Proteomics profiling of cholangiocarcinoma exosomes: A potential role of oncogenic protein transferring in cancer progression.

Cholangiocarcinoma (CCA), a common primary malignant tumor of bile duct epithelia, is highly prevalent in Asian countries and unresponsive to chemotherapeutic drugs. Thus, a newly recognized biological entity for early diagnosis and treatment is highly needed. Exosomes are small membrane bound vesicles found in body fluids and released by most cell types including cancer cells. The vesicles contain specific subset of proteins and nucleic acids corresponding to cell types and play essential roles in pathophysiological processes. The present study aimed to assess the protein profiles of CCA-derived exosomes and their potential roles. We have isolated exosomes from CCA cells namely KKU-M213 and KKU-100 derived from Thai patients and their roles were investigated by incubation with normal human cholangiocyte (H69) cells. Exosomes were internalized into H69 cells and had no effects on viability or proliferation of the host cells. Interestingly, the exosomes from KKU-M213 cells only induced migration and invasion of H69 cells. Proteomic analysis of the exosomes from KKU-M213 cells disclosed multiple cancer related proteins that are not present in H69 exosomes. Consistent with the protein profile, treatment with KKU-M213 exosomes induced ß-catenin and reduced E-cadherin expressions in H69 cells. Collectively, our results suggest that a direct cell-to-cell transfer of oncogenic proteins via exosomal pathway may be a novel mechanism for CCA progression and metastasis.

Discovery, isolation, and structural characterization of cyclotides from Viola sumatrana Miq.

Cyclotides are cyclic peptides from plants in the Violaceae, Rubiaceae, Fabaceae, Cucurbitaceae, and Solanaceae families. They are sparsely distributed in most of these families, but appear to be ubiquitous in the Violaceae, having been found in every plant so far screened from this family. However, not all geographic regions have been examined and here we report the discovery of cyclotides from a Viola species from South-East Asia. Two novel cyclotides (Visu 1 and Visu 2) and two known cyclotides (kalata S and kalata B1) were identified in V. sumatrana. NMR studies revealed that kalata S and kalata B1 had similar secondary structures. Their biological activities were determined in cytotoxicity assays; both had similar cytotoxic activity and were more toxic to U87 cells compared with other cell lines. Overall, the study strongly supports the ubiquity of cyclotides in the Violaceae and adds to our understanding of their distribution and cytotoxic activity.

Comparative protein profiles of Butea superba tubers under seasonal changes.

Seasonal changes are major factors affecting environmental conditions which induce multiple stresses in plants, leading to changes in protein relative abundance in the complex cellular plant metabolic pathways. Proteomics was applied to study variations in proteome composition of Butea. superba tubers during winter, summer and rainy season throughout the year using two-dimensional polyacrylamide gel electrophoresis coupled with a nanoflow liquid chromatography coupled to electrospray ionization quadrupole-time-of-flight tandem mass spectrometry. A total of 191 protein spots were identified and also classified into 12 functional groups. The majority of these were mainly involved in carbohydrate and energy metabolism (30.37 %) and defense and stress (18.32 %). The results exhibited the highest numbers of identified proteins in winter-harvested samples. Forty-five differential proteins were found in different seasons, involving important metabolic pathways. Further analysis indicated that changes in the protein levels were due mainly to temperature stress during summer and to water stress during winter, which affected cellular structure, photosynthesis, signal transduction and homeostasis, amino-acid biosynthesis, protein destination and storage, protein biosynthesis and stimulated defense and stress mechanisms involving glycolytic enzymes and relative oxygen species catabolizing enzymes. The proteins with differential relative abundances might induce an altered physiological status within plant tubers for survival. The work provided new insights into the better understanding of the molecular basis of plant proteomes and stress tolerance mechanisms, especially during seasonal changes. The finding suggested proteins that might potentially be used as protein markers in differing seasons in other plants and aid in selecting B. superba tubers with the most suitable medicinal properties in the future.

Proteome analysis of Pueraria mirifica tubers collected in different seasons.

