Manganese-mineralized cancer cells as immunogenic cancer vaccines for tumor immunotherapy.

The strategy of using tumor cells to construct whole-cell cancer vaccines has received widespread attention. However, the limited immunogenicity of inactivated tumor cells and the challenge of overcoming immune suppression in solid tumors have hindered the application of whole-cell-based cancer immune therapy. Inspired by the regulatory effects of MnO2 and spatiotemporal control capability of material layers in cell surface engineering, we developed a manganese (Mn)-mineralized tumor cell, B16F10@MnO2, by inactivating B16F10 melanoma cells with KMnO4 to generate manganese-mineralized tumor cells. The cell-based composite was formed by combining amorphous MnO2 with the membrane structure of cells based on the redox reaction between KMnO4 and tumor cells. The MnO2 layer induced a stronger phagocytosis of ovalbumin (OVA)-expressing tumor cells by antigen presenting cells than formaldehyde-fixed cells did, resulting in specific antigen-presentation in vitro and in vivo and subsequent immune responses. Intratumoral therapy with B16F10@MnO2 inhibited B16F10 tumor growth. Moreover, the infiltration of CD8+ T cells within B16F10 solid tumors and the proportion of central memory T cells both increased in B16F10@MnO2 treated tumor-bearing mice, indicating enhanced adaptive immunity. This study provides a convenient and effective method to improve whole-cell-based anti-tumor therapy.

Association of the rs1990760, rs3747517, and rs10930046 polymorphisms in the IFIH1 gene with susceptibility to autoimmune diseases: a meta-analysis.

Objective: Interferon induced with helicase C domain 1 (IFIH1) single-nucleotide polymorphisms (SNP) rs1990760, rs3747517, and rs10930046 have been shown to be closely related to the risk of autoimmune diseases. The aim of this study was firstly to examine the association of the rs1990760 with type 1 diabetes (T1D) in a Chinese population. Secondly, to assess the association of SNP rs1990760, rs3747517, and rs10930046 with autoimmune diseases susceptibility. Methods: A total of 1,273 T1D patients and 1,010 healthy control subjects in a Chinese population were enrolled in this case-control study. Subsequently, we performed a meta-analysis on the association of the SNP rs1990760, rs3747517, and rs10930046 in the IFIH1 gene with susceptibility to autoimmune diseases. The random and fixed genetic effects models were used to evaluate the association and the effect sizes, including odds ratios (OR) and 95% confidence intervals (CI). Stratification analyses based on ethnicity and the type of autoimmune diseases were performed. Results: IFIH1 SNP rs1990760 was not associated with a significant risk of T1D in the Chinese population in the case-control study. A total of 35 studies including 70,966 patients and 124,509 controls were identified and included in the meta-analysis. The results displayed significant associations between IFIH1 rs1990760 A allele and rs3747517 C allele and autoimmune diseases risk (OR=1.09, 95% CI: 1.01~1.17; OR=1.24, 95% CI: 1.15~1.25, respectively). Stratified analysis indicated a significant association rs1990760 and rs3747517 with autoimmune diseases risk in the Caucasian population (OR=1.11, 95% CI: 1.02~1.20, OR=1.29, 95% CI: 1.18~1.41, respectively). Conclusions: This study revealed no association between IFIH1 SNP rs1990760 and T1D in Chinese. Furthermore, the meta-analysis indicated that rs1990760 and rs3747517 polymorphisms, confer susceptibility to autoimmune diseases, especially in the Caucasian population.

3D Technique-Based Nonsurgical Correction of Deformational Congenital Auricular Deformities.

INTRODUCTION: Congenital auricular deformity (CAD) is a common postpartum deformity, and nonsurgical correction of CAD has been recognized as a safe and effective approach. Three-dimensional (3D) technique has been used in surgical reconstruction of unilateral microtia; however, 3D technique used in nonsurgical correction for deformational CAD has not been reported. METHODS: In this study, 12 CAD patients aged from 0.6 to 7 months with 16 deformational CAD were treated with 3D technique-based personalized nonsurgical correction (3D-NSC). Patients' CAD was photographed pre- and post-correction, and clinical outcome was evaluated as poor, fair, good, and excellent by comparing pre- and post-correction pictures. Different kinds of tests were used to analyze the data. RESULTS: All patients got an improved auricle shape (10 excellent, 2 good, and 4 fair). Multivariate regression analysis showed that CAD type was significantly associated with correction outcome, sex and age were significantly associated with correction outcome for the 11 constructed types of CAD, and age was significantly associated with the correction outcome when we focused on the male constructed auricles. CONCLUSION: 3D-NSC provided a significant nonsurgical clinical treatment for CAD patients, with younger patients getting better clinical outcomes with shorter correction time.

Development of a Survival Prognostic Model for Non-small Cell Lung Cancer.

