ROS-Responsive Bola-Lipid Nanoparticles as a Codelivery System for Gene/Photodynamic Combination Therapy.

The nonviral delivery systems that combine genes with photosensitizers for multimodal tumor gene/photodynamic therapy (PDT) have attracted much attention. In this study, a series of ROS-sensitive cationic bola-lipids were applied for the gene/photosensitizer codelivery. Zn-DPA was introduced as a cationic headgroup to enhance DNA binding, while the hydrophobic linking chains may facilitate the formation of lipid nanoparticles (LNP) and the encapsulation of photosensitizer Ce6. The length of the hydrophobic chain played an important role in the gene transfection process, and 14-TDZn containing the longest chains showed better DNA condensation, gene transfection, and cellular uptake. 14-TDZn LNPs could well load photosensitizer Ce6 to form 14-TDC without a loss of gene delivery efficiency. 14-TDC was used for codelivery of p53 and Ce6 to achieve enhanced therapeutic effects on the tumor cell proliferation inhibition and apoptosis. Results showed that the codelivery system was more effective in the inhibition of tumor cell proliferation than individual p53 or Ce6 monotherapy. Mechanism studies showed that the production of ROS after Ce6 irradiation could increase the accumulation of p53 protein in tumor cells, thereby promoting caspase-3 activation and inducing apoptosis, indicating some synergistic effect. These results demonstrated that 14-TDC may serve as a promising nanocarrier for gene/PDT combination therapy.

Metal-Organic Framework Photosensitized TiO2 Co-catalyst: A Facile Strategy to Achieve a High Efficiency Photocatalytic System.

A 3D metal-organic framework (ADA-Cd=[Cd2 L2 (DMF)2 ]⋅3 H2 O where H2 L is (2E,2'E)-3,3'-(anthracene-9,10-diyl)diacrylic acid) constructed from diacrylate substituted anthracene, sharing structural characteristics with some frequently employed anthraquinone-type dye sensitizers, was introduced as an effective sensitizer for anatase TiO2 to achieve enhanced visible light photocatalytic performance. A facile mechanical mixing procedure was adopted to prepare the co-catalyst denoted as ADA-Cd/TiO2 , which showed enhanced photodegradation ability, as well as sustainability, towards several dyes under visible light irradiation. Mechanistic studies revealed that ADA-Cd acted as the antenna to harvest visible light energy, generating excited electrons, which were injected to the conduction band (CB) of TiO2 , facilitating the separation efficiency of charge carriers. As suggested by the results of control experiments, combined with the corresponding redox potential of possible oxidative species, . O2- , generated from the oxygen of ambient air at the CB of TiO2 was believed to play a dominant role over . OH and h+ . UV/Vis and photoluminescence technologies were adopted to monitor the generation of . O2- and . OH, respectively. This work presents a facile strategy to achieve a visible light photocatalyst with enhanced catalytic activity and sustainability; the simplicity, efficiency, and stability of this strategy may provide a promising way to achieve environmental remediation.

Bio-reducible polycations from ring-opening polymerization as potential gene delivery vehicles.

Synthetic polycations show great potential for the construction of ideal non-viral gene delivery systems. Several cationic polymers were synthesized by the epoxide ring-opening polymerization between diepoxide and various polyamines. Disulfide bonds were introduced to afford the polymers bio-reducibility, while the oxygen-rich structure might enhance the serum tolerance and biocompatibility. The polycations have much lower molecular weights than PEI 25 kDa, but still could well bind and condense DNA into nano-sized particles. DNA could be released from the polyplexes by addition of reductive DTT. Compared to PEI, the polycations have less cytotoxicity possibly due to their lower molecular weights and oxygen-rich structure. More significantly, these materials exhibit excellent serum tolerance than PEI, and up to 6 times higher transfection efficiency than PEI could be obtained in the presence of serum. The transfection mediated by was seldom affected even at a high concentration of serum. Much lower protein adsorption of polycations than PEI was proved by bovine serum albumin adsorption experiments. Flow cytometry also demonstrates their good serum resistance ability.

TACN-based cationic lipids with amino acid backbone and double tails: materials for non-viral gene delivery.

