Do arachidonic acid and its metabolites, secreted by rheumatoid and osteoarthritic synovial tissue, account for the strong inhibition of DNA synthesis in cultured human articular chondrocytes? A novel approach to the mechanism of tissue damage

AIMS: To further characterize factors secreted in vitro by osteoarthritic and rheumatoid arthritis synovial membranes that inhibit DNA synthesis by cultured human articular chondrocytes, and extend these findings to synovial fluid. MATERIAL AND METHODS: Synovial tissue, synovial fluid and articular cartilage were obtained at surgery from two patients suffering rheumatoid arthritis and two other patients suffering from osteoarthritis. Synovial tissue was incubated in DMEM, then condition media and synovial fluids were extracted with methanol. Methanol extracts and extracted residues (hyaluronic acid, proteins) were assayed for their capacity to inhibit DNA synthesis in articular chondrocytes. Methanol extracts were also fractionated by thin layer chromatography on silica-coated plates and recovered fractions similarly tested. RESULTS: All extracts exhibited strong and concentration-dependent inhibition of [3H]-thymidine incorporation. The most potent inhibition was obtained with the extracts from rheumatoid joints and the least potent inhibition was with synovial fluids. The removal of active substances with methanol leaves an inactive residue. Methanol extraction does not alter the mitogenic activity of five exogenous growth factors and two cytokines, thus suggesting that such activity is entirely due to lipids. The bulk of anti-mitotic factors extracted by methanol co-migrate when fractionated by thin layer chromatography on silica-coated plates with arachidonic acid and its lipo-oxygenase metabolites. IN CONCLUSION: Inflamed synovium produces and releases lipids, most probably arachidonic acid metabolites that inhibit cell proliferation thus limiting inflammation and pannus formation in arthritis.     

Study on the antioxidant activity of Menthae longifoliae folium and effect of pesticide treatment with Topsin M on the antioxidant activity

This study evaluated both the antioxidant activity of Menthae longifoliae folium and the influence of Topsin M (a pesticide which is widely used nowadays) upon the antioxidant activity of the drug. In this respect, there have been used leaves of Mentha longifolia (L.) Huds., both untreated and treated with Topsin M. The leaves were dried, powdered and extracted with aqueous methanol. Taking into account the fact that polyphenols, and especially flavonoids act as antioxidants, these compounds were quantified in the crude extracts. Both the capacity of inhibition of 15-lipoxygenase and the diphenylpicrylhydrazyl radical-scavenging capacity were also evaluated. The undergone study highlighted the fact that crude extracts of Menthae longifoliae folium had a good antioxidant activity; the extracts were active both as diphenylpicrylhydrazyl radical scavengers and as 15-lipoxygenase inhibitors. Treatment with Topsin M did not consistently influence the antioxidant activity of the drug.     

L'acide arachidonique et ses métabolites, sécrétés par le tissu synovial rhumatoïde et arthrosique, interviennent-ils dans l'inhibition de la synthèse d'ADN par les chondrocytes articulaires humains en culture ? Une nouvelle approche d'analyse des mécanismes des lésions tissulaires

Aims. – To further characterize factors secreted in vitro by osteoarthritic and rheumatoid arthritis synovial membranes that inhibit DNA synthesis by cultured human articular chondrocytes, and extend these findings to synovial fluid.Material and methods. – Synovial tissue, synovial fluid and articular cartilage were obtained at surgery from two patients suffering rheumatoid arthritis and two other patients suffering osteoarthritis. Synovial tissue was incubated in DMEM, then condition media and synovial fluids were extracted with methanol. Methanol extracts and extracted residues (hyaluronic acid, proteins) were assayed for their capacity to inhibit DNA synthesis in articular chondrocytes. Methanol extracts were also fractionated by thin layer chromatography on silica-coated plates and recovered fractions similarly tested.Results. – All extracts exhibited strong and concentration-dependent inhibition of [³H]-thymidine incorporation. The most potent inhibition was obtained with the extracts from rheumatoid joints and the least potent inhibition was with synovial fluids. The removal of active substances with methanol leaves an inactive residue. Methanol extraction does not alter the mitogenic activity of five exogenous growth factors and two cytokines, thus suggesting that such activity is entirely due to lipids. The bulk of antimitotic factors extracted by methanol comigrate when fractionated by thin layer chromatography on silica-coated plates with arachidonic acid and its lipooxygenase metabolites.In conclusion. – Inflamed synovium produces and releases lipids, most probably arachidonic acid metabolites that inhibit cell proliferation thus limiting inflammation and pannus formation in arthritis.     

