Sphingolipid-Enriched Porcine Placental Extract Promotes the Expression of Structural Genes and Desquamation Enzyme Genes in Cultured Human Keratinocytes.

Porcine placental extract (PPE) is commonly used in various health foods and cosmetics. PPE use in cosmetics predominantly consist of the water-soluble fraction derived from the entire placenta. In this report, we examined the effect of the hydrophobic constituents of the PPE, specifically the sphingolipid-enriched fraction designated as the sphingolipid-enriched porcine placental extract (SLPPE), on the expression of genes associated with skin function in cultured normal human epidermal keratinocytes. Using quantitative RT-PCR (qRT-PCR) analysis, we found that SLPPE concentrations ranging from 25 to 100 µg/mL upregulated the gene expression of key components associated with the cornified envelope structure (filaggrin (FLG), involucrin (IVL) and loricrin (LOR)), cornification enzymes (transglutaminase 1 (TGM1) and TGM5) and the desquamation enzymes (kallikrein 5 (KLK5) and KLK7). Additionally, KLK5p and FLG protein (FLGp) were detected in the culture supernatants of keratinocytes treated with SLPPE at these concentrations. These findings suggest that SLPPE is possible to promote the cornification and desquamation in epidermal keratinocytes, and it may offer potential benefits in cosmetics.

Anti-inflammatory Effect of the Water-Soluble Portion of Porcine Placental Extract in Lipopolysaccharide-Stimulated RAW264.7 Murine Macrophage Cells.

Porcine placental extract (PPE) is used as a nonprescription drug for analeptics and in health foods and cosmetics in Japan, Korea and China. It was reported that PPE has anti-oxidative and anti-inflammatory activities; however, the mechanisms and the responsible molecules involved in these activities are still unclear. Here, we investigated how enzymatically prepared PPE affects proinflammatory factors such as interleukin (IL)-1ß, IL-6, and tumor necrosis factor (TNF)-α in a cultured macrophage cell line, RAW264.7, when co-stimulated with lipopolysaccharide (LPS). Enhanced production of IL-1ß, IL-6 and TNF-α by LPS was significantly reduced by the addition of PPE and these effects were dose dependent. Nitric oxide (NO) production induced in cultured macrophages by LPS was also inhibited by PPE. Real-time PCR after the reverse transcription of total RNAs isolated from cells treated with PPE revealed that the mRNA expressions of IL-1ß, IL-6, TNFα, and NO synthase (NOS)-2 were reduced. The necessary concentration of PPE prepared by enzymatic digestion to mediate anti-inflammatory effects compared with the reported value of that extracted by phosphate buffered saline without digestion was proportional to the amount of extracted materials from the same amount of placenta (about 10-fold). This suggests that the molecules responsible for the anti-inflammatory activity exists in the placenta and can be extracted by phosphate buffered saline, and thus might survive enzymatic digestion.

Suppression of melanin synthesis by the phenolic constituents of sappanwood (Caesalpinia sappan).

Sappanwood (Caesalpinia sappan Linn.) is used as an herbal medicine. It is sometimes used to treat skin damage or as a facial cleanser. In the present study, the methanol (MeOH) extract of sappanwood was found to inhibit melanin synthesis in cultured human melanoma HMV-II cells stimulated with forskolin, and six active compounds (1-5 and 7) were isolated from the extract along with a non-active compound (6). Compounds 2-7 were identified as sappanchalcone (2), 3'-deoxy-4-O-methylsappanol (3), brazilein, (4), brazilin (5), sappanol (6), and 4-O-methylsappanol (7). Compound 1 was a new compound, and its structure was determined to be (6aS,11bR)-7,11b-dihydro-6H-indeno[2,1-c]chromene-3,6a,10,11-tetrol by spectroscopic analyses. Among the six active compounds, brazilin (5) (EC50: 3.0 ± 0.5 µM) and 4-O-methylsappanol (7) (EC50: 4.6 ± 0.7 µM) strongly suppressed melanin synthesis in HMV-II cells. Bioactive compounds showed moderate cytotoxicities against HMV-II cells with IC50 values of 83.1 ± 4.0 µM (for 2), 72.0 µM ± 2.4 (for 3), 33.8 ± 1.1 µM (for 4), 18.4 ± 0.8 µM (for 5), and 20.2 ± 0.8 (for 7), respectively. Brazilin (5) selectively suppressed the expression of mRNAs for tyrosinase-related protein (TYRP) 2 and tyrosinase but did not influence the expression of TYRP1. These results suggest that brazilin (5) is a new class of melanin inhibitor and that sappanwood could be used as a cosmetic material.