Lung cancer is a leading cause of cancer-related death, and >80% of lung cancer diagnoses are non-small-cell lung cancer (NSCLC). However, when using current staging and prognostic indices, the prognosis can vary significantly. In the present study, we calculated a prognostic index for predicting overall survival (OS) in NSCLC patients. The data of 545 NSCLC patients were retrospectively reviewed. Univariate and multivariate Cox proportional hazards regression analyses were performed to evaluate the prognostic value of clinicopathological factors. Age (hazard ratio [HR] = 1.25, 95% confidence interval [CI] = 1.02-1.54), TNM stage (III, HR = 1.64, 95% CI = 1.08-2.48; IV, HR = 2.33, 95% CI = 1.48-3.69), lung lobectomy (HR = 1.96, 95% CI = 1.45-2.66), chemotherapy (HR = 1.42, 95% CI = 1.15-1.74), and pretreatment hemoglobin level (HR = 1.61, 95% CI = 1.28-2.02) were independent prognosticators. A prognostic index for NSCLC (PInscl, 0-6 points) was calculated based on age (≥65 years, 1 point), tumor-node-metastasis (TNM) stage (III, 1 point; IV, 2 points), lung lobectomy (no, 1 point), chemotherapy (no, 1 point), and pretreatment hemoglobin level (low, 1 point). In comparison with the "PInscl = 0" subgroup (survival time = 2.71 ± 1.86 years), the "PInscl = 2" subgroup (survival time = 1.86 ± 1.24 years), "PInscl = 3" subgroup (survival time = 1.45 ± 1.07 years), "PInscl = 4" subgroup (survival time = 1.17 ± 1.06 years), "PInscl = 5" subgroup (survival time = 0.81 ± 0.78 years), and "PInscl = 6" subgroup (survival time = 0.65 ± 0.56 years) exhibited significantly shorter survival times. Kaplan-Meier survival analysis showed that patients with higher PInscl scores had poorer OS than those with lower scores (log-rank test: χ2 = 155.82, P < 0.0001). The area under the curve of PInscl for predicting the 1-year OS was 0.73 (95 % CI = 0.69-0.77, P < 0.001), and the PInscl had a better diagnostic performance than the Karnofsky performance status or TNM stage (P < 0.01). In conclusion, the PInscl, which is calculated from age, TNM stage, lung lobectomy, chemotherapy, and pretreatment hemoglobin level, significantly predicted OS in NSCLC patients.

Successful Treatment of Chronic Lower Extremity Ulcers with Allogeneic Platelet-Rich Plasma and Artificial Dermis: A Case Report.

Hydroxyurea is an oral medication associated with painful, nonhealing ulcers, for which there is no effective treatment but permanent discontinuation of hydroxyurea. The authors present a case of leg ulcers that likely occurred as a result of hydroxyurea use in a patient with essential thrombocythemia. Topical treatment with allogeneic platelet-rich plasma and artificial dermis completely healed the leg ulcers without hydroxyurea cessation.

Pretreatment Hemoglobin Level Is an Independent Prognostic Factor in Patients with Lung Adenocarcinoma.

Background/Aim: Few studies have reported the prognostic value of pretreatment hemoglobin levels in patients with lung adenocarcinoma (LA). In the present study, we retrospectively reviewed 306 LA patients for their prognosis associated with the pretreatment hemoglobin levels. Methods: Person-years and case fatality rate (CFR) were calculated from May 2010 to June 2017. Hazard ratio (HR) and 95% confidence intervals (CIs) were estimated using the Cox proportional hazards regression analysis. Survival curves were generated using the Kaplan-Meier analysis. Results: Patients with low pretreatment hemoglobin (LPHb) levels had a higher CFR than did patients with normal pretreatment hemoglobin (NPHb) levels (HR = 1.48, 95% CI = 1.06-2.08, and P=0.023). Overall survival of NPHb patients was significantly higher than that of LPHb patients (P < 0.05). Conclusion: Low pretreatment hemoglobin level was demonstrated to be an independent biomarker for poor prognosis in patients with LA.

Pre-treatment hemoglobin levels are an independent prognostic factor in patients with non-small cell lung cancer.

To date, few studies have reported the prognostic value of pre-treatment hemoglobin levels in patients with non-small cell lung cancer (NSCLC). In the present study, 416 patients with NSCLC were retrospectively reviewed. Univariate Cox proportional hazards regression analysis demonstrated that patients with normal pre-treatment hemoglobin (NPHb) levels had a greater chance of surviving for longer period, than did patients with low pre-treatment hemoglobin (LPHb) levels (HR, 2.05; 95% CI, 1.63-2.57; P<0.001). After adjustment for age, sex, tumor-node-metastasis stage, Karnofsky performance status, lung lobectomy, chemotherapy and radiotherapy, multivariate Cox proportional hazards regression analysis revealed that LPHb was an independent predictor for the poor prognosis of patients with NSCLC (HR, 1.86; 95% CI, 1.47-2.36; P<0.001). Estimation of the cumulative survival revealed that the overall survival of NPHb patients was significantly higher than that for LBHb patients (P<0.05), independent of whether the patients had received lung lobectomy or chemotherapy treatments. In conclusion, low pre-treatment hemoglobin levels were demonstrated to be an independent biomarker for poor prognosis in patients with NSCLC.