Pueraria mirifica-derived tuberous powder has been long-term consumed in Thailand as female hormone-replacement traditional remedies. The protein profiles of tubers collected in different seasons were evaluated. Phenol extraction, 2D-PAGE, and mass spectrometry were employed for tuberous proteome analysis. Out of the 322 proteins detected, over 59% were functionally classified as being involved in metabolism. The rest proteins were involved in defense, protein synthesis, cell structure, transportation, stress, storage, and also unidentified function. The proteins were found to be differentially expressed with respect to harvest season. Importantly, chalcone isomerase, isoflavone synthase, cytochrome p450, UDP-glycosyltransferase, and isoflavone reductase, which are all involved in the biosynthesis pathway of bioactive isoflavonoids, were most abundantly expressed in the summer-collected tubers. This is the first report on the proteomic patterns in P. mirifica tubers in relevant with seasonal variation. The study enlights the understanding of variance isoflavonoid production in P. mirifica tubers.

Clinical, radiographic, and histologic analysis of the effects of acemannan used in direct pulp capping of human primary teeth: short-term outcomes.

Acemannan has been previously reported as a direct pulp-capping agent in animal study. This natural material demonstrated its biocompatibility and enhanced reparative dentin formation. The objective of this study was to investigate the action of acemannan as a direct pulp-capping material in human primary teeth with deep caries. Forty-two deeply carious mandibular primary molars from 37 children, aged 7-11 years old diagnosed with reversible pulpitis were studied. After completely removing the infected dentine, teeth with a pinpoint pulpal exposure were randomly divided into two treatment groups: acemannan or calcium hydroxide. A glass-ionomer cement base was applied to all teeth prior to restoration with stainless steel crowns. Clinical and radiographic evaluation was performed 6 months post-treatment. The teeth due to exfoliate were extracted and histopathologically evaluated for inflammation, dentine bridge formation, and soft tissue organization. At 6 months, the overall clinical and radiographic success rates of direct pulp capping with acemannan and calcium hydroxide at 6 months were 72.73 and 70.0 %, respectively. The histopathological results indicated that the acemannan-treated group had significantly better histopathological responses compared with the calcium hydroxide-treated group (p < 0.05). These data suggest acemannan offers a valuable alternative biomaterial for vital pulp therapy in primary teeth.

Antioxidant and antiproliferative activities of protein hydrolysate from the rhizomes of Zingiberaceae plants.

Plant proteins have been investigated for their antioxidant activities, but there are still no reports detailing the antioxidant activity levels of plants in the Zingiberaceae family, which are popular food agents and used in folklore medicine. In this study, the crude rhizome protein extract and associated pepsin/pancreatin protein hydrolysate of 15 plants in the Zingiberaceae family were screened using the DPPH method for antioxidant activity. The protein hydrolysate of C. zedoaria possessed the highest antioxidant activity (IC50of 25.7±6.3µg/mL), which was close to that of the reference ascorbic acid (IC50of 22.3±1.8µg/mL). After enrichment by Q Sepharose ion exchange chromatography using a five step elution gradient of increasing NaCl concentration (0, 0.25, 0.5, 0.75 and 1M), the fraction eluting in the 0.5M NaCl (F50) showed the highest antioxidant activity (IC50 of 41.78±2.9µg/mL), and was found to have weak in vitro cytotoxicity against the HEP-G2 and SW620 cell lines (IC50 of 200.8±11.8 and 241.0±9.3µg/mL, respectively), but not the BT474, CHAGO and KATO-3 cell lines. F50 had an estimated molecular weight by MALDI-TOF mass spectrometry of 12,400-12,800 Da.

Protein profiling of mefloquine resistant Plasmodium falciparum using mass spectrometry-based proteomics.

