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Polylysines have been frequently used in drug delivery and antimicrobial and cell adhesion studies. Because of steric hindrance, chirality plays a major role in the functional difference between poly-l-lysine (PLL) and poly-d-lysine (PDL), especially when they interact with the plasma membranes of mammalian cells. Therefore, it is speculated that the interaction between chiral polylysines and the plasma membrane may cause different cellular behaviors. Here, we carefully investigated the interaction pattern of PLL and PDL with plasma membranes. We found that PDL could be anchored onto the plasma membrane and interact with the membrane lipids, leading to the rapid morphological change and death of A549 cells (a human lung cancer cell line) and HPAEpiCs (a human pulmonary alveolar epithelial cell line). In contrast, PLL exhibited good cytocompatibility and was not anchored onto the plasma membranes of these cells. Unlike PLL, PDL could trigger protective autophagy to prevent cells in a certain degree, and the PDL-caused cell death occurred

Background: Interleukin (IL)-18 is produced mainly in the heart and can be associated with the development of cardiac hypertrophy that leads to cardiac dysfunction. However, the effects of hypoxia on IL-18 expression and atrial natriuretic factor (ANF) secretion remain largely unknown. Aim: The aim of this study was to assess the effect of hypoxia on IL-18 production and its role in ANF secretion by using an isolated perfused beating rat atrial model. Methods: The level of ANF in the perfusates was determined by radioimmunoassay, and the protein levels of Src, IL-18 and its receptors (IL-18-Rα and IL-18-Rβ), Rho guanine nucleotide exchange factor (RhoGEF) and RhoA, activating transcription factor 3 (ATF3), T cell factor (TCF) 3 and 4, and lymphoid enhancer factor (LEF) 1 in atrial tissue samples were detected by Western blotting. Results: Hypoxia significantly upregulated the expression of the non-receptor tyrosine kinase Src, and this effect was blocked by endothelin-1 receptor type A (BQ123) and type B (BQ788) antagonists. Hypoxia also enhanced the expression of RhoGEF and RhoA concomitantly with the upregulation of IL-18, IL-18-Rα and IL-18-Rβ. The hypoxia-induced RhoGEF and RhoA were abolished by Src inhibitor 1 (SrcI), and the protein levels of IL-18 and its two receptors were also blocked by SrcI. Moreover, the hypoxia-induced expression levels of ATF3, TCF3, TCF4 and LEF1 were repealed by IL-18 binding protein, and the hypoxia-promoted secretion of ANF was also obviously attenuated by this binding protein. Conclusions: These findings imply that Src-IL-18 signaling is involved in the release of ANF in hypoxic beating rat atria.

When attacked by herbivores, plants emit volatiles to attract parasitoids and predators of herbivores. However, our understanding of the effect of plant volatiles on the subsequent behaviour of conspecific parasitoids when herbivores on plants are parasitized is limited. In this study, rice plants were infested with gravid females of the brown planthopper (BPH) Nilaparvata lugens for 24 hr followed by another 24 hr in which the BPH eggs on plants were permitted to be parasitized by their egg parasitoid, Anagrus nilaparvatae; volatiles from rice plants that underwent such treatment were less attractive to subsequent conspecific parasitoids compared to the volatiles from plants infested with gravid BPH females alone. Chemical analysis revealed that levels of JA and JA-Ile as well as of four volatile compounds-linalool, MeSA, α-zingiberene and an unknown compound-from plants infested with BPH and parasitized by wasps were significantly higher than levels of these compounds from BPH-infested plants. Laboratory and field bioassays revealed that one of the four increased chemicals-α-zingiberene-reduced the plant's attractiveness to the parasitoid. These results suggest that host plants can fine-tune their volatiles to help egg parasitoids distinguish host habitats with parasitized hosts from those without.

The application of a coordination container in biomedicine is hindered by single binding domains and unsatisfactory biostability and biocompatibility. Herein, we designed a sulfonylcalix[4]arene-based decahexanuclear zinc(II) coordination container employing a flexible tetracarboxylate ligand as a linker and utilized it as a novel drug delivery system. The coordination container consisting of one endo and four exo cavities provides multiple binding domains for efficient encapsulation of drug molecules as clearly revealed by systematic host-guest studies using NMR techniques of 1H NMR titration experiments and 2D NOESY and diffusion-ordered NMR spectroscopy studies. Incorporation of a flexible p-phenylene-bis(methanamino) spacer into the container via the carboxylate linker allowed a stepwise drug loading process through sequential binding at endo and exo cavities, as well as enabling pH-responsive stepwise drug release. The drug-loaded coordination container not only exhibits excellent biostability and biocompatibility but also provides encouraging therapeutic efficiency toward inflammatory macrophages as revealed by in vitro studies. The novel strategy for engineering the endo cavity of a coordination container provides a new approach to achieving controlled drug delivery and opens up new opportunities for designing novel functional supramolecular materials.

More than 95% of all human genes are alternatively spliced after transcription, which enriches the diversity of proteins and regulates transcript and/or protein levels. The splicing isoforms produced from the same gene can manifest distinctly, even exerting opposite effects. Mounting evidence indicates that the alternative splicing (AS) mechanism is ubiquitous in various cancers and drives the generation and maintenance of various hallmarks of cancer, such as enhanced proliferation, inhibited apoptosis, invasion and metastasis, and angiogenesis. Splicing factors (SFs) play pivotal roles in the recognition of splice sites and the assembly of spliceosomes during AS. In this review, we mainly discuss the similarities and differences of SF domains, the details of SF function in AS, the effect of SF-driven pathological AS on different hallmarks of cancer, and the main drivers of SF expression level and subcellular localization. In addition, we briefly introduce the application prospects of targeted therapeutic strategies, including small-molecule inhibitors, siRNAs and splice-switching oligonucleotides (SSOs), from three perspectives (drivers, SFs and pathological AS). Finally, we share our insights into the potential direction of research on SF-centric AS-related regulatory networks.