Bioactive phenylpropanoid glycosides from Tabebuia avellanedae.

Three novel phenylpropanoid glycosides 2, 5, 6 were isolated from water extract of Tabebuia avellanedae, together with three known phenylpropanoid glycosides 1, 3, 4. All compounds were identified on the basis of spectroscopic analysis and chemical methods and, for known compounds, by comparison with published data. All isolated compounds showed strong antioxidant activity in the DPPH assay, and compound 5 give the highest antioxidant activity among all compounds, with an IC50 of 0.12 µM. All compounds exhibited moderate inhibitory effect on cytochrome CYP3A4 enzyme.

Anti-inflammatory and anti-melanogenic proanthocyanidin oligomers from peanut skin.

Peanut skin (Arachis hypogaea L., Fabaceae) is an abundant source for polyphenols, such as proanthocyanidin oligomers. To determine whether proanthocyanidin has beneficial effects on skin, we tested for inhibitory activity of proanthocyanidins isolated from peanut skin on inflammatory cytokine production and melanin synthesis in cultured cell lines. Administration of peanut skin extract (PSE, 200 µg/mL) decreased melanogenesis in cultured human melanoma HMV-II co-stimulated with phorbol-12-myristate-13-acetate. It also decreased production of inflammatory cytokines (PSE at 100 µg/mL), tumor necrosis factor-α and interleukin-6, in cultured human monocytic THP-1 cells in response to lipopolysaccharide. We isolated ten known proanthocyanidins and one new proanthocyanidin trimer from the PSE. The structure of the new compound (5) was determined by 1D- and 2D-NMR and mass spectrometry analyses, and was determined as epicatechin-(2ß→O→7,4ß→6)-epicatechin-(4ß→6)-epicatechin. The other known proanthocyanidins were identified as proanthocyanidin monomers (1), dimers (6-9), trimers (3-5) and tetramers (2, 10, 11). They showed suppressive activities against melanogenesis and cytokine production at concentrations ranging from 0.1-10 µg/mL. Among the tested compounds, suppressive activities of proanthocyanidin dimers or trimers in two assay systems were stronger than those obtained with monomer or tetramers. These data indicate that proanthocyanidin oligomers from peanut skin have the potential to reduce dermatological conditions such as inflammation and melanogenesis.

Phytochemical and in silico studies for potential constituents from Centaurium spicatum as candidates against the SARS-CoV-2 main protease and RNA-dependent RNA polymerase.

In the present study, a new secoiridoid glycoside lisianthoside II 1, along with seven known compounds 2-8, were isolated from Centaurium spicatum L. In-silico molecular docking and molecular dynamic simulation against SARS-CoV-2 Main protease (Mpro) and RNA-dependent RNA polymerase (RdRp) were conducted. The affinity docking scores revealed that 8 is the best bound ligand to Mpro active site with binding energy of -14.9877 kcal/mol (RSMD = 1.16 Å), while 6 was the highest against RdRp (-16.9572 kcal/mol, RMSD = 1.01 Å). Moreover, the molecular dynamic simulation revealed that 8 with a (ΔG) of -7.9 kcal/mol (RMSD value of 2.6 Å) and 6 (RMSD value of 1.6 Å) and binding free energy (ΔG) of -7.1 kcal/mol achieved the highest stability over 50 ns of MDS inside the Mpro and RdRp enzyme's active site, respectively. Hence, the isolated compounds could be a good lead for development of new leads targeting COVID-19.

