Application of Propolis in Protecting Skeletal and Periodontal Health—A Systematic Review

Chronic inflammation and oxidative stress are two major mechanisms leading to the imbalance between bone resorption and bone formation rate, and subsequently, bone loss. Thus, functional foods and dietary compounds with antioxidant and anti-inflammatory could protect skeletal health. This review aims to examine the current evidence on the skeletal protective effects of propolis, a resin produced by bees, known to possess antioxidant and anti-inflammatory activities. A literature search was performed using Pubmed, Scopus, and Web of Science to identify studies on the effects of propolis on bone health. The search string used was (i) propolis AND (ii) (bone OR osteoporosis OR osteoblasts OR osteoclasts OR osteocytes). Eighteen studies were included in the current review. The available experimental studies demonstrated that propolis could prevent bone loss due to periodontitis, dental implantitis, and diabetes in animals. Combined with synthetic and natural grafts, it could also promote fracture healing. Propolis protects bone health by inhibiting osteoclastogenesis and promoting osteoblastogenesis, partly through its antioxidant and anti-inflammatory actions. Despite the promising preclinical results, the skeletal protective effects of propolis are yet to be proven in human studies. This research gap should be bridged before nutraceuticals based on propolis with specific health claims can be developed.     

Biological Effects of β-Glucans on Osteoclastogenesis

Although the anti-tumor and anti-infective properties of β-glucans have been well-discussed, their role in bone metabolism has not been reviewed so far. This review discusses the biological effects of β-glucans on bone metabolisms, especially on bone-resorbing osteoclasts, which are differentiated from hematopoietic precursors. Multiple immunoreceptors that can recognize β-glucans were reported to be expressed in osteoclast precursors. Coordinated co-stimulatory signals mediated by these immunoreceptors are important for the regulation of osteoclastogenesis and bone remodeling. Curdlan from the bacterium Alcaligenes faecalis negatively regulates osteoclast differentiation in vitro by affecting both the osteoclast precursors and osteoclast-supporting cells. We also showed that laminarin, lichenan, and glucan from baker’s yeast, as well as β-1,3-glucan from Euglema gracilisas, inhibit the osteoclast formation in bone marrow cells. Consistent with these findings, systemic and local administration of β-glucan derived from Aureobasidium pullulans and Saccharomyces cerevisiae suppressed bone resorption in vivo. However, zymosan derived from S. cerevisiae stimulated the bone resorption activity and is widely used to induce arthritis in animal models. Additional research concerning the relationship between the molecular structure of β-glucan and its effect on osteoclastic bone resorption will be beneficial for the development of novel treatment strategies for bone-related diseases.     

Biosynthesis, total syntheses, and antitumor activity of tanshinones and their analogs as potential therapeutic agents

Tanshinones are a series of abietane diterpenes, isolated exclusively from Salvia miltiorrhiza and related species. More than 40 tanshinones and their analogs have been isolated since the 1930s. Their biosynthetic pathway correlates with the MEP/DOXP pathway, and many key enzymes, such as mCPS, are responsible for establishing their molecular scaffolds and stereospeci?city. Because of their unique structural characteristics and promising biological activities, total syntheses of various tanshinones have attracted the interest of many synthetic chemists, including R. H. Thomson, H. Kakisawa, R. L. Danheiser, Y. Inouye and J. K. Snyder. Tanshinones and their analogs exhibit interesting and broad antitumor activity in various cell and animal models. Most recently, the tanshinone analog neo-tanshinlactone has shown potent and selective activity against breast cancer. This review will discuss the biosynthesis, total syntheses, and antitumor activities of tanshinones,especially neo-tanshinlactone and its analogs.     