To study the protective effects of azithromycin on renal damage induced by doxorubicin and albumin in mice.Forty male BALB/c mice were randomly divided into blank control group (Ctrl group), renal damage model group (ADR+BSA group), azithromycin treated group (Azm group) and prednisone acetate positive control group (Pdn group) in accordance with random number table method. Mice in ADR+BSA, AZM and Pdn group were injected intravenously with 9.8 kgCompared with the Ctrl group, the 24 h urine protein level of the ADR+BSA group was increased significantly (P＜0.05), and the Ccr was decreased significantly (P＜0.05). After the azithromycin treatment, the 24 h urine protein was decreased significantly (P＜0.05), while the Ccr was increased significantly (P＜0.05) compared with ADR+BSA group.Azithromycin has a protective effects on the renal damage induced by doxorubicin and albumin in mice.

Background/Aims: Thymic stromal lymphopoietin (TSLP) is a cytokine that plays diverse roles in the regulation of immune responses. However, a detailed understanding of the TSLP signaling pathway in asthma remains elusive. In this study, we aimed to investigate the role of the TSLP signaling pathway in asthma and its effect on airway inflammation and remodeling. Methods: Forty Sprague Dawley (SD) rats were evenly classified into control, asthma, IgG2a mAb and anti-TSLP mAb groups. Ovalbumin (OVA)-induced asthma models were successfully established. Blood, bronchoalveolar lavage fluid (BALF) and lung tissue samples were prepared. Total BALF leukocytes were counted, and the proportions of different leukocyte types were determined. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) and immunohistochemistry were performed to determine the mRNA and protein levels of TSLP, OX40L, α-smooth muscle actin (α-SMA, a marker of airway remodeling in asthma) and collagen I in the plasma. Enzyme-linked immunosorbent assay (ELISA) was carried out to measure the concentrations of TSLP, OX40L, and other inflammatory factors, such as interferon (IFN)-γ, interleukin (IL)-4, IL-5 and IL-13, in the plasma. Results: Compared with the control group, there were more leukocytes, increased EOS and LYM proportions, higher Underwood and PAS scores, increased WTt, WTm, WAt/A0, WAm/WAt, WTt/R0, WTm/WTt, TSLP, OX40L, a-SMA and collagen I mRNA and protein levels, and higher SLP, OX40L, IL-4, IL-5 and IL-13 levels, but lower MON proportions and IFN-γ levels in the asthma and IgG2a mAb groups. Compared with the asthma and IgG2a mAb groups, there were less leukocytes, decreased EOS and LYM proportions, lower Underwood and PAS scores, decreased WTt, WTm, WAt/A0, WAm/WAt, WTt/R0, WTm/WTt, TSLP, OX40L, a-SMA and Collagen I mRNA and protein levels, and lower levels of SLP, OX40L, IL-4, IL-5 and IL-13, but higher MON proportions and IFN-γ levels in the anti-TSLP mAb group. WTm and WTt were positively associated with the TSLP, OX40L, α-SMA and collagen-I levels in the rat lung tissues. Conclusion: The results indicate that TSLP may be an important contributor for asthma development as TSLP signaling blockade attenuates airway inflammation and remodeling in asthmatic rats.

Aims Gallic acid (GA) protects against myocardial ischemia-reperfusion (I/R) injury, prevents cardiac hypertrophy and fibrosis, and has anti-inflammatory activity in the heart. However, its effects in regulating atrial natriuretic peptide (ANP) secretion are unknown. The aim of this study was to determine the function of GA in regulating ANP secretion and atrial dynamics in rat atria. Key findings GA (0.01, 0.05, and 0.1 μmol/L) significantly increased atrial ANP secretion and induced positive inotropy dose-dependently. GA (0.1 μmol/L) also increased plasma level of ANP and hemodynamics in rats. These effects were accompanied by upregulation of atrial protein kinase C subtypes β and e (PKC β and PKC e ), which was completely blocked by LY333531 and EAVSLKPT, antagonists of protein PKC β and PKC e , respectively. GA-induced ANP secretion was also attenuated by Go6983 but not rottlerin, antagonists of PKC α and PKC δ , and the positive inotropy was reversed by Go6983. U-73122, a phospholipase C (PLC) antagonist, mitigated the effects of GA on ANP secretion and mechanical dynamics and downregulated Phospho-PLC β at Ser 537 (pPLC β S537), Phospho-PLC β at Ser 1105 (pPLC β S1105), PKC β and PKC e levels, whereas KN62, an inhibitor of Ca 2 + /calmodulin-dependent kinase II, was not modified the GA-induced ANP secretion and suppressed GA-induced mechanical dynamics. Significance GA promotes ANP secretion and effects positive inotropy with regard to mechanical dynamics through the activation of PLC-PKC signaling in rat atria.