Stilbenoids from the melinjo (Gnetum gnemon L.) fruit modulate cytokine production in murine Peyer's patch cells ex vivo.

Melinjo fruit ( Gnetum gnemon L.) has been used as a food in Southeast Asia. To investigate if this fruit has regulatory actions on ileal immune responses, we measured T-helper (Th) cytokine production, i.e., interleukin-2 (IL-2), IL-4, IL-5, and interferon-gamma (IFN- γ), in cultured Peyer's patch (PP) cells from mice orally treated with a methanol extract of melinjo fruit. Oral administration of the 50 % ethanol extract at 100 mg/kg/day significantly enhanced the production of the Th1 cytokines IL-2 and IFN- γ irrespective of concanavalin-A stimulation, whereas the production of the Th2 cytokines IL-4 and IL-5 was not affected. We also isolated seven active constituents accompanied with two new stilbenoids from the ethylacetate fraction of the extracts. The structure of the new stilbene glucosides gnemonoside L (5) and gnemonoside M (7) was determined by 1D and 2D NMR and MS analyses. Five known stilbenoids were identified as resveratrol (1), isorhapontigenin (2), gnemonoside D (4), gnetins C (3) and E (6). Among these tested compounds, only new stilbenoid 7 strongly enhanced Th1 cytokine production in cultured PP cells at 10 mg/kg/day. These results indicated that this melinjo extract and its active constituent 7 potentiated T-cell-dependent immune responses in the ileal mucosa.

Anti-inflammatory and anti-melanogenic steroidal saponin glycosides from Fenugreek (Trigonella foenum-graecum L.) seeds.

Fenugreek seed ( Trigonella foenum-graecum L.) is used as an herbal medicine for treating metabolic and nutritive dysfunctions. To determine if this plant has other beneficial effects, we tested the inhibitory activities of a methanol (MeOH) extract of fenugreek seed on the production of inflammatory cytokines and melanin synthesis in cultured cell lines in vitro. The MeOH extract inhibited the production of phorbol-12-myristate-13-acetate-induced inflammatory cytokines such as tumor necrosis factor (TNF)-α in cultured THP-1 cells, and also restrained the intracellular synthesis of melanin in murine melanoma B16F1 cells. We isolated three active constituents from fenugreek seed extracts. These were identified as the steroidal saponins 26- O-ß-D-glucopyranosyl-(25 R)-furost-5(6)-en-3 ß,22 ß,26-triol-3- O-α-L-rhamno-pyranosyl-(1'' → 2')-O-[ß-D-glucopyranosyl-(1''' → 6')- O]-ß-D-glucopyranoside 1, minutoside B 2, and pseudoprotodioscin 3. Compounds 1 and 2 strongly suppressed the production of inflammatory cytokines, whereas 3 showed a weaker suppressing effect. Melanogenesis in B16F1 cells was significantly suppressed by 1 and 3, and weakly suppressed by 2. All three compounds showed moderate cytotoxicities. These results indicate that fenugreek extract and its active constituents could protect against skin damage.

Possible involvement of the µ opioid receptor in the antinociception induced by sinomenine on formalin-induced nociceptive behavior in mice.

