Search

Bacillus subtilis sups. chungkookjang produces a higher molecular mass poly-γ-glutamic acid (γ-PGA). Recently, previous studies have demonstrated immune stimulation and an antitumor effect of the high molecular mass γ-PGA using various mouse models although these effects have not been shown in other species of animals. Therefore, the current study was conducted to determine the effect of γ-PGA in pigs with and without PRRSV infection. PRRS-negative pigs were intramuscularly injected with 1, 3, or 5 ml of 20 mg/mll γ-PGA, and one group of pigs served as a non-treatment (NT) group. All groups treated with γ-PGA had significantly higher weight gains, and pigs treated with 5 ml of γ-PGA exhibited higher tumor necrosis factor (TNF)-α, interferon (IFN)-α and IFN-β expression levels compared with the NT group. According to the preliminary results, an animal challenge study was conducted with a highly virulent PRRSV strain, MN184, along with γ-PGA treatment at different time points. Pigs treated with γ-PGA had lower levels of viral loads in the sera and in lungs and gained significantly more weight (p

Background: We retrospectively evaluated the usefulness of an elevated glucose-to-lymphocyte ratio (GLR) as a sensitive prognostic biomarker of disease-specific survival in 338 patients who underwent surgical resection of pancreatic ductal adenocarcinoma (PDAC). Methods: The optimal GLR cutoff value was determined using the method of Contal and O'Quigley. Patient demographics, clinical information, and imaging data were analyzed to identify preoperative predictors of long-term survival outcomes. Results: Elevated GLR correlated significantly with aggressive tumor biologic behaviors, such as a high carbohydrate antigen (CA) 19-9 level ( p = 0.003) and large tumor size ( p = 0.011). Multivariate analysis identified (1) GLR > 92.72 [hazard ratio (HR) = 2.475, p < 0.001], (2) CA 19-9 level > 145.35 (HR = 1.577, p = 0.068), and (3) symptoms ( p = 0.064) as independent predictors of long-term, cancer-specific survival. These three risk factors were used to group patients into groups 1 (0 factors), 2 (1-2 factors), and 3 (3 factors), which corresponded to significantly different 5-year overall survival rates (50.2%, 34.6%, and 11.7%, respectively; p < 0.001). Conclusions: An elevated preoperative GLR is associated with aggressive tumor characteristics and is an independent predictor of poor postoperative prognosis in patients with PDAC. Further prospective studies are required to verify these findings.

Background: Lung cancer, especially non-small cell lung cancer (NSCLC), poses a significant health challenge globally due to its high mortality. Afatinib, a second-generation epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI), has shown superior efficacy over traditional chemotherapy in NSCLC treatment. However, issues like secondary resistance and adverse effects call for alternative therapies. HAD-B1, comprising 4 herbal medicines, has shown promise in lung cancer treatment in both preclinical and clinical settings. This study assesses the combination of HAD-B1 and Afatinib in advanced NSCLC patients to potentially improve outcomes by addressing the limitations of current EGFR-TKI therapies., Method: A randomized, open-label trial evaluated the efficacy and safety of HAD-B1 with Afatinib in 90 EGFR-mutation-positive NSCLC patients. Participants were divided into treatment and control groups, receiving Afatinib with or without HAD-B1. The study focused on the initial dose maintenance rate and disease control rate (DCR) of Afatinib, alongside secondary outcomes like survival rates and quality of life, under continuous safety monitoring., Results: Among the 90 participants, no significant difference was found in initial dose maintenance (60.98% in the treatment group vs 52.50% in the control, P  = .4414) or DCR (80.49% vs 90.00%, P  = .2283). Secondary outcomes like PFS, TTP, and OS showed no notable differences. However, physical functioning significantly improved in the treatment group ( P  = .0475, PPS group). The control group experienced higher rates of adverse events of special interest and adverse drug reactions ( P  = .01), suggesting HAD-B1 with Afatinib might enhance physical function without increasing adverse effects., Conclusion: Combining HAD-B1 with Afatinib potentially improves quality of life and reduces adverse events in advanced NSCLC patients. Further research is necessary to confirm the long-term benefits of this combination therapy, aiming to advance NSCLC treatment outcomes., Trial Registration: Clinical Research Information Service (CRIS) of the Republic of Korea, https://cris.nih.go.kr/ (ID: KCT0005414)., Competing Interests: Declaration of Conflicting InterestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