Action of thymic extracts on the growth of different cell populations in vitro]

Thymic, splenic and renal cellular extracts were obtained by tissular grinding and ultra centrifugation, from animals having undergone diverse hormonal treatments. These extracts were added to different cellular populations in culture (carcinomatous HeLa strain, established MRCs strain, primo-culture of human foetal lung strain). The thymic extracts of non treated animals and the splenic extracts rich in FK substance from intact guinea pigs treated by oestrogenes or FSH, provoque an inhibition of tumoral cell growth. The most important effects are obtained with thymus extracts rich in FK substance. The FK substance seems capable of concentrating certain antitumoral thymic fractions.     

Molecular mode of inhibition of glycogenolysis in rat liver by the dihydropyridine derivative, BAY R3401: inhibition and inactivation of glycogen phosphorylase by an activated metabolite

The racemic prodrug BAY R3401 suppresses hepatic glycogenolysis. BAY W1807, the active metabolite of BAY R3401, inhibits muscle glycogen phosphorylase a and b. We investigated whether BAY R3401 reduces hepatic glycogenolysis by allosteric inhibition or by phosphatase-catalyzed inactivation of phosphorylase. In gel-filtered liver extracts, racemic BAY U6751 (containing active BAY W1807) was tested for inhibition of phosphorylase in the glycogenolytic (in which only phosphorylase a is active) and glycogen-synthetic (for the evaluation of a:b ratios) directions. Phosphorylase inactivation by endogenous phosphatase was also studied. In liver extracts, BAY U6751 (0.9-36 micromol/l) inhibited glycogen synthesis by phosphorylase b (notwithstanding the inclusion of AMP), but not by phosphorylase a. Inhibition of phosphorylase-a-catalyzed glycogenolysis was partially relieved by AMP (500 micromol/l). BAY U6751 facilitated phosphorylase-a dephosphorylation. Isolated hepatocytes and perfused livers were tested for BAY R3401-induced changes in phosphorylase-a:b ratios and glycogenolytic output. Though ineffective in extracts, BAY R3401 (0.25 micromol/l-0.5 mmol/l) promoted phosphorylase-a dephosphorylation in hepatocytes. In perfused livers exposed to dibutyryl cAMP (100 micromol/l) for maximal activation of phosphorylase, BAY R3401 (125 micromol/l) inactivated phosphorylase by 63% but glucose output dropped by 83%. Inhibition of glycogenolysis suppressed glucose-6-phosphate (G6P) levels. Activation of glycogen synthase after phosphorylase inactivation depended on the maintenance of G6P levels by supplementing glucose (50 mmol/l). We conclude that the metabolites of BAY R3401 suppress hepatic glycogenolysis by allosteric inhibition and by the dephosphorylation of phosphorylase a.     

Relation between immunity and survival in carcinoma of the colon and rectum.

Twenty-two patients with cancer of the colon and rectum were evaluated before and after surgical treatment by leukocyte migration inhibition assays using autologous and allogeneic tumor extracts. All patients were resected for cure. Patients with cancer of the colon demonstrated significant migration inhibition against autologous extracts before surgery and in the immediate postoperative period. Patients with rectal cancer did not demonstrate significant migration inhibition at any time. The possibility that immunity plays a role in the poorer prognosis of patients with rectal cancer is discussed.     