Sinomenine, an alkaloid originally isolated from the roots and the rhizome of Sinomenium acutum is used as a traditional Chinese herbal medicines for rheumatoid arthritis and neuralgia. The aims of this study were to investigate the effects of oral administration of shinomenine on formalin-induced nociceptive behavior in mice and the opioid receptor subtypes involved in the antinociceptive effects of sinomenine. Our findings showed that a single dose of oral-administrated sinomenine inhibited the formalin induced licking and biting responses in a dose-dependent manner. Intraperitoneal pretreatment with naloxone hydrochloride, an opioid receptor antagonist, and ß-funaltrexamine hydrochloride (ß-FNA), a selective µ-opioid receptor antagonist, significantly attenuated sinomenine induced antinociception, but not by naltrindole, a nonselective δ-opioid receptor antagonist and nor-binaltorphimine, a selective κ-opioid receptor antagonist. Furthermore, in western blot analysis, oral administration of sinomenine resulted in a significant blockage of spinal extracellular signal-regulated protein kinase (ERK1/2) activation induced by formalin. Naloxone hydrochloride and ß-FNA significantly reversed the blockage of spinal ERK1/2 activation induced by sinomenine. These results suggest that sinomenine-induced anti nociceptive effect and blockage of spinal ERK1/2 activation may be triggered by activation of µ-opioid receptors.

Antioxidant C-glycosyl flavones from the leaves of Sasa kurilensis var. gigantea.

C-glycosyl flavones, kurilensin A (1) and B (2), together with two known compounds, tricin-4'-O-beta-d-glucopyranoside (3) and tricin-5-O-beta-d-glucopyranoside (4), were isolated from hot-water extracts of the leaves of Sasa kurilensis. The structure of the compounds was determined by spectroscopic analyses including 1D, 2D NMR and MS. The absolute configuration of sugar moieties in 1 and 2 was determined by chiral HPLC analyses of the benzoyl derivatives of acid hydrolysis. Compounds 1 and 2 exhibited higher radical scavenging activity than ascorbic acid in the 1,1-diphenyl-2-pycrylhydrazyl (DPPH) assay system.

Bioactive monoterpene glycosides conjugated with gallic acid from the leaves of Eucalyptus globulus.

Two monoterpene glycosides, conjugated with gallic acid [globulusin A (1) and B (2)], together with four known compounds, cypellocarpin A (3), eucaglobulin (4), cuniloside (5) and (1S, 2S, 4R)-trans-2-hydroxy-1,8-cineole beta-d-glucopyranoside (6), were isolated from hot-water extracts of the leaves of Eucalyptus globulus. The structures of compounds 1 and 2 were determined by 1D, 2D NMR and MS spectroscopic analyses. The absolute stereochemistry of 1 was determined by correlating the spectroscopic data with those of synthetic compound 6 with a known configuration. Globulusin A (1) and B (2), cypellocarpin A (3) and eucaglobulin (4), scavenged DPPH free radicals and globulusin A (1) showed a higher antioxidant activity than the other tested compounds, with an IC50 of 3.8microM. Globulusin A (1) and eucaglobulin (4) concentration-dependently suppressed inflammatory cytokine production, tumor-necrosis factor-alpha and interleukin-1beta in cultured human myeloma THP-1 cells co-stimulated with phorbol myristate acetate. These compounds also inhibited melanogenesis in cultured murine melanoma B16F1 cells, without any significant cytotoxicity. These results suggested that globulusin A (1) and eucaglobulin (4), which were isolated as antioxidants from E. globulus, also had anti-inflammatory as well as anti-melanogenesis activity.

Capsicum ethanol extracts and capsaicin enhance interleukin-2 and interferon-gamma production in cultured murine Peyer's patch cells ex vivo.