The goal of this paper was to look into the anti-tumor mechanism of Non-Steroidal Anti-Inflammatory Drug (NSAID)-conjugated SN-38 Prodrug in A549 lung cancer cells. We found that Indomethacine-SN-38 (IndoSN-38) and Naproxen-SN-38(NaproSN-38) as a theranostic prodrug targeting cyclooxygenase-2(COX-2) in cancer cells inhibited A549 cell viability in a dose-dependent fashion. IndoSN-38 and NaproSN-38 inhibited A549 cell viability in a dose-dependent fashion. The suppression of A549 cell viability was due to induction of the cell apoptosis by enhancing the activities of Caspase 3 and Caspase 8. The cell cycle arrest of sub-G1 was found in the cells treated with IndoSN-38 or NaproSN-38. Collectively, these data suggested that the anti-proliferative activities of the NSAID-conjugated SN-38 prodrugs were due to promotion of cell death and arresting the cell cycle which was similar with those of SN-38., Competing Interests: The authors declare that they have no competing interests., (© 2023 Published by Elsevier B.V.)

Glycine max Merr . (GM) is a functional food that provides many beneficial phytochemicals. However, scientific evidence of its antidepressive and sedative activities is scarce. The present study was designed to investigate the antidepressive and calmative effects of GM and its biologically active compound, genistein (GE), using electroencephalography (EEG) analysis in an electric foot shock (EFS)-stressed rat. The underlying neural mechanisms of their beneficial effects were determined by assessing corticotropin-releasing factor (CRF), serotonin (5-HT), and c-Fos immunoreactivity in the brain using immunohistochemical methods. In addition, the 5-HT2C receptor binding assay was performed because it is considered a major target of antidepressants and sleep aids. In the binding assay, GM displayed binding affinity to the 5-HT2C receptor (IC50 value of 14.25 ± 11.02 µg/mL). GE exhibited concentration-dependent binding affinity, resulting in the binding of GE to the 5-HT 2C receptor (IC50, 77.28 ± 26.57 mg/mL). Administration of GM (400 mg/kg) increased non-rapid eye movement (NREM) sleep time. Administration of GE (30 mg/kg) decreased wake time and increased rapid eye movement (REM) and NREM sleep in EPS-stressed rats. In addition, treatment with GM and GE significantly decreased c-Fos and CRF expression in the paraventricular nucleus (PVN) and increased 5-HT levels in the dorsal raphe in the brain. Overall, these results suggest that GM and GE have antidepressant-like effects and are effective in sleep maintenance. These results will benefit researchers in developing alternatives to decrease depression and prevent sleep disorders.

Many plants have been used in Korean medicine for treating insomnia. However, scientific evidence for their sedative activity has not been fully investigated. Thus, this study was carried out to investigate the sedative effects of the extracts of medicinal plants, including Yukmijihwang-tang and its various modified forms through the 5-HT2c receptor binding assay, and to further confirm its sleep-promoting effects and the underlying neural mechanism in rats utilizing electroencephalography (EEG) analysis. Enzyme-linked immunosorbent assay (ELISA) was used to measure serotonin (5-HT) in the brain. The water extracts of modified Yukmijihwang-tang (YmP) displayed binding affinity to the 5-HT2C receptor (IC 50 value of 199.9 µg/mL). YmP (50 mg/kg) administration decreased wake time and increased REM and NREM sleep based on EEG data in rats. Additionally, treatment with YmP significantly increased the 5-HT level in the hypothalamus. In conclusion, the sedative effect of YmP can be attributed to the activation of the central serotonergic systems, as evidenced by the high affinity of binding of the 5-HT2C receptor and increased 5-HT levels in the brain of the rat. This study suggests that YmP can be a new material as a sleep inducer in natural products.

Objectives: Snake venom is a complex mixture of various pharmacologically active substances, such as small proteins, peptides, and organic and mineral components. This paper aims to identify and analyse the proteins in common venomous snakes, such as Gloydius blomhoffii ( G. blomhoffii ) and Agkistrodon acutus ( A. acutus ), in Korea., Methods: We used mass spectrometry, electrophoresis, N-terminal sequencing and in-gel digestion to analyse the proteins in these two snake venoms., Results: We identified eight proteins in G. blomhoffii venom and four proteins in A. acutus venom. The proteins detected in G. blomhoffii and A. acutus venoms were phospholipase A2, snake venom metalloproteinase and cysteine-rich secretory protein. Snake C-type lectin (snaclec) was unique to A. acutus venom., Conclusion: These data will contribute to the current knowledge of proteins present in the venoms of viper snakes and provide useful information for investigating their therapeutic potential., Competing Interests: CONFLICT OF INTEREST The authors declare that they have no conflicts of interest., (© 2022 Korean Pharmacopuncture Institute.)