Malaria is a mosquito borne infectious disease caused by protozoa of genus Plasmodium. There are five species of Plasmodium that are found to infect humans. Plasmodium falciparum can cause severe malaria leading to higher morbidity and mortality of malaria than the other four species. Antimalarial resistance is the major obstacle to control malaria. Mefloquine was used in combination with Artesunate for uncomplicated P. falciparum in South East Asia and it has developed and established mefloquine resistance in this region. Here, gel-enhanced liquid chromatography/tandem mass spectrometry (GeLC-MS/MS)-based proteomics and label-free quantification were used to explore the protein profiles of mefloquine-sensitive and -induced resistant P. falciparum. A Thai P. falciparum isolate (S066) was used as a model in this research. Our data revealed for the first time that 69 proteins exhibited at least 2-fold differences in their expression levels between the two parasite lines. Of these, 36 were up-regulated and 33 were down-regulated in the mefloquine-resistant line compared with the mefloquine-sensitive line. These findings are consistent with those of past studies, where the multidrug resistance protein Pgh1 showed an up-regulation pattern consistent with that expected from its average 3-copy pfmdr1 gene number. Pgh1 and eight other up-regulated proteins (i.e., histo-aspartyl protease protein, exportin 1, eukaryotic translation initiation factor 3 subunit 8, peptidyl-prolyl cis-trans isomerase, serine rich protein homologue, exported protein 1, ATP synthase beta chain and phospholipid scramblase 1) were further validated for their expression levels using reverse transcriptase quantitative real-time PCR. The data support the up-regulation status in the mefloquine-resistant parasite line of all the candidate genes referred to above. Therefore, GeLC-MS/MS-based proteomics combined with label-free quantification is a reliable approach for exploring mefloquine resistance biomarkers in P. falciparum. Identification of these proteins leads to better understanding of mefloquine resistant mechanisms in malaria parasites.

Untargeted metabolomics analysis revealed changes in the composition of glycerolipids and phospholipids in Bacillus subtilis under 1-butanol stress.

1-Butanol has been utilized widely in industry and can be produced or transformed by microbes. However, current knowledge about the mechanisms of 1-butanol tolerance in bacteria remains quite limited. Here, we applied untargeted metabolomics to study Bacillus subtilis cells under 1-butanol stress and identified 55 and 37 ions with significantly increased and decreased levels, respectively. Using accurate mass determination, tandem mass spectra, and synthetic standards, 86 % of these ions were characterized. The levels of phosphatidylethanolamine, diglucosyldiacylglycerol, and phosphatidylserine were found to be upregulated upon 1-butanol treatment, whereas those of diacylglycerol and lysyl phosphatidylglycerol were downregulated. Most lipids contained 15:0/15:0, 16:0/15:0, and 17:0/15:0 acyl chains, and all were mapped to membrane lipid biosynthetic pathways. Subsequent two-stage quantitative real-time reverse transcriptase PCR analyses of genes in the two principal membrane lipid biosynthesis pathways revealed elevated levels of ywiE transcripts in the presence of 1-butanol and reduced expression levels of cdsA, pgsA, mprF, clsA, and yfnI transcripts. Thus, the gene transcript levels showed agreement with the metabolomics data. Lastly, the cell morphology was investigated by scanning electron microscopy, which indicated that cells became almost twofold longer after 1.4 % (v/v) 1-butanol stress for 12 h. Overall, the studies uncovered changes in the composition of glycerolipids and phospholipids in B. subtilis under 1-butanol stress, emphasizing the power of untargeted metabolomics in the discovery of new biological insights.

Pueraria mirifica leaves, an alternative potential isoflavonoid source.

We investigated the major leaf isoflavonoid contents of Pueraria mirifica from three different cultivars (PM-III, PM-IV, and PM-V) using reverse RP-HPLC analysis. The proportions and net levels of puerarin, daidzin, genistin, and daidzein in P. mirifica leaves were found to depend on the plant cultivar and to correlate with cultivation temperature and rainfall amount. The crude leaf-extracts were tested using the Yeast Estrogen Screen (YES) assay with both human estrogen receptors (hERα and hERß). Their estrogenic activity was higher when determined by the YES system containing hERß than that with hERα and was also higher when the Δsnq2 than the wildtype yeast was employed. The results open the possibility of selecting and cultivating certain P. mirifica cultivars at a farm scale to produce a sufficient supply of leaf material to act as a starting source for the commercial scale extraction of these major isoflavonoids.

