-1) (from principal component score PC2), attributed to collagen type II, and 1460 cm(-1) (from PC3), attributed to collagen type III, suggested that the LLLT causes acceleration in cellular activity, especially on the cells that repair cartilage, accelerating the breakdown of cartilage destroyed by collagenase and stimulating the fibroblast to synthesize repairing collagen III. CONCLUSIONS: LLLT accelerated the initial breakdown of cartilage destroyed by collagenase and stimulated the fibroblast to synthesize the repairing collagen III, suggesting a beneficial effect of LLLT on OA.

Low-dose UVA1 phototherapy in systemic sclerosis: effects on acrosclerosis.

BACKGROUND: Increased collagen synthesis, vascular damage, and T-lymphocytic infiltration contribute to the development of systemic sclerosis. Preliminary studies revealed the effectiveness of low-dose UVA1 phototherapy in acrosclerosis. OBJECTIVE: We sought to confirm data of a pilot study revealing the efficacy of low-dose UVA1 irradiation in acrosclerosis in a larger number of patients. METHODS: Symptoms of 18 patients receiving low-dose UVA1 phototherapy were evaluated clinically and biometrically in an open, nonrandomized study. A number of pretherapeutic and posttherapeutic biopsy specimens were tested immunohistochemically for matrix-metalloproteinase-1. RESULTS: UVA1 irradiation led to softening of former stiffness reflected by a significant decrease of the hand score, increase of total skin distension, and reduction of skin thickness. Posttherapeutically, matrix-metalloproteinase-1 immunolabeling revealed a significant dermal elevation of collagenase. CONCLUSION: Low-dose UVA1 phototherapy is a capable treatment option for acrosclerosis. Its beneficial effect may be mediated by the induction of collagenases and a reduction of collagen deposition and cellular infiltration.

Cloning of murine membrane-type-1-matrix metalloproteinase (MT-1-MMP) and its metanephric developmental regulation with respect to MMP-2 and its inhibitor.

BACKGROUND: Extracellular matrix macromolecules regulate morphogenesis of embryonic organs, and are developmentally regulated. Their expression and turnover is regulated by matrix metalloproteinases (MMPs). Recently, an epithelial cell "membrane" associated metalloproteinase (MT-1-MMP) has been identified that acts as an activator of a "secreted" MMP-2, and is produced by mesenchymal fibroblasts. The activity of MMP-2 is inhibited by a "soluble" tissue inhibitor of MMP-2, TIMP-2. The role of MT-1-MMP in renal development is unknown. METHODS: MT-1-MMP was cloned from embryonic mouse kidney cDNA library, and its spatio-temporal distribution during development in the context of MMP-2 and tissue inhibitor of metalloproteinase-2 (TIMP-2) was studied. RESULTS: The cloned MT-1-MMP exhibited approximately 86% nucleotide sequence homology with human MT-1-MMP, and had a catalytic domain and a zinc binding site preceded by a RRKR furin recognition motif. A approximately 4.5 Kb MT-1-MMP mRNA transcript was detected, and its expression was developmentally regulated. A parallel developmental regulation of MMP-2 mRNA expression was also observed. TIMP-2 expression was also developmentally regulated, but lagged behind MT-1-MMP and MMP-2. By in situ hybridization, MT-1-MMP mRNA was seen to be confined to ureteric bud epithelia, and was absent in the mesenchyme, while MMP-2 was confined to the mesenchyme. MT-1-MMP protein expression was seen on ureteric bud epithelia, induced mesenchyme and nascent nephrons, and it was highest during mid gestation. Similar spatio-temporal expressions of MMP-2 and TIMP-2 proteins were observed. CONCLUSIONS: mRNAs of MT-MMP-1 and MMP-2 are expressed in the respective epithelial and mesenchymal compartments, while their proteins are co-expressed in the epithelia suggest that MT-1-MMP and MMP-2, in conjunction with TIMP-2, may be involved in paracrine/juxtacrine epithelial:mesenchymal interactions during metanephrogenesis.

[Practical significance of cytokine determination in joint fluid in patients with arthroses or rheumatoid arthritis]. / Praktische Bedeutung der Zytokinbestimmung im Gelenkpunktat von Patienten mit Arthrosen oder rheumatischen Arthritiden.