In epidermal growth factor receptor (EGFR) mutant non-small cell lung cancer (NSCLC), acquired resistance to EGFR tyrosine kinase inhibitors (TKI) leads to disease progression. Strategies to overcome the resistance are required in treatment for advanced lung cancer. In this study, we investigated the therapeutic effect of afatinib and HangAmDan-B1 (HAD-B1) co-administration in gefitinib-resistant NSCLC using HCC827-GR, NSCLC cell line with gefitinib resistance, and the HCC827-GR cell implanted mouse model. HAD-B1 consists of 4 herbs, Panax notoginseng Radix, Cordyceps militaris , Panax ginseng C. A. Mey, and Boswellia carteri Birdwood, and has been reported to be effective in patients with advanced lung cancer in clinical practice. Our findings demonstrated that HAD-B1 combined with afatinib markedly inhibited cell proliferation and induced apoptosis compared to afatinib monotherapy and HAD-B1 monotherapy. Inhibition of HCC827-GR cell proliferation by HAD-B1 occurred through MET amplification and reduced phosphorylation, and the synergistic effect of afatinib and HAD-B1 induced cell cycle arrest and apoptosis in HCC827-GR cells via the downregulation of ERK and mTOR signaling pathways. In hematology and biochemistry tests, HAD-B1 alleviated the toxicity of tumor. In conclusion, HAD-B1 combined with afatinib would be a promising therapeutic strategy for NSCLC with EGFR-TKI resistance.

We investigated the antitumor effect of Cordyceps militaris extract (CME) on A549 cisplatin-resistant (CR) lung cancer cells. The proliferation of A549/CR cells was suppressed by CME. Apoptosis of the cells was induced by CME. The cell cycle arrest was observed in the sub-G1 phase in the cells treated with CME. Proteomic profile analysis showed that H-Ras was downregulated in CME-treated cells and it was confirmed by western blot analysis. Collectively, these data demonstrated that CME is an alternative treatment for the anticancer effect.

The scaffolding protein receptor for activated C-kinase 1 (RACK1) mediates receptor activator of nuclear factor κΒ ligand (RANKL)-dependent activation of p38 MAPK in osteoclast precursors; however, the role of RACK1 in mature osteoclasts is unclear. The aim of our study was to identify the interaction between RACK1 and c-Src that is critical for osteoclast function. A RACK1 mutant protein (mutations of tyrosine 228 and 246 residues to phenylalanine; RACK1 Y228F/Y246F) did not interact with c-Src. The mutant retained its ability to differentiate into osteoclasts; however, the integrity of the RANKL-mediated cytoskeleton, bone resorption activity, and phosphorylation of c-Src was significantly decreased. Importantly, lysine 152 (K152) within the Src homology 2 (SH2) domain of c-Src is involved in RACK1 binding. The c-Src K152R mutant (mutation of lysine 152 into arginine) impaired the resorption of bone by osteoclasts. These findings not only clarify the role of the RACK1-c-Src axis as a key regulator of osteoclast function but will also help to develop new antiresorption therapies to prevent bone loss-related diseases.

[This corrects the article DOI: 10.1155/2018/8249563.].

Periodontitis is associated with a dysbiotic shift in the oral microbiome. Vaccine approaches to prevent microbial shifts from healthy to diseased state in oral biofilms would provide a fundamental therapeutic strategy against periodontitis. Since dental plaque formation is a polymicrobial and multilayered process, vaccines targeting single bacterial species would have limited efficacy in clinical applications. In this study, we developed a divalent mucosal vaccine consisting of a mixture of FlaB-tFomA and Hgp44-FlaB fusion proteins targeting virulence factors of inflammophilic bacteria Fusobacterium nucleatum and Porphyromonas gingivalis, respectively. Introduction of peptide linkers between FlaB and antigen improved the stability and immunogenicity of engineered vaccine antigens. The intranasal immunization of divalent vaccine induced protective immune responses inhibiting alveolar bone loss elicited by F. nucleatum and P. gingivalis infection. The built-in flagellin adjuvant fused to protective antigens enhanced antigen-specific antibody responses and class switch recombination. The divalent vaccine antisera recognized natural forms of surface antigens and reacted with diverse clinical isolates of Fusobacterium subspecies and P. gingivalis. The antisera inhibited F. nucleatum-mediated biofilm formation, co-aggregation of P. gingivalis and Treponema denticola, and P. gingivalis-host cell interactions. Taken together, the built-in adjuvant-engineered mucosal vaccine provides a technological platform for multivalent periodontitis vaccines targeting dysbiotic microbiome.