Cationic lipids have become an efficient type of non-viral vectors for gene delivery. In this Letter, four cationic lipids containing 1,4,7-triazacyclononane (TACN) headgroup, glutamic/aspartic acid backbone and dioleyl tails were designed and synthesized. The TACN headgroup gives these lipids excellent pH buffering capacities, which were higher than branched 25 kDa PEI. Cationic liposomes prepared from these lipids and DOPE showed good DNA affinity, and full DNA condensation was found at N/P ratio of 3 via agarose gel electrophoresis. The lipoplexes were characterized by dynamic light scattering (DLS) assay, which gave proper particle sizes and zeta-potentials for transfection. In vitro gene transfection results in two cell lines reveal that TAN (with aspartic acid and amide bond in the structure) shows the best transfection efficiency, which is close to commercially available transfection agent Lipofectamine 2000.

Effect of distilled water on rapid inactivation of tumour cells attached to surgery instruments.

The aim of the study was to explore the effect of distilled water on killing tumour cells attached to the surgery instruments during operation. Tumour cells were collected from the suspected tumour cell-contaminated surgery instruments and then cultured. Then the tumour cells were treated by distilled water at different gradient temperature for different time periods. The morphology of the tumour cells was observed by inverted microscope after hematoxylin-eosin staining. The results showed that positive tumour cell culture rate was 34.3%. After soaked in distilled water for 60 s at 55°C, the tumour cells were inactive, and the death rate was 100%. We also found that no active cells were seen to grow adherently after recultured. In conclusion, tumour cells can be killed by distilled water for 60 s at 55°C, which provides a new fast and low-cost tumour-free technique to inactivate tumour cells attached to surgery instruments.

TACN-containing cationic lipids with ester bond: preparation and application in gene delivery.

A series of novel cationic lipids based on 1,4,7-triazacyclononane (TACN) with different hydrophobic chains were synthesized via the formation of a biodegradable ester bond. These lipids were found to have good buffering capacity at the pH range of 5.0-6.5, which is similar to that of the acidic endosomal compartments. The liposomes formed from these lipids and DOPE could condense DNA into nanoparticles with proper sizes. In vitro experiments showed moderate to good gene transfection efficiency of the formed lipoplexes. The structure-activity relationships of this type of lipids were discussed.

Diol glycidyl ether-bridged cyclens: preparation and their applications in gene delivery.

Polymeric 1,4,7,10-tetraazacyclododecanes (cyclens) using diol glycidyl ether with different chain length as bridges (5a-e) were designed and synthesized from various diols, 1,7-diprotected cyclen and epichlorohydrin. The molecular weights of the title polymers were measured by GPC with good polydispersity. Agarose gel retardation and fluorescent titration using ethidium bromide showed good DNA-binding ability of 5. They could retard plasmid DNA (pDNA) at an N/P ratio of 4-6 and form polyplexes with sizes around 100-250 nm from an N/P ratio of 10 to 60 and relatively low zeta-potential values (5-22 mV). The cytotoxicity of 5 assayed by MTT is much lower than that of 20 kDa PEI. In vitro transfection against A549 and 293 cells showed that the transfection efficiency (TE) of 5c/DNA polyplexes is close to that of 20 kDa PEI at an N/P ratio of 5. Structure-activity relationship (SAR) of 5 was discussed in their DNA-binding, cytotoxicity, and transfection studies. The TE of 5c/DNA polyplexes could be improved by the introduction of 50 µM of chloroquine, the endosomolytic agents, to pretreated cells. These studies may extend the application areas of macrocyclic polyamines, especially for cyclen.

Novel CO2 Fluorescence Turn-On Quantification Based on a Dynamic AIE-Active Metal-Organic Framework.

Traditional CO2 sensing technologies suffer from the disadvantages of being bulky and cross-sensitive to interferences such as CO and H2O, these issues could be properly tackled by innovating a novel fluorescence-based sensing technology. Metal-organic frameworks (MOFs), which have been widely explored as versatile fluorescence sensors, are still at a standstill for aggregation-induced emission (AIE), and no example of MOFs showing a dynamic AIE activity has been reported yet. Herein, we report a novel MOF, which successfully converts the aggregation-caused quenching of the autologous ligand molecule to be AIE-active upon framework construction and exhibits bright fluorescence in a highly viscous environment, resulting in the first example of MOFs exhibiting a real dynamic AIE activity. Furthermore, a linear CO2 fluorescence quantification for mixed gases in the concentration range of 2.5-100% was thus well-established. These results herald the understanding and advent of a new generation in all solid-state fluorescence fields.

Implication of CD38 gene in autophagic degradation of collagen I in mouse coronary arterial myocytes.