OBJECTIVE: To determine whether the activity of cartilage-degrading enzymes in the synovial fluid (SF) of patients with rheumatoid arthritis and other joint diseases is correlated with the concentration of cytokines in the SF. METHODS: Cytokines and cartilage-degrading enzymes were determined in the SF of 97 patients with various disorders involving the knee joints (rheumatoid arthritis (RA) n 44; osteoarthritis (OA) n 35; meniscal trauma (Men) n 10; reactive arthritides (ReA) n 8). In these samples we measured the concentrations of interleukin-1 alpha and beta, IL-1-receptor antagonist (IL-1ra), IL-6, IL-8, tumor necrosis factor alpha (TNF alpha; all by ELISA), collagenase-activity and caseinase-activity (by substrate assays). RESULTS: With the exception of IL-1 alpha and IL-6, cytokine-concentrations were significantly higher in RA than in OA SF-samples (p < 0.05; ANOVA on ranks). IL-1ra, IL-6, and IL-1 beta were correlated best with the collagenase-activity in the SF (r = 0.63; 0.57; 0.55; Spearman's rank correlation), while IL-1 beta (r = 0.53) and IL-1ra (r = 0.52) were best correlated with the caseinase-activity in the samples. The SF-concentration of IL-1ra was well correlated with the levels of IL-6, IL-1 beta, II-8, and TNF alpha (r from 0.73 to 0.66; all p < 0.005), but not with IL1 alpha. The molar ratio of IL-1 to IL-1ra in the SF was neither correlated with the activity of collagenase nor caseinase. IL-1 beta and IL-1ra in the SF were positively correlated with the erythrocyte sedimentation rate (ESR). CONCLUSIONS: The determination of IL-1 beta and IL-1ra in the SF of patients with joint disorders as examined in this study seems to allow to a certain extent a prediction of the collagenase- and caseinase-activity contained in the diseased joint. We would favor.

Effects of linoleic acid and gamma-linolenic acid on the growth and metastasis of a human breast cancer cell line in nude mice and on its growth and invasive capacity in vitro.

It has been reported that gamma-linolenic acid (GLA)-rich diets suppress mammary carcinogenesis and transplanted tumor growth and that GLA inhibits the growth of cultured human cancer cell lines. We compared the effects of dietary GLA and linoleic acid (LA) on the growth of MDA-MB-435 human breast cancer cells and their expression of the metastatic phenotype in vivo and in vitro. Athymic nude mice (30/dietary group) were fed isocaloric diets containing 20% (wt/wt) fat but providing 8% GLA or LA for 7 days, and 10(6) tumor cells were then injected into a thoracic mammary fat pad. The diets were continued for a further 11 weeks. The primary tumor growth rates were similar in mice from the two dietary groups; there was a nonstatistically significant trend for the incidence of macroscopic lung metastases and the total lung metastatic volumes to be higher in the GLA-fed mice (79% and 40.1 +/- 13.9 mm3) than in the LA-fed mice (64% and 15.5 +/- 5.4 mm3). The tumor cell phospholipids from the 8% GLA-fed mice contained significantly lower LA levels but higher arachidonic acid levels (both p < 0.001) than those from 8% LA-fed mice. Also the arachidonate-derived eicosanoids (prostaglandin E, leukotriene B4, and 5-, 12-, and 15-hydroxyeicosatetraenoic acids) were significantly higher in tumors from the 8% GLA group. Zymography showed higher 92-kDa type IV collagenase activity in tumors from 8% GLA-fed mice. In vitro, GLA and LA, at 0.5-2 micrograms/ml, stimulated MDA-MB-435 cell growth; 10 micrograms/ml was mildly inhibitory. Whereas LA stimulated tumor cell invasion and 92-kDa type IV collagenase production in vitro, GLA inhibited invasion and did not induce activity of the proteolytic enzyme. Our results do not support the hypothesis that supplementation with GLA would exert a beneficial effect on the progression of an existing breast cancer, perhaps because it is metabolized in vivo to arachidonate-derived eicosanoids that are known to be involved in the metastatic process.