Epidermal growth factor receptor mutation-positive non-small cell lung cancer is cared for mainly by target therapeutics in the clinical treatment at present. We investigated the antitumor effect of HangAmDan-B1 (HAD-B1) combined with afatinib on H1975 (L858R/T790M double mutation) lung cancer cells. The combined treatment of HAD-B1 with afatinib inhibited the proliferation of H1975 cells in a dose-dependent manner compared with the treatment of afatinib or HAD-B1 alone. The combined treatment group significantly induced early apoptosis and cell cycle arrest of the cells compared with afatinib- or HAD-B1-treated control group. Profile analysis of cell cycle proteins in H1975 cells treated with the combination of HAD-B1 and afatinib using InnoPharmaScreen antibody microarray showed downregulation of pERK1/2 and upregulation of p16 in the cells. In vivo tumor growth assay in xenograft animal model of human H1975 lung cancer cells revealed that the mean tumor volume in the group treated with the combination of HAD-B1 and afatinib showed a significant reduction compared with the control groups.

Prolonged exposure to stress can affect mood and cognition and lead to mood disorders. Research on stress-associated mood disorders is important in modern society as people are increasingly exposed to unavoidable stressors. We used a mouse model with 2 weeks of exposure to electric foot shock and restraint, to determine the effect of Fraxinus rhynchophylla Hance (FX) extract on chronic stress-induced depression. We measured the effect of FX extract using various physiological, behavioral, and biochemical measures. FX extract ameliorated chronic stress-induced body and relative liver weight loss and improved depressive-like behaviors in the open field and forced swim tests. In addition, plasma cortisol and serotonin levels in stress-induced mice following FX treatment were similar to normal mice, and the elevation of proinflammatory cytokines was prevented. Moreover, FX treatment increased the expression of phosphorylated cyclic adenosine-3',5'-monophosphate response element-binding protein (pCREB)/brain-derived neurotrophic factor (BDNF). Further experiments confirmed the efficacy of FX extract by showing similar results using esculin and esculetin, compounds extracted from FX. Taken together, these results indicate that FX extract has an antidepressant effect on chronic stress-induced depression by associating signaling with neuroinflammation and neurogenesis.

Chronic periodontitis is caused by interactions between the oral polymicrobial community and host factors. Periodontal diseases are associated with dysbiotic shift in oral microbiota. Vaccination against periodontopathic bacteria could be a fundamental therapeutic to modulate polymicrobial biofilms. Because oral cavity is the site of periodontopathic bacterial colonization, mucosal vaccines should provide better protection than vaccines administered systemically. We previously reported that bacterial flagellin is an excellent mucosal adjuvant. In this study, we investigated whether mucosal immunization with a flagellin-adjuvanted polypeptide vaccine induces protective immune responses using a Porphyromonas gingivalis infection model. We used the Hgp44 domain polypeptide of Arg-gingipain A (RgpA) as a mucosal antigen. Intranasal (IN) immunization induced a significantly higher Hgp44-specific IgG titer in the serum of mice than sublingual (SL) administration. The co-administration of flagellin potentiated serum IgG responses for both the IN and SL vaccinations. On the other hand, the anti-Hgp44-specific IgA titer in the saliva was comparable between IN and SL vaccinations, suggesting SL administration as more compliant vaccination route for periodontal vaccines. The co-administration of flagellin significantly potentiated the secretory IgA response in saliva also. Furthermore, mice administered a mixture of Hgp44 and flagellin via the IN and SL routes exhibited significant reductions in alveolar bone loss induced by live P. gingivalis infections. An intranasally administered Hgp44-flagellin fusion protein induced a comparable level of Hgp44-specific antibody responses to the mixture of Hgp44 and flagellin. Overall, a flagellin-adjuvanted Hgp44 antigen would serve an important component for a multivalent mucosal vaccine against polymicrobial periodontitis.