A Gram-stain-negative, aerobic, rod-shaped, yellow-pigment-producing bacterium (designated strain CC-CZW007T) was isolated from seafood samples (sea urchins) at Penghu Island in Taiwan. Strain CC-CZW007Tgrew optimally at pH 7.0 and 30 °C in the presence of 3 % (w/v) NaCl. The novel strain shared highest 16S rRNA gene sequence similarity toVitellibacter vladivostokensisJCM 11732T(96.8 %),Vitellibacter soesokkakensisKCTC 32536T(96.4 %),Vitellibacter nionensisKCTC 32420T(95.8 %) andVitellibacter aestuariiJCM 15496T(95.6 %) and lower sequence similarity to members of other genera. Phylogenetic analyses based on 16S rRNA genes revealed a distinct taxonomic position attained by strain CC-CZW007Twith respect to other species of the genusVitellibacter. The major fatty acids were iso-C15 : 0and iso-C17 : 03-OH. The polar lipid profile was composed of major amounts of phosphatidylethanolamine, unidentified lipids and aminolipids; a moderate amount of aminophospholipid was also detected. The DNA G+C content was 34.7 mol%. The predominant quinone system was menaquinone (MK-6). On the basis of polyphasic taxonomic evidence presented here, strain CC-CZW007Tis proposed to represent a novel species within the genusVitellibacter, for which the nameVitellibacter echinoideorumsp. nov. is proposed. The type strain is CC-CZW007T( = BCRC 80886T = JCM 30378T).

Lipocalin-type prostaglandin D synthase (L-PGDS) and peroxisome proliferator activated receptor γ (PPARγ) play important roles in cardiovascular diseases. Nevertheless, effects of hypoxia-inducible factor 1α (HIF-1α) on L-PGDS and PPARγ protein levels and its role in hypoxia-induced atrial natriuretic peptide (ANP) secretion are unclear. In perfused beating rat atria, we observed that hypoxia significantly increased HIF-1α protein levels and stimulated ANP secretion, while upregulating L-PGDS. Hypoxia-induced ANP secretion was clearly attenuated by HIF-1α antagonist 2-methoxyestradiol, downregulating both HIF-1α and L-PGDS protein levels. It was also attenuated by L-PGDS antagonists, AT-56 and HQL-49, downregulating L-PGDS protein levels. In addition, hypoxia-induced ANP secretion was accompanied by increased PPARγ protein levels and was strongly attenuated by PPARγ antagonist GW9662. Hypoxia-induced increase in atrial PPARγ protein levels were dramatically inhibited by both 2-methoxyestradiol and AT-56. These results indicated that hypoxia promotes ANP secretion, at least in part, by activating HIF-1α-l-PGDS-PPARγ signaling in beating rat atria.

A xenograft brain tumor model was established by the subcutaneous injection of U87MG cells into nude mice to investigate the efficacy of a non-viral vector, arginine-modified polyamidoamine dendrimer (PAMAM-R), in delivering a therapeutic gene, human interferon beta (IFN-β). We used 4',6-diamidino-2-phenylindole staining, the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay, and the caspase-3 activity assay to determine the induction of apoptosis upon transfection with the PAMAM-R/IFN-β gene polyplex in vitro. The polyplex was injected into xenograft brain tumors. Mice treated with PAMAM-R/pORF-IFN-β exhibited a significantly smaller tumor size than control mice and PAMAM-R/pORF treated mice. Hematoxylin/eosin staining and immunohistochemistry with the endothelial growth factor receptor antibody also revealed inhibition of tumor growth. Furthermore, reverse transcription polymerase chain reaction and the TUNEL assay also verified the expression of IFN-β and induction of apoptosis in vivo. These results indicate that the PAMAM-R/pORF-IFN-β polyplex is an effective therapeutic candidate for glioblastoma multiforme due to its selective induction of apoptosis in tumor cells.

Cartilage tissue engineering using cells and biocompatible scaffolds has emerged as a promising approach to repair of cartilage damage. To date, however, no engineered cartilage has proven to be equivalent to native cartilage in terms of biochemical and compression properties, as well as histological features. An alternative strategy for cartilage engineering is to focus on the in vivo regeneration potential of immature engineered cartilage. Here, we used a rabbit model to evaluate the extent to which the maturity of engineered cartilage influenced the remodeling and integration of implanted extracellular matrix scaffolds containing allogenous chondrocytes. Full-thickness osteochondral defects were created in the trochlear groove of New Zealand white rabbits. Left knee defects were left untreated as a control (group 1), and right knee defects were implanted with tissue-engineered cartilage cultured in vitro for 2 days (group 2), 2 weeks (group 3), or 4 weeks (group 4). Histological, chemical, and compression assays of engineered cartilage in vitro showed that biochemical composition became more cartilagenous, and biomechanical property for compression gradually increased with culture time. In an in vivo study, gross imaging and histological observation at 1 and 3 months after implanting in vitro-cultured engineered cartilage showed that defects in groups 3 and 4 were repaired with hyaline cartilage-like tissue, whereas defects were only partially filled with fibrocartilage after 1 month in groups 1 and 2. At 3 months, group 4 showed striking features of hyaline cartilage tissue, with a mature matrix and a columnar arrangement of chondrocytes. Zonal distribution of type II collagen was most prominent, and the International Cartilage Repair Society score was also highest at this time. In addition, the subchondral bone was well ossified. In conclusion, in vivo engineered cartilage was remodeled when implanted; however, its extent to maturity varied with cultivation period. Our results showed that the more matured the engineered cartilage was, the better repaired the osteochondral defect was, highlighting the importance of the in vitro cultivation period.