Impact of molecular weight in four-branched star vectors with narrow molecular weight distribution on gene delivery efficiency.

A series of star-shaped cationic polymers, termed star vectors (SVs), has been developed as effective nonviral gene delivery carriers. In this study, we separated SVs into several fractions having different molecular weights with very narrow molecular weight distributions in order to examine in detail the influence of the molecular weight of the SVs on the gene transfection efficiency. As a model compound for several types of SVs, 4-branched poly(N,N-dimethylaminopropyl acrylamide) having a molecular weight (M(n)) of approximately 35 kDa and polydispersity of 1.6 was prepared by iniferter-based radical polymerization. The SVs were separated using size-exclusion chromatography to obtain seven fractions having M(n) ranging from 27 kDa to 73 kDa with polydispersity ranging from 1.1 to 1.2. All the fractionated SVs have similar pH of 10.2-10.4 and were able to interact with and condense luciferase-encoding plasmid deoxyribonucleic acid (DNA) to yield SV/DNA polyplexes. A water-soluble tetrazolium-1 (WST) assay showed that all SVs had minimal cellular cytotoxicity under an N/P charge ratio of 10. The critical micellar concentration decreased with an increase in the M(n) of the fractionated SVs; however, the particle size of the polyplexes, exclusion activity of ethidium bromide, and zeta-potential of the polyplexes increased. An in vitro evaluation using COS-1 cells at an N/P ratio of 10 showed that transfection activity increased almost linearly with M(n). The highest transfection activity was obtained for SVs with the highest M(n) (73 kDa), which was over 7 times that for the SVs with the lowest M(n) (27 kDa), the nonfractionated original SV, or PEI standard. The transfection efficiency was more correlated with the amphiphilicity or hydrophobicity of the SVs and the surface potential and condensate density of the polyplexes than with the particle size.

Photoinduced cross-linking of star vector for improvement of gene transfer efficiency.

This study aimed to investigate the effect of cross-linking of a cationic nonviral gene carrier on gene expression. As a precursor for photo-cross-linking, a star-shaped, six-branched cationic polymer of poly(N,N-dimethylaminopropylacrylamide) (six-branched star vector, SV), which was previously designed as a gene carrier, was synthesized by iniferter-based living radical polymerization. Upon UV irradiation, the number-average molecular weight (Mn) of the SV increased from ca. 28 kDa to ca. 32 kDa (irradiation time, 180 min) and ca. 46 kDa (240 min) with broadness of the polydispersity due to the coupling reaction between the polymer radicals generated at the terminal ends of each branch of the SVs, resulting in the preparation of cross-linked SVs (CSVs) without the use of any chemical cross-linking agents. Irrespective of cross-linking, all the SVs were able to interact with and condense luciferase-encoding plasmid DNA to yield relatively stable polymer/DNA complexes (polyplexes) of approximate diameter 150 nm with zeta-potential of ca. 20 mV. However, a transfection study using several types of cell lines, HeLa, Hep G2, 293, and COS-1, showed that by cross-linking of SVs the luciferase activity increased drastically. The activity with CSV (Mn=ca. 46 kDa) was increased by at least 1 order of magnitude in the original SV (Mn=ca. 28 kDa), which was several-fold that in the SV with the same molecular weight in all cells. In all SVs, no significant cellular cytotoxicity was observed even at a high charge ratio of 45. The SV-based gene transfection was significantly enhanced by the cross-linking of the SVs.

Enhancement of star vector-based gene delivery to endothelial cells by addition of RGD-peptide.

This study aimed to investigate the feasibility of using a cationic nonviral gene carrier in endothelial cells for enhancing gene expression by the addition of an integrin-binding RGD peptide. A 4-branched cationic polymer of poly( N,N-dimethylaminopropylacrylamide) (star vector), developed as a gene carrier, could complex with the luciferase-encoding plasmid DNA under a charge ratio of 5 (vector/pDNA) to form polymer/DNA complexes (polyplexes). The addition of the RGD-containing peptide (GRGDNP) to the polyplex solution led to a decrease in the zeta-potential from ca. +30 to +20 mV along with the reduction in the particle size from ca. 300 to 200 nm. Additionally, a marked inhibition of polyplex aggregation was observed, indicating the coating of the polyplex surface with RGD peptides. A transfection study on endothelial cells showed that the luciferase activity increased with the amount of RGD peptides added to the polyplexes and exhibited minimal cellular cytotoxicity. The transfection activity further increased when cyclic RGD peptides (RGDFV) were used; the activity with RGD peptide addition was approximately 8-fold compared to that without RGD peptide addition. Gene delivery to endothelial cells was significantly enhanced by only the addition of RGD peptides to star vector-based polyplexes.