Effect of acemannan, an extracted polysaccharide from Aloe vera, on BMSCs proliferation, differentiation, extracellular matrix synthesis, mineralization, and bone formation in a tooth extraction model.

Aloe vera is a traditional wound healing medicine. We hypothesized acemannan, a polysaccharide extracted from Aloe vera gel, could affect bone formation. Primary rat bone marrow stromal cells (BMSCs) were treated with various concentrations of acemannan. New DNA synthesis, VEGF, BMP-2, alkaline phosphatase activity, bone sialoprotein, osteopontin expression, and mineralization were determined by [(3)H] thymidine incorporation assay, ELISA, biochemical assay, western blotting, and Alizarin Red staining, respectively. In an animal study, mandibular right incisors of male Sprague-Dawley rats were extracted and an acemannan treated sponge was placed in the socket. After 1, 2, and 4 weeks, the mandibles were dissected. Bone formation was evaluated by dual-energy X-ray absorptiometry and histopathological examination. The in vitro results revealed acemannan significantly increased BMSC proliferation, VEGF, BMP-2, alkaline phosphatase activity, bone sialoprotein and osteopontin expression, and mineralization. In-vivo results showed acemannan-treated groups had higher bone mineral density and faster bone healing compared with untreated controls. A substantial ingrowth of bone trabeculae was observed in acemannan-treated groups. These data suggest acemannan could function as a bioactive molecule inducing bone formation by stimulating BMSCs proliferation, differentiation into osteoblasts, and extracellular matrix synthesis. Acemannan could be a candidate natural biomaterial for bone regeneration.

Characterization of a bioscaffold containing polysaccharide acemannan and native collagen for pulp tissue regeneration.

Dental pulp regeneration exploits tissue engineering concepts using stem cells/scaffolds/growth-factors. Extracted collagen is commonly used as a biomaterial-scaffold due to its biocompatibility/biodegradability and mimics the natural extracellular matrix. Adding biomolecules into a collagen-scaffold enhanced pulp regeneration. Acemannan, ß-(1-4)-acetylated-polymannose, is a polysaccharide extracted from aloe vera. Acemannan is a regenerative biomaterial. Therefore, acemannan could be a biomolecule in a collagen-scaffold. Here, acemannan and native collagen were obtained and characterized. The AceCol-scaffold's physical properties were investigated using FTIR, SEM, contact angle, swelling, pore size, porosity, compressive modulus, and degradation assays. The AceCol-scaffold's biocompatibility, growth factor secretion, osteogenic protein expression, and calcification were evaluated in vitro. The AceCol-scaffolds demonstrated higher hydrophilicity, swelling, porosity, and larger pore size than the collagen scaffolds (p < 0.05). Better cell-cell and cell-scaffold adhesion, and dentin extracellular matrix protein (BSP/OPN/DSPP) expression were observed in the AceCol-scaffold, however, DSPP expression was not detected in the collagen group. Significantly increased cellular proliferation, VEGF and BMP2 expression, and mineralization were detected in the AceCol-scaffold compared with the collagen-scaffold (p < 0.05). Computer simulation revealed that acemannan's 3D structure changes to bind with collagen. In conclusion, the AceCol-scaffold synergistically provides better physical and biological properties than collagen. The AceCol-scaffold is a promising material for tissue regeneration.

Lipid biomarker profiling of adult Brugia malayi using mass spectrometry detection.

In this article, we first reported the lipid profile of adult Brugia malayi using ultra-performance liquid chromatography electrospray ionization mass spectrometry (UPLC-ESI-MS) to provide a promising drug target for lymphatic filariasis (LF). The MS and MS/MS data analysis indicated that there were target lipids, phosphatidylcholines (PC), at m/z 769.5612 and 831.5767, which were found only in adult males, while at m/z 811.6072, they were found only in females, which is interesting for use as a biomarker in LF disease. In addition, the lipid profile showed that three membrane lipid classes, glycerophospholipids, glycerolipids and sphingolipids, were discovered. Glycerophospholipids were the main components in adult parasites, especially phosphatidylcholine (60%) and phosphatidylethanolamine (27%). Phosphatidylglycerol (5%), phosphatidylserine (4%), phosphatidylinositol (4%) and phosphatidic acid (3%). Consequently, the lipid profile of adult B. malayi is significant and not only provides a promising drug target for LF but also assists in a better understanding of the biological process and mechanism by which parasites interfere with LF disease in the future. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03296-y.