Background Haemodialysis vascular access dysfunction caused by aggressive venous neointimal hyperplasia is a major problem for haemodialysis patients with synthetic arteriovenous (AV) grafts. Several different strategies to prevent venous stenosis by inhibiting smooth muscle cell proliferation and migration using local delivery of potent antiproliferative agents are currently under investigation. We performed this study to evaluate the efficacy of sirolimus-eluting vascular grafts in preventing stenosis and to compare the effectiveness of sirolimus-coated grafts with that of paclitaxel-coated vascular grafts that we characterized in a previous study. Methods AV grafts were implanted laterally between the common carotid artery and external jugular vein of 14 female Landrace pigs. Three types of grafts were implanted: grafts coated with 1.08 μg/mm(2) sirolimus (low dose, n = 6), grafts coated with 2.41 μg/mm(2) sirolimus (high dose, n = 2) and uncoated control grafts (n = 6). Animals were sacrificed 6 weeks after surgery. Cross-sections of the venous anastomoses were analysed to determine the percentage of luminal stenosis in each group, and immunohistochemistry was performed to identify the cellular phenotypes of the neointimal hyperplasia and tissues adjacent to the implanted grafts. Results Compared with the control group, neointimal hyperplasia in the venous anastomoses of the groups implanted with sirolimus-coated vascular grafts was significantly suppressed without infection. The mean ± standard error values for the percentage of luminal stenosis were 75.7 ± 12.7% in the control group and 22.2 ± 1.41% in the low-dose sirolimus-coated group. Myofibroblasts and fibroblasts were the major cell types found in the neointimal hyperplasia. Conclusions Neointimal hyperplasia was effectively suppressed by sirolimus-eluting grafts. However, the inhibitory effects of sirolimus-eluting grafts were weaker than those observed for paclitaxel-coated grafts in our previous study.

Protein phosphatase 1 consists of a secondary structure arrangement, conserved in the serine/threonine protein phosphatase gene family, flanked by nonconserved N-terminal and C-terminal domains. The deletion mutant of PP1 with the 8 nonconserved N-terminal residues removed was designated PP1-(9-330). PP1-(9-330) had a higher activity and affinity than PP1 when assayed against four different substrates, and it also demonstrated a 6-fold higher sensitivity to the inhibitor okadaic acid. This suggested that the N-terminal domain suppressed the activity of PP1 and interfered with its inhibition by okadaic acid. The ANS fluorescence intensity of PP1-(9-330) was greater than that of PP1, which implies that the hydrophobic groove running from active site in the truncated PP1 was more hydrophobic than in PP1. Our findings provide evidence that the nonconserved N-terminus of PP1 functions as an important regulatory domain that influences the active site and its pertinent properties.

We designed chiral 2-nitroimidazole derivatives containing a 2-aminomethylene-4-cyclopentene-1,3-dione moiety as antiangiogenic hypoxic cell radiosensitizers. Based on results of molecular orbital calculations, the 2-aminomethylene-4-cyclopentene-1,3-dione moiety is expected to show high electrophilicity comparable to that of the 2-methylene-4-cyclopentene-1,3-dione moiety included in TX-1123 and tyrphostin AG17. We evaluated the antiangiogenic and radiosensitizing effects of the new compounds, along with other biological properties including their activities as hypoxic cytotoxicities and protein tyrosine kinase (PTK) inhibitory activities. Among the compounds tested, 5 (TX-2036) proved to be the strongest antiangiogenic hypoxic cell radiosensitizer. All the other chiral 2-nitroimidazole derivatives having 2-aminomethylene-4-cyclopentene-1,3-dione moiety tested were also antiangiogenic hypoxic cell radiosensitizers. The PTK inhibitory activity of 5 (TX-2036) showed this to be a promising and potent EGFR kinase inhibitor, having an IC(50) value of lower than 2microM. This compound also was an Flt-1 kinase inhibitor having an IC(50) value of lower than 20microM. Our results show that these chiral 2-nitroimidazole derivatives that contain the 2-aminomethylene-4-cyclopentene-1,3-dione moiety as a potent antiangiogenic pharmacophoric descriptor are promising lead candidates for the development of antiangiogenic hypoxic cell radiosensitizers.

The purpose of this study is to evaluate the feasibility of human amniotic membrane (HAM) as a chondrocyte carrier by assessing cell proliferation and maintenance of phenotype in vitro and cartilage regeneration in vivo. Intact HAM was treated with 0.1% trypsin-ethylenediaminetetraacetic acid (EDTA) for 15 min and the epithelial cells removed to make a denuded HAM. Rabbit articular chondrocytes were then seeded on three different HAM substrates: the epithelial side of intact HAM (IHE), basement side of denuded HAM (DHB), and stromal side of denuded HAM (DHS). These cell-substrate specimens were cultured for up to 4 weeks, and cell proliferation rate and phenotypic stability were examined at weeks 1 and 4. While chondrocytes grew in monolayer fashion on the surface of IHE and DHB substrates, the cells seeded in DHS penetrated and spread into the whole thickness of the stromal layer. The proliferating activity of chondrocytes in DHB was continuously up-regulated. A similar proliferating activity was observed in DHS in the first week, which remained stable for up to 4 weeks. The expression of type II collagen gradually increased with time in the DHS group, while it gradually decreased in the DHB group or was not detected at all in the IHE group. These results suggested that denuded HAM was able to support chondrocyte proliferation and maintenance of phenotype in vitro, seemingly more favorable when DHS was used. Based on this data, the DHS with chondrocytes was used to cover rabbit osteochondral defect with the stromal side facing in. The defect area was successfully regenerated with hyaline cartilage in the Safranin-O stain and International Cartilage Repair Society (ICRS) scoring after 8 weeks of implantation. In conclusion, our findings suggest that denuded HAM could be one of the ideal cell carrier matrices for cartilage regeneration.