Collagen deposition is a hallmark of atherosclerosis. Although compromised collagen I degradation has been implied in the pathogenesis of atherosclerosis, the molecular mechanisms are still unclear. Thus, we determined the role of CD38, an enzyme involved in cellular calcium modulation and autophagic flux, in the regulation of collagen I degradation in coronary arterial myocytes (CAMs).In primary cultured CAMs from CD38-/- mice, collagen I protein accumulation but not mRNA abundance was significantly increased compared with cells from CD38+/+ mice either under control or upon TGF-Beta stimulation. Pharmacological inhibition of the formation of autophagosomes with 3-methyladenine or of autophagolysosomes with a lysosomal functional blocker, bafilomycin A1, induced a similar increase in collagen protein levels, while inhibition of the proteasome by MG132 had no effects on collagen I accumulation. In addition, CD38-deficiency did not change the protein expression of matrix metalloprotein-9 (MMP-9) or tissue inhibitor of metalloproteinase-1 (TIMP-1) in CAMs. Confocal microscopy showed that collagen I deposition was mainly lied within lysosomes or autophagosomes in CD38-/- or TGF-Beta treated CAMs. Collagen I deposition increased when CAMs lack CD38 expression or if autophagy was blocked, which is associated with impaired autophagic degradation of collagen I. This CD38 regulation of autophagic flux may represent a novel mechanism for extracellular matrix (ECM) plasticity of coronary arteries upon atherogenic stimulation.

Analysis of exfoliated gastric carcinoma cells attached on surgical supplies.

Surgery is considered to have a leading role in the treatment of gastric carcinoma. Surgical supplies are used to cut, divide, and ligate during surgery, and are not only in close contact with normal tissues, but may also be contaminated by pathological tissues and cells. This study sought to determine the presence of exfoliated tumor cells on surgical supplies at different stages during the surgical procedure. We collected five types of surgical supplies from 90 patients who underwent D2 radical gastrectomy to find out if there was any cancer cells attached to them. Highest numbers of cancer cells were found on gauze used to clean the surgical instruments and on the gloves of scrub nurses. The likelihood of finding cancer cells increased with advancing clinical stage of disease, lower differentiation of cancer cells, increasing frequency of use of supplies and extent of contact, and was also associated with the characteristic of surgical supplies. Dissemination of tumor cells could be prevented by using a number of methods, depending on the type of surgical supply items.

Cyclen-based lipidic oligomers as potential gene delivery vehicles.

A series of cyclen-based linear oligomers bearing hydrophobic long chains (lipopolymers Cy-LC, where Cy and LC represent cyclen-based linear backbone and hydrophobic long chain substituents, respectively) were designed and synthesized. The effects of type and degree of substitution (DS) of hydrophobic long chains on the transfection efficiency were systematically studied. The nitrogen atoms with relatively strong basicity on the cyclen ensure their good DNA binding ability, which was confirmed by gel retardation and ethidium bromide exclusion assays. Lipopolyplexes could be formed as nanoparticles with suitable sizes and zeta potentials for gene transfection. In vitro gene delivery experiments revealed that the linoleic acid (LIN) substituted material Cy-LIN has better transfection efficiency than 25 kDa polyethylenimine in the absence or in the presence of serum. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and hemolysis assays showed low cytotoxicity and good biocompatibility of the lipopolyplexes. Fluorescent labeled DNA was used to study the cellular uptake and intracellular distribution of transfected DNA. Flow cytometry results suggested that a long chain is necessary for efficient cellular uptake, and images from confocal laser scanning microscopy showed that after 4h transfection, most of the fluorescent labeled DNA accumulated in the perinuclear region, which was required for efficient gene expression. Moreover, it was also found that the DS of the hydrophobic moiety can adjust the balance between DNA binding ability and dissociation of polyplexes, significantly affecting the transfection efficiency.

Ring-opening polymerization for hyperbranched polycationic gene delivery vectors with excellent serum tolerance.