A proteomic analysis of Curcuma comosa Roxb. rhizomes.

BACKGROUND: The similarly in plant physiology and the difficulty of plant classification, in some medicinal plant species, especially plants of the Zingiberaceae family, are a major problem for pharmacologists, leading to mistaken use. To overcome this problem, the proteomic base method was used to study protein profiles of the plant model, Curcuma comosa Roxb., which is a member of the Zingiberaceae and has been used in traditional Thai medicine as an anti-inflammatory agent for the treatment of postpartum uterine bleeding. RESULTS: Due to the complexity of protein extraction from this plant, microscale solution-phase isoelectric focusing (MicroSol-IEF) was used to enrich and improve the separation of Curcuma comosa rhizomes phenol-soluble proteins, prior to resolving and analyzing by two-dimensional polyacrylamide gel electrophoresis and identification by tandem mass spectrometry. The protein patterns showed a high abundance of protein spots in the acidic range, including three lectin proteins. The metabolic and defense enzymes, such as superoxide dismutase (SOD) and ascorbate peroxidase, that are associated with antioxidant activity, were mainly found in the basic region. Furthermore, cysteine protease was found in this plant, as had been previously reported in other Zingiberaceae plants. CONCLUSION: This report presents the protein profiles of the ginger plant, Curcuma comosa. Several interesting proteins were identified in this plant that may be used as a protein marker and aid in identifying plants of the Zingiberaceae family.

Zingipain, A cysteine protease from Zingiber ottensii Valeton rhizomes with antiproliferative activities against fungi and human malignant cell lines.

The objective of this study was to investigate the activity of a protein identified as cysteine protease, purified from Zingiber ottensii Valeton rhizomes, in terms of antiproliferation against fungi, bacteria, and human malignant cell lines. By means of buffer extraction followed by (NH(4))(2)SO(4) precipitation and ion-exchange chromatography, the obtained dominant protein (designated F50) was submitted to non-denaturing and reducing sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), where a single band and three bands were revealed from eletrophoretic patterns, respectively. It could be concluded at this point that the F50 was potentially a heterotrimer or heterodimer composed of either two small (∼13.8 and ∼15.2 kD) subunits or these two together with a larger (∼32.5 kD) one. In-gel digestion was carried out for the most intense band from reducing SDS-PAGE, and to the resulting material was applied liquid chromatography (LC)-mass spectroscopy (MS)/MS. The main F50 subunit was found to contain fragments with 100% similarity to zingipain-1, a cysteine protease first discovered in Zingiber officinale. The activity corresponding to the identified data, cysteine protease, was then confirmed in the F50 by azocasein assay and a positive result was obtained. The F50 then was further investigated for antiproliferation against three plant pathogenic fungi species by disk diffusion test, four bacterial species by direct exposure in liquid culture and dish diffusion tests, and five human malignant cell lines by tissue culture assay. It was found that a dose of 23.6 µg F50/0.3 cm(2) of paper disk exhibited the best inhibitory effect against Collectotrichum cassiicola, while lesser effects were found in Exserohilum turicicum and Fusarium oxysporum, respectively. No inhibitory effect against bacterial proliferation was detected in all studied bacterial strains. However, relatively strong antiproliferative effects were found against five human cell lines, with IC50 values ranging from 1.13 µg/mL (hepatoma cancer; HEP-G2) to 5.37 µg/mL (colon cancer; SW620). By periodic acid-Schiff's staining and phenol-sulfuric acid assay, the F50 was determined as a glycoprotein containing 26.30 ± 1.01% (by weight) of carbohydrate. Thus, a new glycoprotein with protease activity was successfully identified in Zingiber ottensii rhizome. The glycoprotein also contained antiproliferative activity against some plant pathogenic fungi and human cancer cell lines.