A Gram-stain-negative, aerobic, non-motile, rod-shaped, flexirubin-producing bacterium, designated strain CC-CZW010T, was isolated from the edible sea urchinTripneustes gratillain Penghu Island, Taiwan. The isolate grew optimally at pH 7.0 and 30 °C in the presence of 2 % (w/v) NaCl. The most closely related strains in terms of 16S rRNA gene sequence similarity wereChryseobacterium taihuenseNBRC 108747T(97.6 %) andChryseobacterium aquaticumKCTC 12483T(96.7 %). Phylogenetic analyses based on 16S rRNA gene sequences revealed a distinct taxonomic position attained by strain CC-CZW010Twith respect to other species of the genusChryseobacterium. Strain CC-CZW010Tpossessed iso-C15 : 0, anteiso-C15 : 0, iso-C17 : 03-OH, summed feature 3 (comprising C16 : 1ω7c/C16 : 1ω6c) and summed feature 9 (comprising C16 : 010-methyl/iso-C17 : 1ω9c) as predominant fatty acids. The major polar lipid profile consisted of phosphatidylethanolamine, two unidentified lipids and five aminolipids. The polyamine pattern contained the major compoundsym-homospermidine. Menaquinone 6 (MK-6) was the predominant respiratory quinone, and the G+C content of the genomic DNA was 36.4 mol%. According to distinct phylogenetic, phenotypic and chemotaxonomic features, strain CC-CZW010Trepresents a novel species of the genusChryseobacterium, for which the nameChryseobacterium echinoideorumsp. nov. is proposed. The type strain is CC-CZW010T( = BCRC 80786T = JCM 30470T).

A Gram-stain-positive, aerobic, non-motile, rod-shaped, yellow-pigment-producing bacterium, (designated strain CC-CZW004T), was isolated from seafood samples (sea urchins) at Penghu Island in Taiwan. Strain CC-CZW004Tgrew optimally at pH 7.0 and 30 °C in the presence of 3 % (w/v) NaCl. Phylogenetic analyses based on 16S rRNA genes revealed a distinct taxonomic position attained by strain CC-CZW004Twith respect to other members of the genusNocardioides. The novel strain shared highest 16S rRNA gene sequence similarities toNocardioides daejeonensisJCM 16922T(96.4 %),Nocardioides pacificusJCM 19260T(96.3 %), andMarmoricola scoriaeJCM 17444T(96.2 %). The major fatty acids of strain CC-CZW004Tconsisted of C17:0, C16:1ω5c, C17:1ω8c`, iso-C16:0and C19:1ω11c/C19:1ω9c(summed feature 6). The diagnostic diamino acid in the cell wall wasll-2,6-diaminopimelic acid. The polar lipid profile was composed of major amounts of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol and aminophospholipid. The DNA G+C content was 73.3 mol%. The predominant quinone system was menaquinone (MK-8). On the basis of polyphasic taxonomic evidences presented here, strain CC-CZW004Trepresents a novel species of the genusNocardioides, for which the nameNocardioides echinoideorumsp. nov. is proposed. The type strain is CC-CZW004T( = BCRC 16974T = JCM 30276T).

Pulp regeneration caused by endogenous cells homing has become the new research spot in endodontics. However, the source of functional cells that are involved in and contributed to the reconstituting process has not been identified. In this study, the possible role of systemical BMSC in pulp regeneration and the effect of stromal cell-derived factor-1 (SDF-1) on stem cell recruitment and angiogenesis were evaluated. 54 mice were divided into three groups: SDF-1 group (subcutaneous pockets containing roots with SDF-1 absorbed neutralized collagen gel and the green fluorescent protein (GFP) positive BMSCs transplantation via the tail vein), SDF-1-free group (pockets containing roots with gel alone and GFP + BMSCs transplantation) and Control group (pockets containing roots with gel alone). The animals were sacrificed after the roots were implanted into subcutaneous pockets for 3 weeks. Histomorphometric analysis was performed to evaluate the regenerated tissue in the canal by hematoxylin and eosin (HE) staining. The homing of the transplanted BMSCs was monitored with a fluorescence microscope and immunohistochemical analysis. The expression of ALP in new formed tissue was detected immunohistochemically. Dental-pulp-like tissue and new vessels were regenerated and GFP-positive BMSCs and expression of ALP could be observed in both SDF-1 group and SDF-1-free group. Furthermore, more GFP+ cells, stronger expression of ALP and stronger angiogenesis were found in the SDF-1 group than in the SDF-1-free group. To conclude, systemic BMSC can home to the root canal and participate in dental-pulp-like tissue regeneration. Intracanal application of SDF-1 may enhance BMSC homing efficiency and angiogenesis.

High molecular weight (MW) hyaluronan (HA) preparation is considered to be more biologically active than HAs of lower MWs. However, many of the HA preparations currently used to treat osteoarthritis (OA) have lower MWs by the enhanced penetration of HA molecules into the synovial lining cells. In this study, we determined the effectiveness of sonophoresis on the delivery of high MW HA into synovial membrane using an animal model of OA. A total of 1000 kDa (HA1000) and 3000 kDa (HA3000) HA were labeled with fluorescein and injected into the knees of rabbits. Low-intensity continuous ultrasound at 1 MHz, 400 mW/cm2 was applied to the knees for 10 min treatment bid. Synovial fluid analysis revealed increased absorption and fluorescence microscopy showed deeper penetration of both HA1000 and HA3000, more so with the latter. Histologic examination indicated that ultrasound treatment resulted in no apparent damage to the synovial membrane. These results suggest that simultaneous sonication with HA injection might compensate for the short half-life of HA. Consequently, this dual treatment would render HA a far more effective tool in the management of OA.