β-Boswellic Acid Inhibits RANKL-Induced Osteoclast Differentiation and Function by Attenuating NF-κB and Btk-PLCγ2 Signaling Pathways

Osteoporosis is a systemic metabolic bone disorder that is caused by an imbalance in the functions of osteoclasts and osteoblasts and is characterized by excessive bone resorption by osteoclasts. Targeting osteoclast differentiation and bone resorption is considered a good fundamental solution for overcoming bone diseases. β-boswellic acid (βBA) is a natural compound found in Boswellia serrata, which is an active ingredient with anti-inflammatory, anti-rheumatic, and anti-cancer effects. Here, we explored the anti-resorptive effect of βBA on osteoclastogenesis. βBA significantly inhibited the formation of tartrate-resistant acid phosphatase-positive osteoclasts induced by receptor activator of nuclear factor-B ligand (RANKL) and suppressed bone resorption without any cytotoxicity. Interestingly, βBA significantly inhibited the phosphorylation of IκB, Btk, and PLCγ2 and the degradation of IκB. Additionally, βBA strongly inhibited the mRNA and protein expression of c-Fos and NFATc1 induced by RANKL and subsequently attenuated the expression of osteoclast marker genes, such as OC-STAMP, DC-STAMP, β3-integrin, MMP9, ATP6v0d2, and CtsK. These results suggest that βBA is a potential therapeutic candidate for the treatment of excessive osteoclast-induced bone diseases such as osteoporosis.     

Deficiency of optineurin enhances osteoclast differentiation by attenuating the NRF2-mediated antioxidant response

Abnormally increased resorption contributes to bone degenerative diseases such as Paget’s disease of bone (PDB) through unclear mechanisms. Recently, the optineurin (OPTN) gene has been implicated in PDB, and global OPTN knockout mice (Optn^−/−) were shown to exhibit increased formation of osteoclasts (osteoclastogenesis). Growing evidence, including our own, has demonstrated that intracellular reactive oxygen species (ROS) stimulated by receptor activator of nuclear factor kappa-B ligand (RANKL) can act as signaling molecules to promote osteoclastogenesis. Here, we report that OPTN interacts with nuclear factor erythroid-derived factor 2-related factor 2 (NRF2), the master regulator of the antioxidant response, defining a pathway through which RANKL-induced ROS could be regulated for osteoclastogenesis. In this study, monocytes from Optn^−/− and wild-type (Optn^+/+) mice were utilized to differentiate into osteoclasts, and both qRT-PCR and tartrate-resistant acid phosphatase (TRAP) staining showed that the Optn^−/− monocytes exhibited enhanced osteoclastogenesis compared to the Optn^+/+ cells. CellROX^® staining, qRT-PCR, and Western blotting indicated that OPTN deficiency reduced the basal expression of Nrf2, inhibited the expression of NRF2-responsive antioxidants, and increased basal and RANKL-induced intracellular ROS levels, leading to enhanced osteoclastogenesis. Coimmunoprecipitation (co-IP) showed direct interaction, and immunofluorescence staining showed perinuclear colocalization of the OPTN-NRF2 granular structures during differentiation. Finally, curcumin and the other NRF2 activators attenuated the hyperactive osteoclastogenesis induced by OPTN deficiency. Collectively, our findings reveal a novel OPTN-mediated mechanism for regulating the NRF2-mediated antioxidant response in osteoclasts and extend the therapeutic potential of OPTN in the aging process resulting from ROS-triggered oxidative stress, which is associated with PDB and many other degenerative diseases.Subject terms: Mechanisms of disease, Stress signalling     

Characterization of tanshinones in the roots of Salvia miltiorrhiza (Dan-shen) by high-performance liquid chromatography with electrospray ionization tandem mass spectrometry