Acemannan-induced tooth socket healing: A 12-month randomized controlled trial.

BACKGROUND/PURPOSE: Natural compounds have become alternatives for bone regeneration. Acemannan, the main polysaccharide extracted from Aloe vera, has been demonstrated as a promising osteoinductive material in vitro and in vivo. This clinical study investigated the effect of acemannan on tooth socket healing. MATERIALS AND METHODS: Thirty-five otherwise healthy patients, 18-25 years old and diagnosed with horizontal or vertical partial impaction of the lower third molars, were enrolled in this randomized controlled trial. After removing the teeth, the sockets randomly received one of the following treatments: spontaneous blood-clotting (control), 20 mg acemannan sponge, or 50 mg acemannan sponge. Cone-beam computed tomography of the mandible was performed immediately (baseline), and at 3-, 6-, and 12-months postoperatively; the data were analyzed using the OsiriX MD program. Bone healing in the socket was determined measuring the socket volume. One-way ANOVA was used to analyze the differences within each group and between groups. RESULTS: Thirty-five patients with 43 partially impacted lower third molars participated in this study. No patients exhibited alveolar osteitis or secondary infection. Compared with baseline, all groups showed significant reduction in socket volume at all observation time-points (p < 0.05). The 50 mg acemannan group had a significantly greater reduction in socket volume compared with the control at all postoperative time-points (p < 0.05). The 20 mg group had a significantly greater reduction in socket volume compared with the control at 3-months postoperatively (p < 0.05). CONCLUSION: We conclude that acemannan increases bone healing at 3-, 6-, and 12-months after removal of partially impacted mandibular third molars.

Global Analysis of Protein Expression in A549 Cells After Prolonged Nicotine Exposure by Using Label-free Quantification.

BACKGROUND/AIM: Lung cancer is the leading cause of cancer death worldwide. Cigarette smoke is the most important risk factor for cancer development. Growing evidence indicates that prolonged nicotine exposure is a potential factor associated with tumorigenesis. Here, the effect of prolonged nicotine exposure on A549 cells was investigated, using label-free quantitative proteomics. MATERIALS AND METHODS: Selection of an invasive subpopulation from the A549 cell line was performed to reveal the differential expression of proteins in relation to prolonged nicotine exposure, using Boyden chamber assays in combination with a proteomics approach. RESULTS: One hundred proteins from the NicoA549-L5 subline showed significant change in expression compared to those from the A549-L5 subline and their A549 parental cell line. Heat shock protein, protein disulfide isomerase A3, profilin-1 and legumain were expressed at higher levels in A549 cells after prolonged nicotine exposure. CONCLUSION: These aberrant proteins might serve as novel cancer biomarkers for cigarette smokers.

A thermostable lectin from the rhizomes of Kaempferia parviflora.

BACKGROUND: Kaempferia parviflora, or black galingale (Kra-Chai-Dam), belongs to the Zingiberaceae family and is used as both a food ingredient and a medicinal plant. There are diverse reports on the biological activities of compounds extracted from the plant, such as antimalarial, antifungal and an effective sexual-enhancing role, but not on the lectins. RESULTS: A lectin was isolated from the rhizomes of Kaempferia parviflora using affinity chromatography on Concanavalin A followed by gel filtration chromatography on Sephacryl S-100. The molecular weight of the purified lectin was about 41.7 kDa. This lectin showed haemagglutinating activity against erythrocytes from several sources, with the highest level being against those from rabbits. Moreover, the lectin was thermostable, with significant haemagglutinating activity detectable up to 75 degrees C. The results of trypsin digestion and liquid chromatography/tandem mass spectrometry analysis suggested that this protein could be a member of the lectin/endochitnase1 family. CONCLUSION: A lectin that showed thermotolerant haemagglutinating activity against erythrocytes from several sources was successfully purified from K. paviflora rhizomes. Peptide sequence analysis indicated that this lectin is similar to lectin/endochitinase 1 (Urtica dioica) or Hevein-like protein (Hevea brasiliensis).