(R)- and (S)-Epichlorohydrins were used to prepare the enantiomers of sterically diverse haloacetylcarbamoyl-2-nitroimidazoles that function as hypoxic cell radiosensitizers. The synthetic design allowed for introduction of a side chain of varying bulk that permitted an examination of the steric effects on enantio-discrimination in biological assay systems. The single stereocenter also connected the two pharmacophores––a 2-nitroimidazole moiety critical to hypoxic cell radiosensitization, and a haloacetylcarbamoyl group to function as an anti-angiogenesis pharmacophore. In the chick embryo chorioallantoic membrane (CAM) assay, the R-enantiomers possessing the bulky p-tert-butylphenyl group showed higher anti-angiogenic activity than the corresponding S-enantiomers, while there were no differences in the activity between the enantiomers containing the less bulky methyl and tert-butyl groups. Among the compounds we report, R-p-tert-butylphenyl-bromoacetylcarbamoyl-2-nitroimidazole, TX-1898, was found to be the most promising candidate for further development of as anti-angiogenic hypoxic cell radiosensitizer.

We designed, synthesized, and evaluated haloacetylcarbamoyl-2-nitroimidazoles, including chloro (KIN-1800, TX-1835, and TX-1836) and bromo derivatives (TX-1844, TX-1845, and TX-1846), as potential hypoxic cell radiosensitizers with antiangiogenic activities. To establish biological function owing to the haloacetylcarbamoyl group in the side-chain, we compared their in vitro radiosensitizing activities with those of their parent 2-nitroimidazoles without haloacetylcarbamoyl groups: misonidazole (MISO), TX-1831, and TX-1832, respectively. Both tert-butoxy substituted derivatives. TX-1835 and TX-1845, were more potent radiosensitizers than TX-1831. The p-tert-butylphenoxy-substituted derivatives, TX-1836 and TX-1846, and the methoxysubstituted derivatives, KIN-1800 and TX-1844, were stronger radiosensitizers than TX-1832 and MISO. We examined the anti-angiogenic activities of these 2-nitroimidazole derivatives containing haloacetylcarbamoyl group by the rat lung endothelial (RLE) cell proliferation assay and chick embryo chorioallantoic membrane (chick CAM) angiogenesis assay and showed that haloacetylcarbamoyl-2-nitroimidazoles were more potent angiogenic inhibitors than the corresponding desacetylcarbamoyl-2-nitroimidazoles. The in vivo chick CAM angiogenesis assay showed that the strong bromoacetylcarbamoyl-2-nitroimidazole radiosensitizers, such as TX-1845 and TX-1846, were the strongest angiogenic inhibitors among them. We concluded that the bromoacetylcarbamoyl-2-nitroimidazole radiosensitizers, such as TX-1845 and TX-1846, are promising as anti-angiogenic hypoxic cell radiosensitizers.

BACKGROUND: Calcium phosphate cement (CPC) can be molded or injected to form a scaffold in situ, which intimately conforms to complex bone defects. Bioactive glass (BG) is known for its unique ability to bond to living bone and promote bone growth. However, it was not until recently that literature was available regarding CPC-BG applied as an injectable graft. In this paper, we reported a novel injectable CPC-BG composite with improved properties caused by the incorporation of BG into CPC. MATERIALS AND METHODS: The novel injectable bioactive cement was evaluated to determine its composition, microstructure, setting time, injectability, compressive strength and behavior in a simulated body fluid (SBF). The in vitro cellular responses of osteoblasts and in vivo tissue responses after the implantation of CPC-BG in femoral condyle defects of rabbits were also investigated. RESULTS: CPC-BG possessed a retarded setting time and markedly better injectability and mechanical properties than CPC. Moreover, a new Ca-deficient apatite layer was deposited on the composite surface after immersing immersion in SBF for 7 days. CPC-BG samples showed significantly improved degradability and bioactivity compared to CPC in simulated body fluid (SBF). In addition, the degrees of cell attachment, proliferation and differentiation on CPC-BG were higher than those on CPC. Macroscopic evaluation, histological evaluation, and micro-computed tomography (micro-CT) analysis showed that CPC-BG enhanced the efficiency of new bone formation in comparison with CPC. CONCLUSIONS: A novel CPC-BG composite has been synthesized with improved properties exhibiting promising prospects for bone regeneration.

Summary Objectives Functional role of Annexin A1 in tumorigenesis is poorly understood. The aim of this study was to investigate the relationship between Annexin A1 protein expression and pathological differentiation grade in biopsy samples from a large cohort of patients with oral squamous cell carcinoma (OSCC); and to evaluate the potential role of Annexin A1 on cell proliferation and tumorigenesis of OSCC. Materials and methods We investigated the relationship between Annexin A1 expression by immunohistochemical staining and pathological differentiation grade of biopsy samples from 232 OSCC patients, and the relationship between Annexin A1 expression and cell proliferation as well as tumor formation using both in vitro and in vivo OSCC models. Results Annexin A1 expression correlated significantly with pathological differentiation grade in OSCC patients, a lower Annexin A1 expression correlating with a poorer differentiation grade. Forced Annexin A1 overexpression in OSCC cell lines, CAL27 and Tca8113, significantly reduced the cell proliferation whereas down-regulation of Annexin A1 expression in OSCC cell line, HB96, significantly increased proliferation of HB96 cells. Tumors formed from CAL27 cells overexpressing Annexin A1 grown significantly slower compared to the parental CAL27 cells in nude mice and showed a significantly reduced nuclear Ki-67 labeling index. Interestingly, these tumors also showed a well differentiated histology pattern whereas the tumors formed from the parental cells were consistently moderately differentiated. Conclusions These data support a significant correlation between Annexin A1 expression and pathological differentiation grade, and a functional role of Annexin A1 in inhibiting cell proliferation and cell differentiation in OSCC.