Demonstration of a dog liver immunosuppressive factor active in vitro

These studies define the presence of an immunosuppressive factor in dog liver that is active in vitro. A crude dog liver extract was prepared by saline extraction and ultracentrifugation. In vitro immunosuppressive activity of the extract was assayed by its influence on standard phytohemagglutinin mitogen stimulation assays (PHA), two-way mixed lymphocyte culture (MLC) reactions, and primed lymphocyte culture (PLC). In six experiments with four extracts, the mean percentage of inhibition (MPI) of dog lymphocytes in PHA mitogen stimulation assays was 102.0 +/- 3.6. In two human MLC experiments, the extract produced an MPI of 102.3 +/- 0.3. In one canine MLC the MPI was 102.5. In three PLC experiments the MPI was 92.7 +/- 8.0, indicating that the extract inhibited sensitization. Cytotoxicity of the extract was shown not to be the mechanism of proliferation inhibition by restimulation of cells washed free of extract, concurrent eosin viability studies, and documentation of normal base proliferation of cells after extract was washed from them. We concluded that there is a naturally occurring immunosuppressive agent in dog liver that is active in vitro as demonstrated by inhibition of PHA MLC, and PLC cellular proliferation assays. The activity is not attributable to cytotoxicity.     

The effect of essential oils on methicillin-resistant Staphylococcus aureus using a dressing model

Patchouli, tea tree, geranium, lavender essential oils and Citricidal™ (grapefruit seed extract) were used singly and in combination to assess their anti-bacterial activity against three strains of Staphylococcus aureus: Oxford S. aureus NCTC 6571 (Oxford strain), Epidemic methicillin-resistant S. aureus (EMRSA 15) and MRSA (untypable). The individual essential oils, extracts and combinations were impregnated into filter paper discs and placed on the surface of agar plates, pre-seeded with the appropriate strain of Staphylococcus. The effects of the vapours of the oils and oil combinations were also assessed using impregnated filter paper discs that were placed on the underside of the Petri dish lid at a distance of 8 mm from the bacteria. The most inhibitory combinations of oils for each strain were used in a dressing model constructed using a four layers of dressings: the primary layer consisted of either Jelonet™ or TelfaClear™ with or without Flamazine™; the second was a layer of gauze, the third a layer of Gamgee and the final layer was Crepe bandage. The oil combinations were placed in either the gauze or the Gamgee layer. This four-layered dressing was placed over the seeded agar plate, incubated for 24 h at 37 °C and the zones of inhibition measured. All experiments were repeated on three separate occasions. No anti-bacterial effects were observed when Flamazine™ was smeared on the gauze in the dressing model. When Telfaclear™ was used as the primary layer in the dressing model compared to Jelonet™, greater zones of inhibition were observed. A combination of Citricidal™ and geranium oil showed the greatest-anti-bacterial effects against MRSA, whilst a combination of geranium and tea tree oil was most active against the methicillin-sensitive S. aureus (Oxford strain). This study demonstrates the potential of essential oils and essential oil vapours as antibacterial agents and for use in the treatment of MRSA infection.     

具有透皮美白功效的短肽筛选和评价研究

目的：筛选具有透皮性能的短肽并评价其美白功效。方法：采用噬茵体表面展示技术透WiStar大鼠腹部皮肤筛选透皮短肽,以L-多巴为底物,检测短肽对酪氨酸酶活性的抑制作用。体外培养B16小鼠黑素瘤细胞,检测短肽对细胞内黑素生成的抑制作用。以豚鼠为实验对象,考察短肽对豚鼠皮肤内黑素的生成抑制作用。结果：通过噬菌体表面展示技术筛选出4条透皮短肽,其中一条氨基酸序列为ACSSQPPYACG的短肽P1,其对酪氨酸酶活性的半数抑制浓度IC50值为25．64ug／ml,能够明显抑制B16小鼠黑素瘤细胞和豚鼠皮肤内黑素生成。结论：成功建立了利用噬菌体表面展示技术筛选具有美白功效短肽的方法,为美白成分的筛选提供了新的途径。     

Immunological screening of SPARC/Osteonectin in nonmineralized tissues.