Cell proliferative effect of polyxyloses extracted from the rhizomes of wild turmeric, Curcuma aromatica.

Hot water-soluble crude polysaccharides were extracted from the rhizomes of wild turmeric, Curcuma aromatica Salisb. (Zingiberaceae), using dry grinding, boiling water extraction, and then ethanol precipitation. The crude polysaccharide extract was then fractionated by DEAE-cellulose ion exchange column chromatography, and subsequently further purified by Superdex G-200 gel filtration column chromatography, giving two relatively abundant polysaccharide fractions, called P11 and P21, and a much less common fraction P22 obtained in insufficient amounts for further analysis. The two main polysaccharide fractions were evaluated for monosaccharide composition by acid hydrolysis and high performance liquid chromatography (HPLC), whilst the molecular weight and functional groups were determined by gel permeable chromatography (GPC) and FT-IR, respectively. Fractions P11 and P21 were found to be polyxyloses with molecular weight-averages of 469,171 and 157,665 Da, respectively. P11 (100 microg/mL) could significantly induce human gingival fibroblast cells proliferation by 30%, while P21 (100 microg/mL) could significantly inhibit gingival fibroblast cells proliferation by 92%. The in vitro human primary gingival fibroblast cell proliferation in cell culture at a concentration of 100 microg/mL.

Hemagglutinating activity of proteins from Parkia speciosa seeds.

Proteins from Parkia speciosa Hassk. (Fabaceae) seeds were extracted and stepwise precipitated using ammonium sulfate. Proteins precipitated with 25% ammonium sulfate were separated by affinity chromatography on Affi-Gel Blue gel followed by protein liquid chromatography on Superdex 200. The protein Gj, which was identified as a protein similar to putative aristolochene synthase, 3'-partial from Oryza sativa L. (Poaceae), had hemagglutinating activity of 0.39 mug/muL. Moreover, fraction C2 from the proteins precipitated with 60% ammonium sulfate, separated by lectin-specific adsorption chromatography using Con A Sepharose, had hemagglutinating activity of 1.17 mug/muL. Using gel electrophoresis, two proteins C2a and C2b were separated, having molecular weights of 45 kDa and 23 kDa, respectively. From protein identification, C2a was found to be similar to the hypothetical protein B1342F01.11 from Oryza sativa, and C2b was similar to the hypothetical protein At1g51560 from Arabidopsis thaliana (L.) Heynh. (Brassicaceae).

Acemannan induces rapid early osseous defect healing after apical surgery: A 12-month follow-up of a randomized controlled trial.

BACKGROUND/PURPOSE: Acemannan is an osteoinductive material. This study's objective was to compare the outcomes of bone defect healing using 3-dimensional images after apical surgery with or without adding acemannan sponges. MATERIALS AND METHODS: Twenty-two anterior teeth from 9 males and 13 females requiring apical surgery were included in this randomized controlled trial. Post-surgery, the bone defects were randomly divided into three groups: blood clot control, 5-, or 10-mg acemannan sponge groups. CBCT scans were taken immediately (baseline), 3-, 6-, and 12-month post-surgery. Sagittal serial sections (1 mm thick slices parallel to the long axis of the tooth) of the defect image were created. The defect boundary was located and the total bone defect volume (BDV) was calculated from the sum of the volume of the serial defect sections. The bone healing was assessed by the percentage of total bone defect volume reduction (%ΔBDV). The paired t-test and one-way ANOVA were used to analyze the differences within each group and between groups, respectively. RESULTS: The baseline mean BDV of the control, 5-, and 10-mg acemannan groups were not significantly different (p > 0.05). After treatment, the mean BDV for each group was reduced in a time-dependent manner. Compared with the control group, the 5- and 10-mg acemannan groups had a significantly greater %ΔBDV (approximately 2- and 1.89-fold) at 3-months post-surgery, respectively (p < 0.05). However, at the 6- and 12- month follow-up, the %ΔBDV was not significantly different between the groups. CONCLUSION: These data suggest acemannan enhanced early bone healing after apical surgery.