We investigated the antiangiogenic effects of Lindera obtusiloba Blume (Hwangmaemok, HMM), which is a plant in the Lauraceae family that is commonly used to treat colds and gastritis. Moreover, given that a recent study reported the inhibitory effects of HMM extract on cancer metastasis, we hypothesized that HMM extract might possess and antiangiogenic function. Thus, this study was conducted to investigate the effects of HMM extract on endothelial cell proliferation, migration, and neovascularization in chick chorioallantoic membrane (CAM), and investigated the molecular mechanism of antiangiogenesis using a ProteoChip-based proteomics technology. To examine the effects of HMM extracts on endothelial cell proliferation and migration, we conducted basic fibroblast growth factor (bFGF)-induced human umbilical vein endothelial cell (HUVEC) proliferation and migration. To assess the molecular mechanism of the antiangiogenic effects of HMM extract, a ProteoChip-based forwarded phase antibody array was employed to identify the differential expression of cell cycle proteins in HMM-treated HUVECs. HMM extract inhibited bFGF-induced HUVEC proliferation and migration in a dose-dependent manner and CAM angiogenesis. The ProteoChip-based antibody microarray data showed upregulation of Nibrin/NBS1 and downregulation of Plk-1 and Cyclin E, which are involved in cell division and controlling the cell cycle in bFGF-induced HUVECs. These data suggest that HMM may be a potent antitumor medicinal herb. The present study demonstrates that the antiangiogenic effect of HMM may be due to suppression of endothelial cell proliferation and migration. Taken together, these results emphasize the potential to use HMM extract as a potent angiogenesis inhibitor to treat cancer.

Small molecule-inhibition targeting protein-protein interaction (PPI) is now recognized as an emerging and challenging area in drug design. We developed a novel interactive drug discovery methodology known as Protein Chip technology (ProteoChip) as a cutting-edge PPI assay system applicable for unique PPI-targeting therapeutics integrated with computer-aided drug design (CADD). Here, we describe a novel small molecular PPI inhibitor, IPS-02001, which the blocks integrin αvβ3-osteopontin interface a novel PPI inhibitor identified by the interactive methodology of both ProteoChip- and CADD-based PPI assay. IPS-02001 (6,7-Dichloro-2,3,5,8-tetrahydroxy-1,4-naphthoquinone) was screened from different compound libraries (InterBioScreen, Commercial libraries) using an in silico structure-based molecular docking simulation method and a protein chip-based protein-protein interaction assay system. Additionally, integrin αvβ3, an adhesion receptor expressed in osteoclasts (OCs), was implicated in the regulation of OC function via regulation of the cytoskeletal organization of OCs. IPS-02001 blocked OC maturation from murine bone marrow-derived macrophages, as well as the resorptive function of OCs. Moreover, treatment with IPS-02001 impaired downstream signaling of integrin αvβ3 linked to Pyk2, c-Src, PLCγ2, and Vav3 and disrupted the actin cytoskeleton in mature OCs. Furthermore, IPS-02001 blocked RANKL-induced bone destruction by reducing the number of OCs and protected against ovariectomy-induced bone loss in mice. Thus, IPS-02001 may represent a promising new class of anti-resorptive drugs for treatment of bone diseases associated with increased OC function., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Purpose: The aim of this study was to evaluate antibacterial activity and osteoblast-like cell viability according to the ratio of titanium nitride and zirconium nitride coating on commercially pure titanium using an arc ion plating system., Materials and Methods: Polished titanium surfaces were used as controls. Surface topography was observed by scanning electron microscopy, and surface roughness was measured using a two-dimensional contact stylus profilometer. Antibacterial activity was evaluated against Streptococcus mutans and Porphyromonas gingivalis with the colony-forming unit assay. Cell compatibility, mRNA expression, and morphology related to human osteoblast-like cells (MG-63) on the coated specimens were determined by the XTT assay and reverse transcriptase-polymerase chain reaction., Results: The number of S. mutans colonies on the TiN, ZrN and (Ti1-xZrx)N coated surface decreased significantly compared to those on the non-coated titanium surface (P<0.05)., Conclusion: The number of P. gingivalis colonies on all surfaces showed no significant differences. TiN, ZrN and (Ti1-xZrx)N coated titanium showed antibacterial activity against S. mutans related to initial biofilm formation but not P. gingivalis associated with advanced periimplantitis, and did not influence osteoblast-like cell viability.