The qualitative analysis of tanshinones in the roots of Salvia miltiorrhiza (Dan-shen in Chinese) was performed using high-performance liquid chromatography with electrospray ionization tandem mass spectrometry (ESI-MS(n)). Tanshinones are the major bioactive constituents of Dan-shen, which is used in China for the treatment of haematological abnormalities and cardiovascular diseases. The ESI-MS(n) fragmentation behavior of tanshinones was investigated. For tanshinones with the tanshinone I nucleus, the fragmentation was triggered by loss of a molecule of CO except bearing a substituent at C17 or C18, followed by sequential eliminations of CO. If C(15-16) was a saturated bond, the fragmentation was triggered by elimination of a molecule of H2O. For tanshinones with the tanshinone IIA nucleus, the fragmentation was triggered by loss of a molecule of H2O, followed by successive eliminations of CO. Ions corresponding to loss of a molecule of propylene (Delta m = 42) were also observed. Moreover, when C(15-16) was a saturated bond, ions corresponding to losses of CH3, H2O and propylene were more abundant. If no D-ring existed, the presence of isopropyl resulted in an elimination of a molecule of H2O with an adjacent CO or OH. In addition, the extension of the pi-conjugation in the A-ring (especially at C(1-2)) induced the fragmentation by loss of a molecule of CO. These fragmentation rules were applied to the identification of tanshinones in a chloroform/methanol (3:7) extract of Dan-shen, which was separated on a C18 column with gradient elution. A total of 27 tanshinones were identified, including five new constituents. The established method could be used for the sensitive and rapid identification of tanshinones in the Dan-shen drug and its pharmaceutical preparations.     

A triterpene glycoside from black cohosh that inhibits osteoclastogenesis by modulating RANKL and TNFalpha signaling pathways

Osteoporosis is a major age-related source of morbidity and mortality. Increased bone resorption mediated by osteoclasts is central to its pathogenesis. Cytokines, particularly RANKL and TNFalpha, are often increased under pathologic conditions, leading to enhanced osteoclastogenesis. Black cohosh (Actaea/Cimicifuga racemosa L), a popular herbal supplement for the treatment of menopausal symptoms, was recently shown to have the beneficial effect of preventing bone loss. Here, we demonstrate that 25-acetylcimigenol xylopyranoside (ACCX), a triterpenoid glycoside isolated from black cohosh, potently blocks in vitro osteoclastogenesis induced by either RANKL or TNFalpha. This blockage of osteoclastogenesis elicited by ACCX results from abrogation of the NF-kappaB and ERK pathways induced by either RANKL or TNFalpha, respectively. Importantly, this compound attenuates TNFalpha-induced bone loss in vivo. Therefore, ACCX represents a potential lead for the development of a new class of antiosteoporosis agents.     

Simvastatin inhibits osteoclast differentiation by scavenging reactive oxygen species

Osteoclasts, together with osteoblasts, control the amount of bone tissue and regulate bone remodeling. Osteoclast differentiation is an important factor related to the pathogenesis of bone-loss related diseases. Reactive oxygen species (ROS) acts as a signal mediator in osteoclast differentiation. Simvastatin, which inhibits 3-hydroxy-3-methylglutaryl coenzyme A, is a hypolipidemic drug which is known to affect bone metabolism and suppresses osteoclastogenesis induced by receptor activator of nuclear factor-κB ligand (RANKL). In this study, we analyzed whether simvastatin can inhibit RANKL-induced osteoclastogenesis through suppression of the subsequently formed ROS and investigated whether simvastatin can inhibit H2O2-induced signaling pathways in osteoclast differentiation. We found that simvastatin decreased expression of tartrate-resistant acid phosphatase (TRAP), a genetic marker of osteoclast differentiation, and inhibited intracellular ROS generation in RAW 264.7 cell lines. ROS generation activated NF-κB, protein kinases B (AKT), mitogen-activated protein kinases signaling pathways such as c-JUN N-terminal kinases, p38 MAP kinases as well as extracellular signal- regulated kinase. Simvastatin was found to suppress these H2O2-induced signaling pathways in osteoclastogenesis. Together, these results indicate that simvastatin acts as an osteoclastogenesis inhibitor through suppression of ROS-mediated signaling pathways. This indicates that simvastatin has potential usefulness for osteoporosis and pathological bone resorption.     