We investigated the effects of red pepper (Capsicum annuum Lin.) extracts (capsicum extract) and its main pungent capsaicin on T helper 1 (Th1) and 2 (Th2) cytokine production in cultured murine Peyer's patch (PP) cells in vitro and ex vivo. Direct administration of capsicum extract (1 and 10 mug/ml) and capsaicin (3 and 30 muM) resulted in suppression of interleukin (IL)-2, interferon (IFN)-gamma, IL-4 and IL-5 production. In an ex vivo experiment using PP cells removed from the mice after oral administration of capsicum extract (10 mg/kg/day for 4 consecutive days), IL-2, IFN-gamma and IL-5 increased in response to concanavalin A (Con A). Oral administration of 3 mg/kg/day capsaicin, one active constituent of the extract, also enhanced IL-2, INF-gamma and IL-4 production in response to Con A stimulation but did not influence the production of IL-5. Orally administered capsazepine (3 mg/kg/day), a selective transient receptor potential vanilloid 1 (TRPV1) antagonist, slightly enhanced IL-2 production also irrespective of Con A stimulation. The capsaicin-induced enhancement of both IL-2 and IFN-gamma production was not reduced by oral administration of capsazepine (3 mg/kg/day), suggesting a TRPV1 receptor-independent mechanism. Flow cytometric analysis revealed that the population of CD3(+) cells in the PP cells was significantly reduced while CD19(+) cells increased after oral administration of capsicum extract (1 and 10 mg/kg/day) and capsaicin (0.3 and 3 mg/kg/day). Capsazepine (3 mg/kg/day) weakly but significantly reversed these effects. Orally administered capsicum extract and capsaicin did not change the T cell subset (CD4(+) and CD8(+)), Th1 (IFN-gamma(+)) and T2 (IL-4(+)) ratio. These findings indicate that capsicum extract and capsaicin modulate T cell-immune responses, and their immunomodulatory effects on murine PP cells are partly due to both TRPV1-dependent and -independent pathway.

The liquid culture filtrates of Paecilomyces tenuipes (Peck) Samson (=Isaria japonica Yasuda) and Paecilomyces cicadae (Miquel) Samson (=Isaria sinclairii (Berk.) Llond) regulate Th1 and Th2 cytokine response in murine Peyer's patch cells in vitro and ex vivo.

The effects of liquid culture filtrates of medicinal entomogenous fungi, Paecilomyces tenuipes (Peck) Samson (=Isaria japonica Yasuda or Isaria tenuipes) (PTCF) and Paecilomyces cicadae (Miquel) Samson (=Isaria sinclairii (Berk.) Llond) (PCCF), on cytokine productions in cultured Peyer's patches (PP) from C57BL/6J mice were investigated in vitro and ex vivo. In an in vitro experiment, PTCF (100 and 10 microg/ml) enhanced the production of T helper 1 (Th1) cytokines, interleukin (IL)-2 and interferon (IFN)-gamma, in cultured PP cells stimulated with 5 microg/ml concanavalin A (Con A) but did not influence on the production of T helper 2 (Th2) cytokines, IL-4 and IL-5. PTCF also enhanced the production of granulocyte macrophage colony-stimulating factor (GM-CSF) and IL-10 in the cultured PP cells. While, PCCF enhanced the production of IFN-gamma but did not alter the level of IL-2 in the PP cells. In an ex vivo experiment using PP cells removed from the mice after oral treatment of PTCF (10 and 100 mg/kg daily for 7 consecutive days), the production of IL-2 and IFN-gamma were increased in response to Con A. On the other hand, orally treated PCCF (10 mg/kg/day) suppressed IL-2 production but did not change the levels of IFN-gamma and IL-10 in the isolated PP cells. The flow cytometric analysis revealed that the population of CD3(+) cells in the PP cells slightly but significantly increased after oral administration of PCCF. Orally administered PTCF did not change the population of T (CD3(+)), B (CD19(+)), T cell subset (CD4(+)and CD8(+)) and Th1 (IFN-gamma(+)) and Th2 (IL-4(+)). From PTCF, the fraction rich in proteoglycans was separated as active fraction that stimulates Th1 immune response. These results indicate that the mode of action of PTCF and PCCF on mucosal immune response is different and this is contributed to their metabolites. Taken together, there is a possibility of PTCF and PCCF being therapeutic or preventive agents for immune diseases such as cancer, allergy and parasitic disease through activation of mucosal immune response.

Water-soluble extracts from Angelica acutiloba Kitagawa enhance hematopoiesis by activating immature erythroid cells in mice with 5-fluorouracil-induced anemia.

