In vivo immunological activity of chitosan-derived nanoparticles.

Chitosan has been studied as an immunomodulator, but few studies have used chitosan derivatives as adjuvants alone. After a preliminary study, we found that nanoparticles prepared from chitosan derivatives had better cellular immune activity when used as an adjuvant. Therefore, animal experiments were conducted to further investigate the performance and mechanism of these nanoparticles as immune adjuvants. We injected mice with the chitosan nanoparticle vaccine and measured the expression levels of immunoglobulins, immune factors, and immune genes in tissues and tissue sections. The results showed that C236-HACC-OVA (C2,3,6-chitosan sulfate-chitosan quaternary ammonium salt-ovalbumin) and NO-HACC-OVA (NO-carboxymethyl chitosan-chitosan quaternary ammonium salt-ovalbumin) nanoparticles can significantly improve the secretion of the immune factors IL-6, TNF, and IL-1ß. The level of IgG1 was highly significant after administering both nanoparticles, but IgG2 was not significant in mice. Three immune factors (IL-4, IL-6, and IL-17) were secreted at high levels in mouse serum at a nanoparticle dose of 0.3 mg/mouse. These nanoparticles also have high safety in the liver, kidney, and spleen of mice. This study proves the possibility of using chitosan derivative nanoparticles as vaccine adjuvants. These data further indicate that chitosan derivative nanoparticles have potential for use as vaccine adjuvants and demonstrate that polysaccharides have a unique position in green vaccine research.

Evaluation of dosimetric predictors of toxicity after IMRT with concurrent chemotherapy for anal cancer.

BACKGROUND: This study investigates the impact of dosimetric parameters on acute and late toxicity for patients with anal squamous cell carcinoma (SCC) treated with image-guided intensity modulated radiation therapy (IG-IMRT) and concurrent chemotherapy. MATERIALS AND METHODS: Patients were enrolled in an observational cohort study between 2008 and 2013 (median follow-up 3.4 years). They were treated with standardized target and organ-at-risk (OAR) contouring, planning, and IG-IMRT. Radiotherapy dose, based on clinicopathologic features, ranged from 45 Gy to 63 Gy to gross targets and 27 Gy to 36 Gy to elective targets. Chemotherapy was concurrent 5-fluorouracil and mitomycin C (weeks 1&5). Toxicity was prospectively graded using NCI CTCAE v.3 and RTOG scales. Logistic regression was used to assess the association between dose/volume parameters (e.g small bowel V5) and corresponding grade 2 + and 3+ (G2+/3 + ) toxicities (e.g. diarrhea). RESULTS: In total, 87 and 79 patients were included in the acute and late toxicity analyses, respectively. The most common acute G2 + toxicities were skin (dermatitis in 87 % [inguino-genital skin], 91 % [perianal skin]) and hematologic in 58 %. G2 + late anal toxicity (sphincter dysfunction), gastrointestinal toxicity, and skin toxicity were respectively experienced by 49 %, 38 %, and 44 % of patients. Statistically significant associations were observed between: G2 + acute diarrhea and small bowel V35; G2 + acute genitourinary toxicity and bladder D0.5cc; G2 + inguino-genital skin toxicity and anterior skin V35; G2 + perianal skin toxicity and posterior skin V15; G2 + anemia and lower pelvis bone V45. D0.5 cc was significantly predictive of late toxicity (G2 + anal dysfunction, intestinal toxicity, and inguino-genital/perianal dermatitis). Maximum skin toxicity grade was significantly correlated with the requirement for a treatment break. CONCLUSION: Statistically significant dose-volume parameters were identified and may be used to offer individualized risk prediction and to inform treatment planning. Additional validation of the results is required.

Dosimetric evaluation of adult and paediatric brain tumours planned using mask-based cobalt-60 fractionated stereotactic radiotherapy compared to linear accelerator-based volumetric modulated arc therapy.

INTRODUCTION: We conducted a study to evaluate the dosimetric feasibility of mask-based cobalt-60 fractionated stereotactic radiotherapy (mcfSRT) with the Leksell Gamma Knife® Icon™ device. METHODS: Eleven patients with intracranial tumours were selected for this dosimetry study. These patients, previously treated with volumetric arc therapy (VMAT), were re-planned using mcfSRT. Target volume coverage, conformity/gradient indices, doses to organs at risk and treatment times were compared between the mcfSRT and VMAT plans. Two-sided paired Wilcoxon signed-rank test was used to compare differences between the two plans. RESULTS: The V95 for PTV was similar between fractionated mcfSRT and VMAT (P = 0.47). The conformity index and gradient indices were 0.9 and 3.3, respectively, for mcfSRT compared to 0.7 and 4.2, respectively, for VMAT (P < 0.001 and 0.004, respectively). The radiation exposure to normal brain was lower for mcfSRT across V10, V25 and V50 compared with VMAT (P = 0.007, <0.001 and <0.001, respectively). The median D0.1cc for optic nerve and chiasm as well as the median D50 to the hippocampi were lower for mcfSRT compared to VMAT. Median beam-on time for mcfSRT was 9.7 min per fraction, compared to 0.9 min for VMAT (P = 0.002). CONCLUSION: mcfSRT plans achieve equivalent target volume coverage, improved conformity and gradient indices, and reduced radiation doses to organs at risk as compared with VMAT plans. These results suggest superior dosimetric parameters for mcfSRT plans and can form the basis for future prospective studies.