SPARC/Osteonectin is a major bone-related protein that is also present in nonmineralized tissues and in platelets. As compared to bone SPARC/Osteonectin, SPARC/Osteonectin from platelets presents a slightly lower electrophoretic mobility in SDS-PAGE and a 100-fold decreased affinity for a unique monoclonal antibody, Mab2 (Malaval et al. 1991). To check the tissular diversity of SPARC/Osteonectin, protein extracts from bovine bone, nonmineralized tissues, and platelets were screened by immunoblotting and immunoradiometric assay, with Mab2 and three other monoclonal antibodies recognizing distinct epitopes. The SPARC/Osteonectin secreted by a human osteosarcoma cell line (MG63) was also tested. In all the nonmineralized tissues tested (gut, bone marrow, tendon, mesentery, artera, lens, skin, liver, and cornea), SPARC/Osteonectin presents the same immunoreactivity and electrophoretic mobility as in bone. The heavier molecular weight and Mab2-negative form present in platelets seems to be unique to this cell type. Osteosarcoma cell extracts and conditioned media give the same results as bone extracts, indicating that the low molecular weight and Mab2-positive form of SPARC/Osteonectin present in most tissues does not result from proteolytic cleavage in the matrix, but is secreted as such. Bone and platelet SPARC/Osteonectin present different patterns of sensitivity to glycosidases, suggestive of a difference in N-glycosylation. However, these treatments do not affect the decreased affinity of Mab2 for platelet SPARC/Osteonectin, which is not likely to be related to difference in N-glycosylation.     

Effect of extracts from the Chinese and European mole cricket on wound epithelialization and neovascularization: in vivo studies in the hairless mouse ear wound model

Until the end of World War II, oily extracts from the European mole cricket, Gryllotalpa gryllotalpa Linné , were used for treating nonhealing wounds and burns. In traditional Chinese medicine, extracts from the Chinese mole cricket, Gryllotalpa africana Beauvois , have been used to treat boils, abscesses, and ulcers successfully for over two centuries and are still being used today. The aim of this study was twofold: first, to measure the effect mole cricket extracts have on wound epithelialization and neovascularization, and second, to identify the active compounds in the Chinese and German mole cricket extracts. For the first aim, the hairless mouse ear wound model was used. The findings showed that wounds treated with the mole cricket extracts epithelialized significantly faster than control wounds 12.7±0.9 and 13.2±1.4 days vs. 16.3±2.2 days (mean±SD, p<0.05), respectively. While the rate of wound neovascularization was significantly increased in the first 3 days postwounding from that point on, the rate in treated wounds was the same as in controls. To identify the active compounds in the mole cricket extracts, the extracts were fractionated and tested in a foreskin basal keratinocyte cell culture assay. In this assay, the migration of keratinocytes is similar to skin cell migration or reepithelialization in a healing wound. Using this method, we found the active compound in the mole cricket extracts to be linoleic acid methyl ester. All other fatty acid structures that were isolated were found to be inactive.     

白花蛇舌草对肿瘤细胞信号传导通路的抑制

研究了白花蛇舌草（Hedyotis diffusa）提取物对肿瘤细胞信号传导通路的影响。首先采取四甲基偶氮唑盐微量酶反应比色法（MTT）法测定各提取物对体外培养人宫颈癌细胞株HeLa增殖的抑制；进一步通过免疫印迹法分析了各提取物对EGF介导的Erk信号传导通路中Erk1／2蛋白磷酸化的影响。结果显示，煎煮及冷水提取物对HeLa细胞的增殖均有抑制，但两者无显著差别，而乙醇提取物显著抑制HeLa细胞增殖。对EGF介导Erk信号传导通路的影响显示。白花蛇舌苹提取物处理细胞90min后，EGF诱导的Erk1/2磷酸化水平显著降低，且表现浓度依赖性。抑制效果：乙醇提取物〉冷水提取物〉煎煮提取物。研究证明。中药白花蛇舌草含有信号传导通路抑制剂，可能在抗肿瘤过程中发挥重要作用。     