P11, a novel peptide ligand containing a PDZ-binding motif (Ser-Asp-Val) with high affinity to integrin α(v)β(3) was identified from a hexapeptide library (PS-SPCL) using a protein microarray chip-based screening system. Here, we investigated the inhibitory mechanism of P11 (HSDVHK) on tumor-induced angiogenesis via a pharmacoproteomic approach. P11 was rapidly internalized by, human umbilical vein endothelial cells (HUVECs) via an integrin α(v)β(3)-mediated event. Caveolin and clathrin appeared to be involved in the P11 uptake process. The cell-penetrating P11 resulted in suppression of bFGF-induced HUVEC proliferation in a dose-dependent manner. Phosphorylation of extracellular-signal regulated kinase (ERK1/2) and mitogen-activated protein kinase kinase (MEK) in bFGF-stimulated HUVECs was inhibited by cell-permeable P11. Proteomic analysis via antibody microarray showed up-regulation of p53 in P11-treated HUVECs, resulting in induction of apoptosis via activation of caspases-3, -8, and -9. Several lines of experimental evidence strongly suggest that the molecular mechanism of P11, a novel anti-angiogenic agent, inhibits bFGF-induced HUVEC proliferation via mitogen-activated protein kinase kinase and extracellular-signal regulated kinase inhibition as well as p53-mediated apoptosis related with activation of caspases.

Cisplatin causes auditory impairment due to the apoptosis of auditory hair cells. There is no strategy to regulate ototoxicity by cisplatin thus far. Dansam-Eum (DSE) has been used for treating the central nerve system injury including hearing loss in Korea. However, disease-related scientific investigation by DSE has not been elucidated. Here, we demonstrated that DSE and its component rosmarinic acid (RA) were shown to inhibit apoptosis of the primary organ of Corti explants as well as the auditory cells. Administration of DSE and RA reduced the thresholds of the auditory brainstem response in cisplatin-injected mice. A molecular docking simulation and a kinetic assay show that RA controls the activity of caspase-1 by interaction with the active site of caspase-1. Pretreatment of RA inhibited caspase-1 downstream signal pathway, such as the activation of caspase-3 and 9, release of cytochrome c, translocation of apoptosis-inducing factor, up-regulation of Bax, down-regulation of Bcl-2, generation of reactive oxygen species, and activation of nuclear factor-κB. Anticancer activity by cisplatin was not affected by treatment with RA in SNU668, A549, HCT116, and HeLa cells but not B16F10 cells. These findings show that blocking a critical step by RA in apoptosis may be useful strategy to prevent harmful side effects of ototoxicity in patients with having to undergo chemotherapy.

We investigated the anti-angiogenic effects of the water extract of HangAmDan (WEHAD), which is a crude extract of nine Korean medicinal substances of animal and plant origin. In human umbilical vein endothelial cells, WEHAD significantly inhibited bFGF-induced proliferation, adhesion, migration, and capillary tube formation. We used an antibody array to perform an analysis of signaling proteins, which showed up-regulated expression of various proteins including RAD51, RAD52, and p73, and down-regulated expression of pFAK. Blood vessel formation in a chick chorioallantoic membrane (CAM) treated with WEHAD was markedly reduced in length compared with a PBS-treated control group. These results suggest that inhibition of angiogenesis by WEHAD may be the mechanism of action for the anti-cancer effects of HAD.

Pharmacological agents that are safe and can sensitize the lung cancer are urgently required. We investigated whether Atractylenolide III (ATL-III), the major component of Atractylodes rhizome can induce apoptosis of the lung carcinoma cells. ATL-III inhibited cell growth, increased lactate dehydrogenase release and modulated cell cycle on human lung carcinoma A549 cells. ALT-III induced the activation of caspase-3 and caspase-9 and cleavage of poly-(ADP)-ribose polymerase. ATL-III induced the release of cytochrome c, upregulation of bax expression, and translocation of apoptosis-inducing factor. In addition, ATL-III inhibited the proliferation and capillary tube formation of human umbilical vein endothelial cells. These data indicate that ATL-III is a potential candidate for treatment of human lung carcinoma., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

USP17, a deubiquitinating enzyme (DUB), belongs to the ubiquitin-specific protease protein family. USP17 has highly conserved Cys, His, and Asp domains responsible for the deubiquitinating activity, and two hyaluronan binding motifs in its sequence. We reported that the presence of hyaluronan binding motifs in USP17 is responsible for the cell viability of cancerous cells. Although many if not all of the DUBs contain hyaluronan binding motifs in its sequence, the actual mechanism of hyaluronan binding motifs regulating signal transduction remains unclear. USP17 also regulates Ras activation and cell proliferation by inhibiting RCE1 activity. Thus USP17 may have a critical role in regulating tumor pathway through its deubiquitinating activity on various tumor-related proteins. This study reports the generation and characterization of polyclonal and six monoclonal antibodies against USP17 through immunoblotting, immunoprecipitation, and immunofluorescence microscopy analyses. These antibodies of USP17 will be useful as tools to investigate the mechanism of hyaluronan-mediated tumor suppression and also to screen interacting oncogenic substrates by immunoprecipitation assay.