High extracellular Ca2+ alone stimulates osteoclast formation but inhibits in the presence of other osteoclastogenic factors

High ambient Ca2+ at bone resorption sites have been implicated to play an important role in the regulation of bone remodeling. The present study was performed to clarify the mode of high extracellular Ca2+ (Ca2+(e))-induced modulation of osteoclastogenesis and the expression of receptor activator of nuclear factor-kappaB ligand (RANKL) and osteoprotegerin (OPG), thereby to define its role in osteoclast formation. Mouse bone marrow cells were cocultured with osteoblastic cells in the absence or presence of osteoclastogenic factors such as 1,25-dihydroxyvitaminD3 (1,25-(OH)2vitD3)and macrophage colony-stimulating factor/soluble RANKL. Ca2+ concentration in media (1.8 mM) was adjusted to 3, 5, 7 or 10 mM. Osteoclast formation was confirmed by the appearance of tartrate-resistant acid phosphatase (TRAP)-positive multinuclear cells and the expression of osteoclast phenotypic markers (calcitonin receptor, vitronectin receptor, cathepsin K, matrix metalloproteinase-9, carbonic anhydrase 2). High Ca2+(e) alone significantly stimulated osteoclast formation in a dose-dependent manner. However, in the presence of highly osteoclastogenic factors, high Ca2+(e) significantly inhibited osteoclastogenesis. High Ca2+(e) alone continuously up-regulated RANKL expression while only transiently increased OPG expression. However, in the presence of 1,25-(OH)(2)vitD(3), high Ca2+(e) did not change the 1,25-(OH)2vitD3-induced RANKL expression while increased OPG expression. Taken together, these findings suggest that high Ca2+(e) alone increase osteoclastogenesis but inhibit in the presence of other osteoclastogenic factors. In addition, high CaCa2+(e)-induced osteoclastogenesis may be mediated by osteoblasts via up-regulation of RANKL expression. Meanwhile up-regulated OPG might participate in the inhibitory effect of high Ca2+(e) on 1,25-(OH)2vitD3-induced osteoclastogenesis.     

Ethanol Extract of Amomum tsao-ko Ameliorates Ovariectomy-Induced Trabecular Loss and Fat Accumulation

In Asia, Amomum tsao-ko has long been used as a spice or seasoning in food to stimulate digestion. In the present study, we evaluated the effects of ethanol extract of Amomum tsao-ko (EEAT) on menopausal osteoporosis and obesity. After the administration of EEAT in ovariectomy (OVX) mice models for five weeks, microcomputed tomography and a histological analysis were performed to assess, respectively, the trabecular structure and the fat accumulation in adipose, liver, and bone tissues. We also examined the effects of EEAT on a bone marrow macrophage model of osteoclastogenesis by in vitro stimulation from the receptor activator of nuclear factor-kappa Β ligand (RANKL) through real-time PCR and Western blot analysis. In addition, ultrahigh performance liquid chromatography–tandem mass spectrometry (UHPLC–MS/MS) with authentic standards was applied to characterize the phytochemical profiling of EEAT. We found that EEAT significantly decreased OVX-induced body weight gain and fat accumulation, significantly prevented OVX-induced deterioration of bone mineral density and microstructure of trabecular tissues, and significantly inhibited osteoclast differentiation by downregulating NF-κB/Fos/NFATc1 signaling in osteoclasts. Furthermore, UHPLC–MS/MS identified eight beneficial phytochemicals in EEAT. Collectively, these results suggest that EEAT might be an effective nutraceutical candidate to attenuate menopausal osteoporosis by inhibiting osteoclastogenesis and to prevent obesity by suppressing fat accumulation.     

New Monoterpenoid Indoles with Osteoclast Activities from Gelsemium elegans

The well-known toxic medicine Gelsemium elegans is widely and historically used to treat bone fracture and skin ulcers by the folk people of China. Two new monoterpenoid indole alkaloids, gelselegandines D and E, together with the known analogue gelegamine A were isolated from G. elegans. Their structures were elucidated by means of spectroscopic techniques and quantum chemical calculations. All isolated compounds were tested for the effects on RANKL-induced osteoclast formation. Interestingly, gelselegandine E and gelegamine A, respectively, showed significant promoting and inhibitory activities on osteoclastogenesis, while gelselegandine D had no activity under the same concentration. This work suggested the different configurations for the carbons near the C-19/20 oxygen rings of the isolated compounds may be the key active groups on osteoclast formation and provided the evidence for the rationality as the traditional treatment for bone-related diseases of G. elegans.     