In order to improve the transfection efficiency (TE) and biocompatibility, we synthesized a series of hyperbranched cationic polymers by ring-opening polymerization between diepoxide and several polyamines. These materials can condense plasmid DNA efficiently into nanoparticles that have much lower cytotoxicity than those derived from bPEI. In vitro transfection experiments showed that polymers prepared from branched or cyclic polyamine (P1 and P5) exhibited TE several times higher than 25KDa bPEI. More significantly, serum seemed to have no negative effect on P1-P5 mediated transfection. On the contrary, the TE of P1 improved, even when the serum concentration reached 70%. Several assays demonstrated the excellent serum tolerance of such polycationic vectors: bovine serum albumin (BSA) adsorption assay revealed considerably lower protein adsorption of P1-P5 than PEI; P1 showed better DNA protection ability from degradation by DNase I than PEI; flow cytometry results suggested that any concentration of serum may not decrease the cellular uptake of P1/DNA polyplex; and confocal laser scanning microscopy also found that serum has little effect on the transfection. By using specific cellular uptake inhibitors, we found that the polyplexes enter the cells mainly via caveolae and microtubule-mediated pathways. We believe that this ring-opening polymerization may be an effective synthetic approach toward gene delivery materials with high biological activity.

Low molecular weight PEI-appended polyesters as non-viral gene delivery vectors.

Routine clinical implementation of human gene therapy requires safe and efficient gene delivery methods. Linear biodegradable polyesters with carbon-carbon double bonds are prepared from unsaturated diacids and diols. Subsequent appending of low molecular weight PEI by Michael addition gives target cationic polymers efficiently. Agarose gel retardation and fluorescence quenching assays show that these materials have good DNA binding ability and can completely retard plasmid DNA at weight ratio of 0.8. The formed polyplexes have appropriate sizes around 275 nm and zeta-potential values about +20-35 mV. The cytotoxicities of these polymers assayed by MTT are much lower than that of 25 kDa PEI. In vitro transfection toward 7402, HEK293 and U-2OS cells show that polymer P1 may give dramatically higher transfection efficiency (TE) than 25 kDa PEI, especially in U-2OS cells, suggesting that such polymer might be promising non-viral gene vectors.

[Contamination of surgical materials by exfoliated cancer cells during gastric carcinoma operation].

OBJECTIVE: To explore the exfoliated cancer cell contamination in different surgical materials during the malignant gastrectomy. METHODS: Ninety gastric cancer patients undergoing gastrectomy were prospectively enrolled in this study. The operation materials of these 90 gastrectomy were divided into 5 groups: surgical instruments (A), gloves for surgeons (B), gloves and gauzes of scrub nurse (C), gauzes for hemostasis (D), anastomosis instrument (E). The rinse fluid of materials was cultured to verify positive cancer cells. Associations among different pathological stages, differentiations, materials and positive cancer cells rates were examined. RESULTS: Stage II and III patients had higher positive rates of exfoliated cancer cell contamination than stage I patients [26.5 (9/34) and 47.5% (21/46) vs. 10.0% (1/10),P=0.046]. Low differentiated adenocarcinoma group had higher positive rate than moderately and well differentiated adenocarcinoma groups [44.8% (26/58) vs. 16.7% (4/24) and 12.5% (1/8), P=0.020]. Positive cancer cell rates of 5 kinds of materials were as follows: 12.2% (11/90) in A group, 6.7% (6/90) in B group, 22.2% (20/90) in C group, 15.6% (14/90) in D group and 3.3% (3/90) in E group, and the differences were significant (P<0.01). CONCLUSION: Different operation materials have different risks to be contaminated by cancer cells, which is associated with the contact frequency, cancer staging and pathological classification.

Linear polycations by ring-opening polymerization as non-viral gene delivery vectors.

For a clinically effective non-viral gene delivery system, a non-toxic and highly efficient vector is of great importance. A series of linear cationic polymers were synthesized by the ring-opening polymerization between diglycidyl ethers and diamines. Their structure-activity relationships as gene delivery vectors were systematically studied. Besides the amino groups with various densities, these polymers have uniform distribution of hydroxyl groups, which were formed in the polymerization and may benefit their biocompatibility and serum-tolerance. These polymers have good DNA binding ability and could condense DNA into nanoparticles with proper sizes and zeta-potentials. MTT assay revealed that polyplexes formed from title polymers have lower cytotoxicity than that derived from PEI. Most of the polymers have higher transfection efficiency than 25 kDa PEI in the in vitro transfection experiments. Polymers prepared from diglycidyl ethers with less or no N atom (2a and 2b) gave dramatically decreased TE, indicating that secondary amine on the backbone is highly required for efficient gene transfection, and compound 2 may be a good building block in the design of cationic polymers for gene delivery. More importantly, these polymers showed much better serum tolerance. Unlike PEI, the transfection mediated by P5 was seldom affected by the presence of 10% serum. Cellular uptake and intracellular distribution studies also confirmed the good performance of P5 in the transfection process with serum.