The extract from the root of Angelica acutiloba Kitagawa (AR), which is used as herbal medicine in Japan, has been reported to be clinically effective for postmenstrual blood loss and erythropoietin (EPO)-resistant anemia in chronic renal failure, although the pharmacological mechanisms underlying its clinical efficacy are unknown. We prepared an animal model of anemia by bolus injection of 5-fluorouracil (5FU) at 150 mg/kg to mice (8- to 12-week-old female C57BL/6J), and then administered orally the water-soluble fraction of AR to the anemic mice for 10 days. After confirming the anti-anemic effect of the water-soluble fraction of AR (AR-3) containing polysaccharides, we examined the effects of AR-3 on immature erythroid cell activity, EPO production, and plasma cytokine levels. AR-3 administration at 50 mg/kg activated erythroid progenitor cells in bone marrow on day 10, increased the percentage of peripheral reticulocytes in red blood cells on day 15, and led to the recovery of red blood cell count to a value that was almost equal to the basal level on day 20. Although EPO production, which was determined by examining EPO mRNA expression in kidney and liver, remained unaltered by AR-3 administration, this treatment significantly lowered plasma interferon-gamma level, which may suppress the activity of erythroid progenitor cells. These results suggest that the polysaccharides in AR promote hematopoiesis by activating immature erythroid cells, in part, by suppressing cytokine secretion. Since the hematopoietic effect was achieved by high-dose AR-3, identification of specific polysaccharides is still required for the development of a novel medicine for anemia caused by a malignancy or chemotherapy.

Protective effect of (+)-catechin against 5-fluorouracil-induced myelosuppression in mice.

The object of this study was to investigate the efficacy of (+)-catechin, which was isolated from Actinidia arguta Planch (Actinidiaceae), as a bone marrow cell proliferation-promoting compound against the hematotoxicity of 5-fluorouracil (5-FU) in mice. Intraperitoneally injected (+)-catechin (1 and 10 mg/kg per day) accelerated the recovery of the number of white blood cells (WBC) and platelets (PLT) but did not affect the number of circulating red blood cells (RBC). (+)-Catechin also augmented the number of myelocytes and splenocytes. Dual color flow cytometric analysis revealed that (+)-catechin reversed the reduction of the population of leukocytes (CD11b+ monocytes, Gr-1+ granulocytes and CD3+ T and CD45RA+ B lymphocytes) in whole blood, spleen and bone marrow caused by 5-FU. (+)-Catechin (1 and 10 mg/kg per day) showed remarkable recovery of Gr-1+ cells in all three types of tissues and of CD11b+ cells in the bone marrow cells. These findings suggest that (+)-catechin selectively enhances the recovery of the population of granulocytes reduced by 5-FU in mice.

Anti-inflammatory constituents from Tabebuia avellanedae.

Five novel compounds were isolated from the water extract of Tabebuia avellanedae, and their structures were established by analysis of NMR spectroscopy and mass spectrometry. Compounds 1-5 at 25µM showed strong inhibitory activity on the inflammatory cytokine, tumor-necrosis factor-α and interleukin-1ß production in cultured human myeloma THP-1 cells co-stimulated with lipopolysaccharide without any significant cytotoxicity, and their anti-allergic and antioxidant activities were evaluated.

Phenolic lipid ingredients from cashew nuts.

Five new phenolic lipids, 2-(8"Z-eicosenoyl)-6-(8'Z-pentadecenyl) salicylic acid (3), 2-(9"Z-hexadecenoyl)-6-(8'Z, 11'Z-pentadecadienyl) methyl salicylate (5), 2-(10"Z, 13"Z-nonadecadienoyl)-6-(8'Z, 11'Z-pentadecadienyl) salicylic acid (6), 2-(16"Z-pentacosenoyl)-6-(8'Z-pentadecenyl) salicylic acid (7) and 2-(9"Z-octadecenoyl)-6-(8'Z, 11'Z-pentadecadienyl) methyl salicylate (8), and three known compounds, cardols (1), anacardic acid (2) and cardanols (4), were isolated from the nuts of the cashew, Anacardium occidentale L. The structures were established on the basis of detailed MS and NMR spectroscopic analyses. Compound 1 highly enhanced both Th-1 (IL-2, IFN-γ) and Th-2 (IL-4, IL-5) cytokine production, and compounds 7 and 8 highly increased cytokine IL-2 and IFN-γ production in response to concanavalin A in cultured murine Peyer's patch cells ex vivo. The isolated compounds showed moderate inhibitory activities on cytochrome CYP3A4 enzyme.