Inhibitory Effect of LGS and ODE Isolated from the Twigs of Syringa oblata subsp. dilatata on RANKL-Induced Osteoclastogenesis in Macrophage Cells

Osteoblasts and osteoclasts play a pivotal role in maintaining bone homeostasis, of which excessive bone resorption by osteoclasts can cause osteoporosis and various bone diseases. However, current osteoporosis treatments have many side effects, and research on new treatments that can replace these treatments is ongoing. Therefore, in this study, the roles of ligustroside (LGS) and oleoside dimethylester (ODE), a natural product-derived compound isolated from Syringa oblata subsp. dilatata as a novel, natural product-derived osteoporosis treatments were investigated. In the results of this study, LGS and ODE inhibited the differentiation of receptor activator of nuclear factor kappa-Β ligand (RANKL)-induced RAW264.7 cells into osteoclasts without cytotoxicity, and down-regulated the activity of TRAP, a specific biomarker of osteoclasts. In addition, it inhibited bone resorption and actin ring formation, which are important functions and features of osteoclasts. Also, the effects of LGS and ODE on the mitogen-activated protein kinase (MAPK) and nuclear factor kappa-light-chain-enhancer of activated B (NF-κB) and phosphoinositide 3-kinases (PI3K)/ protein kinase B (Akt)/mechanistic target of rapamycin (mTOR) signaling pathways that play important roles in osteoclast differentiation were evaluated. In the results, LGS and ODE downregulated the phosphorylation of RANKL-induced MAPK and PI3K/Akt/mTOR proteins in a concentration-dependent manner, translocation of NF-κB into the nucleus was inhibited. As a result, the compounds LGS and ODE isolated from S. oblate subsp. dilatata effectively regulated the differentiation of RANKL-induced osteoclasts and inhibited the phosphorylation of signaling pathways that play a pivotal role in osteoclast differentiation. Therefore, these results suggest the possibility of LGS and ODE as new natural product treatments for bone diseases caused by excessive osteoclasts.     

A-type cranberry proanthocyanidins inhibit the RANKL-dependent differentiation and function of human osteoclasts

This study investigated the effect of A-type cranberry proanthocyanidins (AC-PACs) on osteoclast formation and bone resorption activity. The differentiation of human pre-osteoclastic cells was assessed by tartrate-resistant acid phosphatase (TRAP) staining, while the secretion of interleukin-8 (IL-8) and matrix metalloproteinases (MMPs) was measured by ELISA. Bone resorption activity was investigated by using a human bone plate coupled with an immunoassay that detected the release of collagen helical peptides. AC-PACs up to 100 μg/mL were atoxic for osteoclastic cells. TRAP staining evidenced a dose-dependent inhibition of osteoclastogenesis. More specifically, AC-PACs at 50 μg/mL caused a 95% inhibition of RANKL-dependent osteoclast differentiation. This concentration of AC-PACs also significantly increased the secretion of IL-8 (6-fold) and inhibited the secretion of both MMP-2 and MMP-9. Lastly, AC-PACs (10, 25, 50 and 100 μg/ml) affected bone degradation mediated by mature osteoclasts by significantly decreasing the release of collagen helical peptides. This study suggests that AC-PACs can interfere with osteoclastic cell maturation and physiology as well as prevent bone resorption. These compounds may be considered as therapeutic agents for the prevention and treatment of periodontitis.     