In the present study, we extracted Corydalis heterocarpa with various solvents in order to find the bioactive constituents that demonstrated anti-inflammatory effects. We isolated the active compound, Columbianetin. Anti-inflammatory effect of Columbianetin has been reported but the precise effects of Columbianetin in experimental models have remained unknown. In the present study, we investigate the effect of Columbianetin on the production of histamine, interleukin (IL)-1beta, IL-6, IL-8, and tumor necrosis factor (TNF)-alpha and expression of cyclooxygenase-2 (COX-2) by using the human mast cell line (HMC-1). Various concentrations of Columbianetin were treated before the activation of HMC-1 cells with phorbol 12-myristate 13-acetate (PMA) plus calcium ionophore, A23187. PMA plus A23187 significantly increased IL-1beta, IL-6, IL-8, and TNF-alpha production compared with media control (p<0.05). We also show that the increased cytokines IL-1beta, IL-6, IL-8, and TNF-alpha level was significantly inhibited by Columbianetin in a dose-dependent manner (p<0.05). Maximal inhibition rates of IL-1beta, IL-6, IL-8, and TNF-alpha production by Columbianetin were about 102.6%, 101.1%, 95.8%, and 103.9%, respectively. Columbianetin inhibited expression of COX-2. In addition, the effect of Columbianetin was investigated on the histamine release from HMC-1 stimulated by substance P, which promotes histamine release. Columbianetin also inhibited the histamine release by substance P. In conclusion, these results indicate that Columbianetin may be helpful in regulating mast cell-mediated allergic inflammatory responses.

Histidine decarboxylase (HDC) catalyzes the formation of histamine from histidine. Histamine has various effects in physiological and pathological reactions, such as inflammation, cell growth, and neuro-transmission. We investigated the role of hypoxia-inducible factor (HIF)-1 on hypoxia-induced HDC expression in human mast cell line, HMC-1 cells and mouse bone marrow-derived mast cells (BMMCs). Hypoxia significantly increased histamine production. HDC expression and activity were induced by hypoxia. Additionally, when cells were transfected with a native form of HIF-1alpha, hypoxia could induce higher HDC expression than in the nontransfected cell. HIF-1 binding activity for HDC 5' flanking region (HFR) was similar to that for the hypoxia-responsive element. Using HDC promoter deletion analysis, we also demonstrated that HFR was regulated by HIF-1 activation. In addition, depletion of HIF-1alpha prevents hypoxic induction of HDC in BMMCs. In conclusion, these results demonstrate that hypoxia induces HDC expression by transcriptional mechanisms dependent upon HIF-1.

Protein microarray is an emerging technology that makes high-throughput analysis possible for protein-protein interactions and analysis of proteome and biomarkers in parallel. The authors investigated the application of a novel protein microarray chip, ProteoChip, in new drug discovery. Integrin alpha(v)beta(3) microarray immobilized on the ProteoChip was employed to screen new active peptides against the integrin from multiple hexapeptide sublibraries of a positional scanning synthetic peptide combinatorial library (PS-SPCL). The integrin alpha(v)beta(3)-vitronectin interaction was successfully demonstrated on the integrin microarray in a dose-dependent manner and was inhibited not only by the synthetic RGD peptide but also by various integrin antagonists on the integrin microarray chip. Novel peptide ligands with high affinity to the integrin were also identified from the peptide libraries with this chip-based screening system by a competitive inhibition assay in a simultaneous and high-throughput fashion. The authors have confirmed antiangiogenic functions of the novel peptides thus screened through an in vitro and in vivo angiogenesis assay. These results provide evidence that the ProteoChip is a promising tool for high-throughput screening of lead molecules in new drug development.