A cell-derived extracellular matrix (ECM) scaffold was constructed using cultured porcine chondrocytes via a freeze-drying method, and its ability to promote cartilage formation was evaluated in vitro. Scanning electron microscope (SEM) revealed that the scaffold had highly uniform porous microstructure. Then, rabbit chondrocytes were seeded dynamically on ECM scaffold and cultured for 2 days, 1, 2, and 4 weeks in vitro for analysis. Polyglycolic acid (PGA) scaffold was used as a control. On gross observation of neocartilage tissue, a silvery white cartilage-like tissue was observed after 1 week of culture in ECM scaffold, while similar morphology was seen only after 4 weeks in PGA scaffold. The volume of neocartilage-like tissue was significantly increased in both ECM and PGA groups. The compressive strength was gradually increased with time in ECM group, while gradually decreased in PGA group. DNA, glycosaminoglycan (GAG) and collagen contents also increased gradually with time in both groups, but showed more significant increase in ECM group. Histological staining for GAG (Safranin O staining) and type II collagen (immunohistochemistry) showed sustained accumulation of ECM molecules with time, which gradually and uniformly filled porous space in ECM scaffold. On the contrary, they accumulated only at the peripheral area of PGA scaffold. These results suggest that a novel cell-derived ECM scaffold can provide a promising environment for generating a high quality cartilage in vitro.

We investigated the synergistic effect of pidotimod and red ginseng acidic polysaccharide (RGAP) from Panax ginseng C.A. Meyer on humoral immune response challenged by lipopolysaccharide (LPS) and sheep red blood cells (SRBC) in immunosuppressed mice. Combined treatment with pidotimod and RGAP significantly increased the number of plaque-forming cells in the spleen in response to both LPS and SRBC, while treatment with either pidotimod or RGAP individually had no such effect. IgG levels in serum were augmented for secondary responses to SRBC in co-treated mice, but not in mice treated with either drug alone. Microscopic studies revealed that architecture of the spleen, thymus, and lymph nodes was conserved. GPT and creatinine in serum as indicators of hepatic and renal functions showed no difference compared to the control group. These results indicate that combined treatment with pidotimod and RGAP has an immunostimulatory effect in a synergistic manner on antibody response to challenge with LPS and SRBC without toxic changes.

We investigated the synergistic effect of combined treatment with red ginseng acidic polysaccharide (RGAP) from Panax ginseng C.A. Meyer and pidotimod in cyclophosphamide-treated mice. The combination of pidotimod and RGAP restored concanavalin A-induced splenic T cell proliferation and LPS-stimulated B cell proliferation significantly. The production of nitric oxide from peritoneal macrophages was increased by the combinations. NK cell activity was increased by RGAP alone or in combination with pidotimod. A synergistic increase in the level of serum IL-12 and interferongamm was observed when the combination of the two was used. RGAP alone or in combination with pidotimod modulated the level of serum C-reactive protein to a near-normal level. These results indicate that combinations of pidotimod and RGAP are synergistic and suggest that combination therapy using pidotimod and RGAP for improving immune activity may provide an additional benefit over the use of the two drugs by themselves.

We have observed in our previous study that a cell-derived extracellular matrix (ECM) scaffold could assure the growth of a cartilage tissue construct in vitro. The purpose of the present study was to evaluate the feasibility of a chondrocyte-seeded cell-derived ECM scaffold by implanting it in vivo in nude mouse. A porous cell-derived ECM scaffold was prepared with a freeze-drying protocol using porcine chondrocytes. Rabbit articular chondrocytes were seeded onto the scaffold and cultured for 2 days in vitro, and then implanted into the nude mouse subcutaneously. They were retrieved at 1, 2, and 3 weeks postimplantation. Under macroscopic analysis, the cartilage-like tissue formation matured with time and developed a smooth, white surface. Contrary to the control (in which no cells were seeded), the size of the neocartilage tissue increased slightly by the third week and remained more stable. Total glycosaminoglycan (GAG) content and the GAG/DNA ratio increased significantly with time in the chemical analysis. The histology exhibited a sustained accumulation of newly synthesized sulfated proteoglycans. Immunohistochemistry, Western blot, and reverse transcriptase-polymerase chain reaction (RT-PCR) clearly identified type II collagen at all time points. Compressive strength of in vivo neocartilage increased from 0.45 +/- 0.06 MPa at 1 week to 1.18 +/- 0.17 MPa at 3 weeks. In conclusion, this study demonstrated that the cell-derived ECM scaffold could provide chondrocytes with favorable in vivo environment to produce a hyaline-like cartilage tissue.

2-Bromopropane (2-BP), a halogenated propane analogue, is a substitute for chlorofluorocarbones. The present study was carried out to investigate the potential adverse effects of 2-BP on pregnant dams and embryo-fetal development after maternal exposure on gestational days (GD) 6 through 19 in Sprague-Dawley rats. The test chemical was administered subcutaneously to pregnant rats at dose levels of 0, 250, 500, and 1000 mg/kg per day. All dams were subjected to caesarean section on GD 20 and their fetuses were examined for external, visceral and skeletal abnormalities. In the 1000 mg/kg group, maternal toxicity included an increase in the incidence of abnormal clinical signs, a suppression in the body weight and body weight gain, and a decrease in the food intake. Developmental toxicity included an increase in the fetal deaths, a decrease in the litter size, and a reduction in the fetal body weight. In addition, an increase in the incidence of fetal external, visceral, and skeletal abnormalities was seen. In the 500 mg/kg group, minimal developmental toxicity including decreased fetal body weight and increased fetal ossification delay was observed. There were no adverse effects on either pregnant dams or embryo-fetal development in the 250 mg/kg group. These findings suggest that a 14-day subcutaneous dose of 2-BP is embryotoxic and teratogenic at a maternally toxic dose (i.e., 1000 mg/kg per day) and is minimally embryotoxic at a nonmaternally toxic dose (i.e., 500 mg/kg per day) in Sprague-Dawley rats. In the present experimental conditions, the no-observed-adverse-effect level (NOAEL) of 2-BP is considered to be 500 mg/kg per day for dams and 250 mg/kg per day for embryo-fetal development.