The Biological Activities and Therapeutic Potentials of Baicalein Extracted from Oroxylum indicum: A Systematic Review

In Southeast Asia, traditional medicine has a longestablished history and plays an important role in the health care system. Various traditional medicinal plants have been used to treat diseases since ancient times and much of this traditional knowledge remains preserved today. Oroxylum indicum (beko plant) is one of the medicinal herb plants that is widely distributed throughout Asia. It is a versatile plant and almost every part of the plant is reported to possess a wide range of pharmacological activities. Many of the important bioactivities of this medicinal plant is related to the most abundant bioactive constituent found in this plant—the baicalein. Nonetheless, there is still no systematic review to report and vindicate the biological activities and therapeutic potential of baicalein extracted from O. indicum to treat human diseases. In this review, we aimed to systematically present in vivo and in vitro studies searched from PubMed, ScienceDirect, Scopus and Google Scholar database up to 31 March 2020 based on keywords “Oroxylum indicum” and “baicalein”. After an initial screening of titles and abstracts, followed by a full-text analysis and validation, 20 articles that fulfilled all the inclusion and exclusion criteria were included in this systematic review. The searched data comprehensively reported the biological activities and therapeutic potential of baicalein originating from the O. indicum plant for anti-cancer, antibacterial, anti-hyperglycemia, neurogenesis, cardioprotective, anti-adipogenesis, anti-inflammatory and wound healing effects. Nonetheless, we noticed that there was a scarcity of evidence on the efficacy of this natural active compound in human clinical studies. In conclusion, this systematic review article provides new insight into O. indicum and its active constituent baicalein as a prospective complementary therapy from the perspective of modern and scientific aspect. We indicate the potential of this natural product to be developed into more conscientious and judicious evidencebased medicine in the future. However, we also recommend more clinical research to confirm the efficacy and safety of baicalein as therapeutic medicine for patients.     

Profiling the metabolic difference of seven tanshinones using high-performance liquid chromatography/multi-stage mass spectrometry with data-dependent acquisition

Tanshinones are a class of bioactive constituents in the roots of Salvia miltiorrhiza named Dan-Shen in Chinese, which possess diverse pharmacological activities. In this study, we employed a sensitive high-performance liquid chromatography/multi-stage mass spectrometry (HPLC/MS(n)) method with data-dependent acquisition and a dynamic exclusion program for the identification of phase I metabolites of seven tanshinones in rat bile after intravenous administration. These seven tanshinones are tanshinone IIA, sodium tanshinone IIA sulfonate (abbreviated as STS, a water-soluble derivate of tanshinone IIA), cryptotanshinone, 15,16-dihydrotanshinone I, tanshinone IIB, przewaquinone A and tanshinone I. Altogether 33 metabolites underwent monohydroxylation, dihydroxylation, dehydrogenation, D-ring hydrolysis or oxidation reactions in the C-4 or C-15 side chain which were characterized by analyzing the LC/MS(n) data. Different metabolic reactions for tanshinones were dependent on the degree of saturation and the substituent group in the skeleton. Dehydrogenation was the major metabolic modification for cryptotanshinone with saturated A and D rings. 15,16-Dihydrotanshinone I containing a saturated D ring was mainly metabolized through D-ring hydrolysis. For tanshinone IIA, possessing a saturated A ring, hydroxylation was the major metabolic pathway. When there was hydroxyl group substitution in the C-17 or C-18 position, such as przewaquinone A and tanshinone IIB, or sulfonic group substitution in the C-16 position, such as STS, higher metabolic stability than that of tanshinone IIA was shown and only trace metabolites were generated. Oxidation in the C-4 or C-15 side chain was a characteristic reaction for tanshinone IIA and hydroxylated tanshinone IIA. For tanshinone I, bearing unsaturated A and D rings simultaneously, no metabolites were detected.     

Eleutherococcus sessiliflorus Inhibits Receptor Activator of Nuclear Factor Kappa-B Ligand (RANKL)-Induced Osteoclast Differentiation and Prevents Ovariectomy (OVX)-Induced Bone Loss