The First Stage of Transforming Growth Factor β1 Activation is Release of the Large Latent Complex from the Extracellular Matrix of Growth Plate Chondrocytes by Matrix Vesicle Stromelysin-1 (MMP-3)

Transforming growth factor beta-1 (TGF-beta1) is secreted in a biologically inactive form and stored in the extracellular matrix as a 290 kDa complex consisting of the mature TGF-beta1 homodimer (Mr 25 kDa), the latency-associated peptide (LAP; Mr 75 kDa), and the latent TGF-beta1 binding protein-1 (LTBP1; Mr 190 kDa). Latent TGF-beta1, composed of these three components, is known as the "large latent TGF-beta1 complex." In contrast, latent TGF-beta1 without LTBP1 is known as "small latent TGF-beta1." For all latent forms, dissociation of the TGF-beta1 homodimer from LAP is necessary for growth factor activation and acquisition of biological activity. Matrix vesicles produced by growth plate chondrocytes contain matrix metalloproteinases that can activate small latent TGF-beta1. The enzyme responsible for this is matrix metalloproteinase-3 (MMP-3), although matrix vesicles also contain MMP-2 and plasminogen activator. The present study tested the hypothesis that matrix vesicle enzymes are also involved in the release of the large latent TGF-beta1 complex stored in the extracellular matrix. Matrix vesicles were isolated from cultures of resting zone and growth zone chondrocytes and metalloproteinases present in the matrix vesicles extracted with guanidine-HCl. Chondrocyte extracellular matrices were prepared by lysing confluent cultures and removing the lysed cells. The matrices were incubated with matrix vesicle extracts and the release of total and active TGF-beta1 was determined. To determine if MMP-2 or MMP-3 was involved in the release, matrix vesicle extracts were preincubated with anti-MMP-2 antibody or anti-MMP-3 antibody to selectively deplete the enzyme activity. Matrices were also treated with rhMMP-2 or rhMMP-3. To determine the identity of the released protein(s), digests were separated on SDS-polyacrylamide gels and Western blotting analysis was performed using a specific antibody to LTBP1. Matrix vesicle extracts released both active and total (=latent + active) TGF-beta1 in a time-dependent manner, with peak release after 1 hour of incubation. The amount of total TGF-beta1 released was 10 times higher than the release of active TGF-beta1. The effect of the matrix vesicle extracts was dose-dependent; in addition, the amount and ratio of active to total TGF-b1 released was very similar, irrespective of the source of matrix or matrix vesicle extracts. Pre-incubation of matrix vesicle extracts with anti-MMP-3 antibody blocked the release of active and total TGF-beta1, whereas pre-incubation with pre-immune IgG or anti-MMP-2 antibody had no effect. The addition of rhMMP-3, but not rhMMP-2, caused a dose-dependent increase in the release of total, but not active, TGF-beta1. Western analysis confirmed that both matrix vesicle extracts and rhMMP-3 released the large latent TGF-beta1 complex from the matrix. In addition to the expected 290, 230, and 190 kDa bands, samples run without reduction also contained proteins of molecular weights 110 and 50 kDa that reacted with the anti-LTBP1 antibody. When these same samples were electrophoresed after reduction, the high molecular weight immunoreactive bands disappeared and three bands of molecular weight 75, 32, and 25 kDa were observed. These results indicate that matrix vesicles contain enzymes, especially MMP-3, which are responsible for the release of TGF-beta1 from the matrix, most of which is in latent form. Further, the data suggest that release of the large complex occurs via cleavage at several novel sites in the 130 kDa LTBP1 molecule. Since matrix vesicle MMP-3 is also able to activate small latent TGF-beta1, these results suggest that the large latent TGF-beta1 complex protects against activation of the small latent TGF-beta1. Thus, the data suggest that release of the large latent TGF-bl complex from the matrix and activation of the latent growth factor are only two steps of what must be at least a three-step process.     