Synthesis and in vitro evaluation of novel star-shaped block copolymers (blocked star vectors) for efficient gene delivery.

Novel 4-branched diblock copolymers consisting of cationic chains as an inner domain and nonionic chains as an outer domain were prepared by iniferter-based living radial polymerization and evaluated as a polymeric transfectant. The cationic polymerization of 3-(N,N-dimethylamino)propyl acrylamide (DMAPAAm) using 1,2,4,5-tetrakis( N,N-diethyldithiocarbamylmethyl)benzene as a 4-functional iniferter followed by the nonionic block polymerization of N,N-dimethylacrylamide (DMAAm) afforded 4-branched diblock copolymers with controlled compositions. By changing the solution or irradiation conditions, 4-branched PDMAPAAms with molecular weights of 10,000, 20,000, and 50,000 were synthesized. In addition, by graft polymerization, PDMAPAAm-PDMAAm blocked copolymers with copolymer composition (unit ratio of DMAAm/DMAPAAm) ranging from 0.18 to 1.0 for each cationic polymer were synthesized. All polymers were shown to interact with and condense plasmid DNA to yield polymer/DNA complexes (polyplexes). A transfection study on COS-1 cells showed that the polyplexes from block copolymers with cationic chain length of approximately 50,000 and a nonionic chain length of 30,000, which were approximately 200 nm in diameter and very stable in aqueous media, had the most efficient luciferase activity with minimal cellular cytotoxicity under a charge ratio of 20 (vector/pDNA). The PDMAPAAm-PDMAAm-blocked, star-shaped polymers are an attractive novel class of nonviral gene delivery systems.

Deposition transfection technology using a DNA complex with a thermoresponsive cationic star polymer.

A novel non-viral gene transfection method in which DNA complexes were kept in contact with a deposition surface (deposition transfection) was developed. We designed a novel aqueous thermoresponsive adsorbent material for DNA deposition, which was a star-shaped copolymer with 4-branched chains. Each chain is comprised of a cationic poly(N,N-dimethylaminopropyl acrylamide) (PDMAPAAm) block (Mn: ca. 3000 g x mol(-1)), which formed an inner domain for DNA binding and a thermoresponsive poly(N-isopropylacrylamide) (PNIPAM) block (Mn: ca. 6000 g x mol(-1)), which formed an outer domain for surface adsorption. Complex formation between the copolymer and the luciferase-encoding plasmid DNA occurred immediately upon simple mixing in an aqueous medium; polyplexes approximately 100 nm in size were formed. Because the lower critical solution temperature of the polyplexes was approximately 35 degrees C, they could deposit on the substrate by precipitation from an aqueous solution upon warming, which was confirmed by quartz crystal microbalance (QCM) method and water contact angle measurement. When COS-1 cells were cultured on the polyplex-deposited substrate in a culture medium, the luciferase activity observed was higher than that observed on a DNA-coated substrate with or without the cationic polymer before and after complete adhesion and by conventional solution transfection using the polyplexes. The activity was enhanced with an increase in the charge ratio (surfactant/pDNA) with permissible cellular cytotoxicity.

Serine protease inhibitors inhibit superoxide release and adherence in human neutrophils stimulated by granulocyte-macrophage colony-stimulating factor and tumor necrosis factor-alpha.

Stimulation of human neutrophils with granulocyte-macrophage colony-stimulating factor (GM-CSF) or tumor necrosis factor-alpha (TNF) results in increased superoxide (O2-) release and adherence. O2- release and adherence are dependent on activation of extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein kinase (MAPK). Possible participation of serine proteases in GM-CSF- or TNF-induced activation of human neutrophils was explored with various serine protease inhibitors, including phenylmethylsulfonyl fluoride, L-1-tosylamido-2-phenylethyl-chloromethyl ketone and N-alpha-p-tosyl-L-lysine-chloromethyl ketone. GM-CSF- or TNF-induced O2- release and adherence were inhibited in parallel by pretreatment of neutrophils with these inhibitors. On the other hand, GM-CSF- or TNF-induced phosphorylation of ERK and p38 MAPK was unaffected by these inhibitors at the concentrations effective for the inhibition of O2- release and adherence. These findings suggest that serine proteases are involved in GM-CSF- and TNF-induced O2- release and adherence in human neutrophils and that serine proteases function downstream or independently of the activation of ERK and p38 MAPK.