The aim of this study was to evaluate the effects of root bark of Eleutherococcus sessiliflorus (ES) on osteoclast differentiation and function in vitro and in vivo. In vitro, we found that ES significantly inhibited the RANKL-induced formation of TRAP-positive multinucleated osteoclasts and osteoclastic bone resorption without cytotoxic effects. ES markedly downregulated the expression of nuclear factor of activated T cells cytoplasmic 1 (NFATc1); c-Fos; and osteoclast-related marker genes, such as TRAP, osteoclast-associated receptor (OSCAR), matrix metalloproteinase-9 (MMP-9), calcitonin receptor, cathepsin K, the 38 kDa d2 subunit of the vacuolar H⁺-transporting lysosomal ATPase (Atp6v0d2), dendritic cell-specific transmembrane protein (DC-STAMP), and osteoclast-stimulatory transmembrane protein (OC-STAMP). These effects were achieved by inhibiting the RANKL-mediated activation of MAPK signaling pathway proteins, including p38, ERK, and JNK. In vivo, ES attenuated OVX-induced decrease in bone volume to tissue volume ratio (BV/TV), trabecular thickness (Tb.Th), trabecular number (Tb.N), and bone mineral density, but increased trabecular separation (Tb.Sp) in the femur. Collectively, our findings showed that ES inhibited RANKL-activated osteoclast differentiation in bone marrow macrophages and prevented OVX-mediated bone loss in rats. These findings suggest that ES has the potential to be used as a therapeutic agent for bone-related diseases, such as osteoporosis.     

Protease Produced by Endophytic Fungi: A Systematic Review

The purpose of this systematic review was to identify the available literature of production, purification, and characterization of proteases by endophytic fungi. There are few complete studies that entirely exhibit the production, characterization, and purification of proteases from endophytic fungi. This study followed the PRISMA, and the search was conducted on five databases: PubMed, PMC, Science Direct, Scopus Articles, and Web of Science up until 18 May 2021, with no time or language restrictions. The methodology of the selected studies was evaluated using GRADE. Protease production, optimization, purification, and characterization were the main evaluated outcomes. Of the 5540 initially gathered studies, 15 met the inclusion criteria after a two-step selection process. Only two studies optimized the protease production using statistical design and two reported enzyme purification and characterization. The genus Penicillium and Aspergillus were the most cited among the eleven different genera of endophytic fungi evaluated in the selected articles. Six studies proved the ability of some endophytic fungi to produce fibrinolytic proteases, demonstrating that endophytic fungi can be exploited for the further production of agents used in thrombolytic therapy. However, further characterization and physicochemical studies are required to evaluate the real potential of endophytic fungi as sources of industrial enzymes.     

Micelle-mediated extraction of tanshinones from Salvia miltiorrhiza bunge with analysis by high-performance liquid chromatography

The feasibility of employing non-ionic surfactant oligoethylene glycol monoalkyl ether (Genapol X-080) as an alternative and effective solvent for the extraction of tanshinones from Salvia miltiorrhiza bunge was studied for the first time. Various experimental conditions were investigated to optimize the extraction. Under optimum conditions, i.e. 10% Genapol X-080 (w/v), liquid/solid ratio of 20:1 (ml g⁻¹), ultrasonic-assisted extraction for 45 min, the extraction recovery of the tanshinones reached the highest value. When compared with commonly used solvents, 10% Genapol X-080 yielded almost the same extraction efficiency as methanol and dichloromethane-methanol (1:4). For the pre-concentration of tanshinones by cloud-point extraction (CPE), sodium chloride was added to the solution to facilitate the phase separation and increase the pre-concentration factor by reducing the volume of the surfactant-rich phase.     

Ubiquitin-mediated signalling and Paget's disease of bone

Multiple steps in the RANK-NF-kappaB signalling pathway are regulated by ubiquitylation. Mutations affecting different components of this pathway, including the ubiquitin binding p62 signalling adapter protein, are found in patients with Paget's disease of bone or related syndromes. Here, we review the molecular defects and potential disease mechanisms in these conditions and conclude that the mutations may confer a common increased sensitivity of osteoclasts to cytokines, resulting in disordered NF-kappaB-dependent osteoclast function. Modulation of the osteoclast RANK-NF-kappaB signalling axis may represent a viable therapeutic strategy for Paget's disease and other conditions where excessive bone resorption or remodelling is a feature. Publication history: Republished from Current BioData's Targeted Proteins database (TPdb; http://www.targetedproteinsdb.com).