Control of glucose metabolism in pancreatic beta-cells by glucokinase, hexokinase, and phosphofructokinase. Model study with cell lines derived from beta-cells

Glucose usage by soluble fractions of cell extracts from two insulin-producing cell lines, RINm5F and HIT, was investigated. Analysis of enzyme activities indicated that glucose phosphorylation and phosphofructokinase are likely to be the rate-limiting steps of glycolysis in both RINm5F and HIT cell extracts. RINm5F extracts, which lack glucokinase, exhibited relatively flat concentration-dependency curves of glucose usage and showed substantial inhibition of hexokinase. HIT cell extracts, which contain glucokinase but lack hexokinase, exhibited sigmoidal concentration-dependency curves of glucose usage, reflecting almost fully expressed glucokinase activity. A reconstituted system prepared from RINm5F and HIT cell extracts exhibited a composite concentration-dependency curve of glucose usage and showed substantial inhibition of hexokinase and almost fully expressed glucokinase. However, conditions that activate phosphofructokinase, such as addition of ammonium sulfate or fructose 2,6-bisphosphate or alkalization, removed the inhibition of hexokinase without noticeably affecting the glucokinase component of usage. Results obtained with a reconstituted system containing RINm5F cell extract and purified glucokinase were consistent with these findings. The data presented here indicate that this reconstituted cell-free system serves as a valid model for the study of aspects of glycolytic control in the islet. This model illustrates the preeminent role of glucokinase in the control of glycolysis, consistent with its glucose-sensor function in the islet. In addition, these studies help to define the contribution of phosphofructokinase to the control of glycolysis and the mechanism whereby changes in phosphofructokinase activity could modulate, via changes in the glucose 6-phosphate concentration, the activity of hexokinase and hence the net glycolytic flux.     

Screening of substances and extracts of plant origin in Walker carcinosarcoma 256]

The authors realized a series of tests with extracts or plants or substances of plant origin in the experimental tumor Walker 256 to determine whether the extracts show anticancer activity. The samples tested were obtained in the authors laboratory or came from other centers. Thirty extracts, 26 of which were inactive and 4 active, were tested. The results shown in the tables are primary screens.     

Botanical Extracts Used in the Treatment of Cellulite

Background. Cellulite is defined as skin relief alterations that give the skin an orange peel or mattress appearance. The lesions tend to be asymptomatic and may be considered the anatomic expressions of the structures in the affected area, such as the fat and subcutaneous septa.
Objective. The present article reviews the most important botanical extracts used as active ingredients in the treatment of cellulite, as well as the steps to obtain these botanicals as raw material and their standardization and quality control, which are important to guarantee their therapeutic action.
Methods. The current literature was reviewed, and we also obtained information from the manufacturers of the prducts that contained botanicals because of the few publications about this subject.
Conclusions. The reduction in fat deposits through the continuous use of anticellulite products depends on the availability of the active ingredient at the action site, the concentration of the ingredient in the formulation, and the physiochemical characteristics particular to each active ingredient. The botanicals used in topical products must have standardized extracts, which would permit each phytomedicine to have the same effect anywhere in the world. New scientific research is necessary to verify the efficacy and ideal concentrations of such substances.
DORIS HEXSEL, MD, CECILIA ORLANDI, MD, AND DEBORA ZECHMEISTER DO PRADO, PHARM, HAVE INDICATED NO SIGNIFICANT INTEREST WITH COMMERCIAL SUPPORTERS.     