An after-cataract is caused by the proliferation of residual cells over the equator of the lens. These cells subsequently migrate to the posterior lens capsule, where they undergo aberrant differentiation into fiber-like cells or transdifferentiation into fibroblast-like cells. To study the precise molecular mechanisms of transdifferentiation, an attempt was made to establish an in vitro system, in which the lens epithelial cells (LECs) of the pre-equatorial zone could be transdifferentiated into fibroblast-like cells. The required conditions for culturing the LECs were identified as consisting of four phases; intact bovine explants, explant-cultured, serum-modulated and additionally modulated LECs. The LECs of each phase were compared by examining changes in the expression of the epithelial-mesenchymal transition (EMT)-related genes and changes in cellular morphology and adhesion. The explants that were cultured in a medium containing 10% fetal bovine serum (FBS) for 2 weeks, showed changes in the expression of the EMT-related genes, although the other explant-cultured cells maintained an epithelial morphology. To introduce a transition into mesenchymal cells, the explant cultures were subcultured in a medium containing 20% FBS for six passages. These cells displayed an elongated morphology and were able to grow and migrate in a similar way to fibroblast cells. The expression of the EMT-related genes, such as, extracellular matrix proteins and integrins, was altered. This was similar to the alteration of the 3-dimensional collagen gels model previously reported. During a further process of EMT by additional serum modulation, the inhibitory effect of disintegrin on cell adhesion was gradually decreased, integrin expression was differentially regulated and alpha-smooth muscle actin was post-translationally modified from the point of passage number six. Overall, it can be concluded that terminal transdifferentiation accompanies changes in the cytoskeletal proteins and cell surface molecules. These are modulated in systematic patterns of post-transcriptional and post-translational regulation and patterns of gene regulation, by the synergic effects of several transforming factors contained in serum. Therefore, posterior capsular opacification may also be accompanied by this molecular mechanism.

The effects of LY294002 (LY29) and wortmannin (WM), inhibitors of phosphatidylinositol 3-kinase (PI3K), on monocyte chemoattractant protein-1 (MCP-1) expression by human umbilical vein endothelial cells were investigated. Complete inhibition of interleukin (IL)-1beta-induced Akt phosphorylation occurred at 50 microM LY29 or 100 nM WM. At these concentrations, LY29, but not WM, significantly inhibited constitutive and IL-1beta-induced MCP-1 expression at both protein and mRNA levels. LY303511 (LY30), an inactive analogue of LY29, also inhibited MCP-1 expression. LY29 and LY30 inhibited activation of nuclear factor-kappaB (NF-kappaB). These results suggest that LY29 inhibits MCP-1 expression at least in part via suppression of NF-kappaB, independent of PI3K, and the structure of LY29 and LY30 may be a novel template for development of new anti-inflammatory drugs.

Salmosin is a snake venom-derived novel disintegrin that antagonizes platelet aggregation. In this study, we investigated its functional specificity in tumor angiogenesis. Salmosin significantly inhibited bovine capillary endothelial cell proliferation induced by basic fibroblast growth factor but had no effect on normal growth of the cell. The basic fibroblast growth factor-induced in vivo angiogenesis in the chorioallantoic membrane was disrupted by salmosin treatment without affecting normal embryonic angiogenesis. Adhesion of the bovine capillary endothelial cells to vitronectin was also inhibited by the binding of salmosin to the alpha(v)beta3 integrin. Both the metastatic-tumor growth and the solid-tumor growth that developed in mice were effectively suppressed by salmosin treatment. Several lines of experimental evidence strongly suggest that the tumor-specific antiangiogenic activity of salmosin disrupts tumor growth by blocking the alpha(v)beta3 integrin that is expressed on the vascular endothelial cell surface.

Reactive oxygen species such as superoxides, hydrogen peroxide (H2O2) and hydroxyl radicals have been suggested to be involved in the catalytic action of nitric oxide synthase (NOS) to produce NO from L-arginine. An examination was conducted on the effects of oxygen radical scavengers and oxygen radical-generating systems on the activity of neuronal NOS and guanylate cyclase (GC) in rat brains and NOS from the activated murine macrophage cell line J774. Catalase and superoxide dismutase (SOD) showed no significant effects on NOS or GC activity. Nitroblue tetrazolium (NBT, known as a superoxide radical scavenger) and peroxidase (POD) inhibited NOS, but their inhibitory actions were removed by increasing the concentration of arginine or NADPH respectively, in the reaction mixture. NOS and NO-dependent GC were inactivated by ascorbate/FeSO4 (a metal-catalyzed oxidation system), 2'2'-azobis-amidinopropane (a peroxy radical producer), and xanthine/xanthine oxidase (a superoxide generating system). The effects of oxygen radicals or antioxidants on the two isoforms of NOS were almost similar. However, H2O2 activated GC in a dose-dependent manner from 100 microM to 1 mM without significant effects on NOS. H2O2-induced GC activation was blocked by catalase. These results suggested that oxygen radicals inhibited NOS and GC, but H2O2 could activate GC directly.