Two new acetylated flavonoid glycosides from Centaurium spicatum L.

Two new acetylated flavonol glycosides, quercetin 3-O-[(2,4-diacetyl-α-L-rhamnopyranosyl)-(1→6)]-2,4-diacetyl-ß-D-galactopyranoside (1) and quercetin 3-O-[(2,4-diacetyl-α-L-rhamnopyranosyl)-(1→6)]-3,4-diacetyl-ß-D-galactopyranoside (2), in addition to two known acetylated quercetin glycosides quercetin 3-O-[(2,3,4-triacetyl-α-L-rhamnopyranosyl)-(1→6)-ß-D-galactopyranoside (3) and quercetin 3-O-[(2,3,4-triacetyl-α-L-rhamnopyranosyl)-(1→6)-3-acetyl-ß-D-galactopyranoside (4), were isolated from the aerial part of Centaurium spicatum (L.) Fritsch (Gentianaceae). Structure elucidation, especially the localization of the acetyl groups, and complete (1)H and (13)C NMR assignments of these biologically active compounds were carried out using one- and two-dimensional NMR measurements, including (1)H- and (13)C-NMR, DEPT-135, H-H COSY, HMQC and HMBC, in addition to HR-FAB/MS experiments.

Frequency of myeloid dendritic cells can predict the efficacy of Wilms' tumor 1 peptide vaccination.

BACKGROUND: The object of this study was to investigate the clinical predictive capability of peripheral myeloid dendritic cells (DCs) in Wilms' tumor 1 (WT1) vaccine therapy for patients with gynaecological cancer. PATIENTS AND METHODS: Six patients with WT1/human leukocyte antigen (HLA)-A*2402-positive gynaecological cancer were included in this study. The patients received intradermal injections of a modified 9-mer WT1 peptide every week for 12 weeks. Peripheral blood samples were obtained at 0, 4, 8 and 12 weeks after the initial vaccination. Circulating DCs were detected by flow cytometry. RESULTS: The frequencies of CD14(+)CD16(+)CD33(+)CD85(+) myeloid DCs were significantly higher in the therapeutically effective group than in therapeutically inert group (p<0.05). CONCLUSION: These results suggested that myeloid DCs, which should be associated with inducing cytotoxic T-cells, provided additional prognostic information in the use of cancer peptide vaccine.

Scanning electron microscopy with an ionic liquid reveals the loss of mitotic protrusions of cells during the epithelial-mesenchymal transition.

Epithelial-mesenchymal transition (EMT) is a key event in cancer metastasis and is characterized by increase in cell motility, increase in expression of mesenchymal cell markers, loss of proteins from cell-to-cell junction complexes, and changes in cell morphology. Here, the morphological effects of a representative EMT inducer, transforming growth factor (TGF)-ß1, were investigated in human lung adenocarcinoma (A549) cells and pancreatic carcinoma (Panc-1) cells. TGF-ß1 caused morphological changes characteristic of EMT, and immunostaining showed loss of E-cadherin from cell-to-cell junction complexes in addition to the upregulation of the mesenchymal marker vimentin. During scanning electron microscopy (SEM) with an ionic liquid, we observed EMT-specific morphological changes, including the formation of various cell protrusions. Interestingly, filopodia in mitotic cells were clearly observed by SEM, and the number of these filopodia in TFG-ß1-treated mitotic cells was reduced significantly. We conclude that this reduction in such mitotic protrusions is a novel effect of TGF-ß1 and may contribute to EMT.