104. Zur Frage der Verwendung von Gewebsextrakten in der Krebsbehandlung

Zusammenfassung Für die Therapie maligner Tumoren mit Gewebsextrakten ist eine spezifisch wachstumshemmende Wirkung Voraussetzung. Das in den Untersuchungen Bullough's wirksame sogenannte Chalon schien diese Forderung zu erfüllen. Chalon, eine hypothetische, von differenzierten Zellen gebildete gewebs- aber nicht speciesspezifische Substanz soll konzentrationsabhängig die Mitoseaktivität in Organen und Geweben kontrollieren. Behandlungserfolge nach Verabreichung von Granuloeytenextrakten an Ratten mit chloroleukämisehen Transplantationstumoren, sowie die Regression übertragbarer Melanome bei Hamstern und Mäusen nach Injektion von Schweinehautextrakten wurden zunächst im Sinne einer Chalonwirkung gedeutet. Inzwischen ist es jedoch gelungen, aus derartigen Extrakten Mikroorganismen (Clostridien) zu isolieren und ihre onkolytischen Eigenschaften an zahlreichen Transplantationstumoren nachzuweisen. Diese Wirkung pathogener und apathogener Anaerobier ist seit den Arbeiten von Möse bekannt. Die aus in vitro-Befunden abgeleitete Theorie einer gewebsspezifischen Wachstumskontrolle hat für die Anwendung von Gewebsextrakten in der Krebsbehandlung heute noch keine Bedeutung.

---

The use of tissue extracts in the treatment of cancer

Summary An optimal treatment of neoplasia presupposes a specific inhibition of tumor proliferation. Experiments with tissue extracts carried out mainly by Bullough seemed to suggest a fulfilment of this postulate, for he supposed a tissue-but not species-specific control of the mitotic activity in organs and tissues by an internal secretion product, the chalone. Decrease of cell production in a rat chloroleukaemia by means of granulocytic extract and also melanoma regression after treatment of tumor-bearing hamsters and mice with pig epidermis extracts were explained to be the result of chalone action. In the meantime onkolytic Clostridia have been demonstrated in the active extracts. The tumor-destroying effect of similar anaerobic spores has already been established by the work of Möse. From the above mentioned findings it remains doubtful whether the theoretical defined inhibition of mitotic activity by tissue extracts will be significant for the treatment of cancer.

     

山药-黄芪药对治疗2型糖尿病性骨质疏松的网络药理学机制研究

目的 筛选山药-黄芪药对的化学成分并预测其作用靶点,建立药物活性成分-作用靶点-信号通路网络,分析山药-黄芪药对治疗2型糖尿病性骨质疏松(type 2 diabetic osteoporosis,T2DOP)的作用机制.方法 通过中药系统药理数据库和分析平台(TCMSP)检索山药、黄芪的所有潜在活性成分及其对应的作用靶...     

Novel small molecule inhibitors for prostate-specific antigen

BACKGROUND: Prostate-specific antigen (PSA or KLK3) has been shown to inhibit angiogenesis, but it might also have tumor promoting activities. Thus, it may be possible to modulate prostate cancer growth by stimulating or inhibiting the activity of PSA. To this end we have previously identified peptides that stimulate the activity of PSA. As peptides have several limitations as drug molecules, we screened a chemical library to find drug-like compounds that could be used to modulate the function(s) of PSA. METHODS: Almost 50,000 compounds were analyzed for their ability to modulate PSA activity towards a fluorescent PSA-substrate. The ability of the most active compounds to affect the anti-angiogenic activity of PSA was analyzed by human umbilical vein endothelial cell (HUVEC) tube formation assay. RESULTS: In the initial screening we identified two compounds that inhibited PSA activity. Based on these, similar compounds were selected and tested for activity to define structure-activity relationships. Several compounds with micromolar IC50-values were found, but they were not entirely specific towards PSA, e.g., they inhibited chymotrypsin, which has similar substrate specificity as PSA. However, it was possibly to improve the selectivity of the compounds towards PSA by small structural changes. These compounds inhibited the anti-angiogenic activity of PSA in the HUVEC model, proving that the proteolytic activity of PSA is essential for inhibition of angiogenesis. CONCLUSIONS: We found several PSA inhibitors that could be useful tools for studying the role of PSA in cancer models and in normal physiology as showed in angiogenesis model.