New N-thiadiazolylanilines were designed and synthesized to develop mitochondrial cytotoxins superior to SF 6847. The mitochondrial cytotoxin N-thiadiazolylanilines, TX-108 and TX-109, inhibited EMT6/KU mammary sarcoma cell growth at a low micromolar concentration. Their inhibitory activities were parallel to their mitochondrial cytotoxicity, such as uncoupling oxidative phosphorylation and inhibiting ATP synthesis. This report also supports the notion that the inhibition of tumor cell growth of inhibitor of protein tyrosine kinase AG17, which is identical to SF 6847, may be due to its mitochondrial cytotoxicity.

ObjectivesPaclitaxel coating of hemodialysis grafts is effective in suppressing neointimal hyperplasia in the graft and vascular anastomosis sites. However, paclitaxel can have unwanted effects on the surrounding tissues. To reduce such problems, we developed a method to coat the drug only on the luminal surface of the graft, with little loading on the outer surface.MethodsA peristaltic pump and a double-solvent (water and acetone) system were used to achieve an inner coating of paclitaxel. At the ratio of 90% acetone, paclitaxel was homogeneously coated only on the luminal surface of the graft without changing the physical properties. To determine its effect, grafts were implanted between the common carotid artery and the external jugular vein in pigs using uncoated control grafts (n = 6) and low-dose (n = 6, 0.22 μg/mm2) and high-dose (n = 6, 0.69 μg/mm2) paclitaxel inner-coated grafts. Cross-sections of graft–venous anastomoses were analyzed histomorphometrically 6 weeks after placement to measure the patency rate, percentage of luminal stenosis, and neointimal area.ResultsNo signs of infection or bacterial contamination were observed in the paclitaxel inner-coated groups. Only one of the six control grafts was patent, but all of the paclitaxel-coated grafts were patent, with little neointima. The mean ± standard error values of percentage luminal stenosis were 75.7% ± 12.7% (control), 17.5% ± 3.1% (low dose), and 19.7% ± 3.0% (high dose). The values for the neointimal area (in mm2) were 8.77 ± 1.66 (control), 3.53 ± 0.73 (lose dose), and 4.24 ± 0.99 (high dose). Compared with the control group, paclitaxel inner-coated vascular grafts significantly suppressed neointimal hyperplasia (low dose, P = .001; high dose, P = .002). Myofibroblast proliferation and migration into the graft interstices confirmed the firm attachment of the implanted graft to the surrounding tissue.ConclusionsPaclitaxel coating on the inner luminal surface of vascular grafts was effective in suppressing neointimal hyperplasia, with little inhibition of myofibroblast infiltration within the graft wall.Clinical RelevancePaclitaxel-coated vascular grafts effectively inhibited the neointimal hyperplasia of hemodialysis grafts. However, paclitaxel on the outer surface of expanded polytetrafluoroethylene grafts might prevent myofibroblast proliferation, which might cause hematomas, seromas, infections, or pseudoaneurysms in the space between the graft and surrounding tissue and result in incomplete hemodialysis needle insertion. Therefore, paclitaxel inner-coated expanded polytetrafluoroethylene grafts can reduce the coated amount of this potentially toxic drug to avoid such unwanted effects, whilst preserving its efficacy in inhibiting stenosis.

Background Numerous studies have demonstrated that titanium dioxide nanoparticles (TiO2 NPs) induced nephrotoxicity in animals. However, the nephrotoxic multiple molecular mechanisms are not clearly understood. Methods Mice were exposed to 2.5, 5 and 10 mg/kg TiO2 NPs by intragastric administration for 90 consecutive days, and their growth, element distribution, and oxidative stress in kidney as well as kidney gene expression profile were investigated using whole-genome microarray analysis technique. Results Our findings suggest that TiO2 NPs resulted in significant reduction of renal glomerulus number, apoptosis, infiltration of inflammatory cells, tissue necrosis or disorganization of renal tubules, coupled with decreased body weight, increased kidney indices, unbalance of element distribution, production of reactive oxygen species and peroxidation of lipid, protein and DNA in mouse kidney tissue. Furthermore, microarray analysis showed significant alterations in the expression of 1, 246 genes in the 10 mg/kg TiO2 NPs-exposed kidney. Of the genes altered, 1006 genes were associated with immune/inflammatory responses, apoptosis, biological processes, oxidative stress, ion transport, metabolic processes, the cell cycle, signal transduction, cell component, transcription, translation and cell differentiation, respectively. Specifically, the vital up-regulation of Bcl6, Cfi and Cfd caused immune/ inflammatory responses, the significant alterations of Axud1, Cyp4a12a, Cyp4a12b, Cyp4a14, and Cyp2d9 expression resulted in severe oxidative stress, and great suppression of Birc5, Crap2, and Tfrc expression led to renal cell apoptosis. Conclusions Axud1, Bcl6, Cf1, Cfd, Cyp4a12a, Cyp4a12b, Cyp2d9, Birc5, Crap2, and Tfrc may be potential biomarkers of kidney toxicity caused by TiO2 NPs exposure.