The synthesis, characterization and immunological activity of mucopolysaccharide-quaternized chitosan nanoparticles.

In this study, nanoparticles were prepared by using positively charged quaternized chitosan and negatively charged mucopolysaccharide such as chondroitin sulfate, heparin and hyaluronic acid. The nanoparticles have a stable nanostructure with particle size in 336.2-424.5 nm, potential in 18.5-31.1 mV and polydispersity index PDI of 0.172-0.335. Moreover, their encapsulation efficiency was 68.77 % and 64.89 %, and they have low endotoxin and good stability. It can significantly promote the expression of IL-6, TNF-α, and IL-1ß of DCS cells. Moreover, the in vivo immune activity of heparin-quaternized chitosan-OVA nanoparticles against BALB/C mice was showed that, the nanoparticles could significantly promote the secretion of immunoglobulins in mice including IgG1 and IgG2. And nanoparticle also can promote the production of immune factors. Meanwhile, the expression of immune factor genes was also elevated. Furthermore, the results of tissue section experiments showed that the nanoparticles are safety of the body.

In-situ generation of H2O2 by zero valent iron to control depolymerization of lignocellulose in composting niche.

Lignocellulosic degradation is a bottleneck of bioconversion during the composting process. In-situ generation of H2O2 in the composting system was an ideal method for efficiently promoting lignocellulase degradation, and zero valent iron (ZVI) was concerned because it can generate H2O2 by reducing dissolved oxygen. This study focused on the effects of ZVI treatment on lignocellulose degradation, microbial communities, and carbohydrate-active enzymes (CAZymes) genes during composting. Its results indicated that ZVI increased H2O2 content during composting, accompanied by the formation of •OH. The degradation rates of lignin, cellulose and hemicellulose in ZVI group (20.77%, 30.35% and 44.7%) were significantly higher than in CK group (17.01%, 26.12% and 38.5%). Metagenomic analysis showed that ZVI induced microbial growth that favored lignocellulose degradation, which increased the abundance of Actinobacteria and Firmicutes but reduced Proteobacteria. At the genus level, the abundance of Thermomonospora, Streptomyces, and Bacillus significantly increased. In addition, glycoside hydrolases and auxiliary activities were important CAZymes families of lignocellulose degradation, and their abundance was higher in the ZVI group. Redundancy analysis showed that the increased H2O2 and •OH content was a critical factor in improving lignocellulose degradation. Overall, H2O2 as a co-substrate enhanced the enzymatic efficiency, •OH unspecifically attacked lignocellulose, and the increase in functional microbial abundance was the main reason for promoting lignocellulose degradation in composting.

The Small Open Reading Frame-Encoded Peptides: Advances in Methodologies and Functional Studies.

Small open reading frames (sORFs) are an important class of genes with less than 100 codons. They were historically annotated as noncoding or even junk sequences. In recent years, accumulating evidence suggests that sORFs could encode a considerable number of polypeptides, many of which play important roles in both physiology and disease pathology. However, it has been technically challenging to directly detect sORF-encoded peptides (SEPs). Here, we discuss the latest advances in methodologies for identifying SEPs with mass spectrometry, as well as the progress on functional studies of SEPs.

Contemporary dose-escalation methods for early phase studies in the immunotherapeutics era.

Phase 1 dose-escalation trials are crucial to drug development by providing a framework to assess the toxicity of novel agents in a stepwise and monitored fashion. Despite widely adopted, rule-based dose-escalation methods (such as 3 + 3) are limited in finding the maximum tolerated dose (MTD) and tend to treat a significant number of patients at subtherapeutic doses. Newer methods of dose escalation, such as model-based and model-assisted designs, have emerged and are more accurate in finding MTD. However, these designs have not yet been broadly embraced by investigators. In this review, we summarise the advantages and disadvantages of contemporary dose-escalation methods, with emphasis on model-assisted designs, including time-to-event designs and hybrid methods involving optimal biological dose (OBD). The methods reviewed include mTPI, keyboard, BOIN, and their variations. In addition, the challenges of drug development (and dose-escalation) in the era of immunotherapeutics are discussed, where many of these agents typically have a wide therapeutic window. Fictional examples of how the dose-escalation method chosen can alter the outcomes of a phase 1 study are described, including the number of patients enrolled, the trial's timeframe, and the dose level chosen as MTD. Finally, the recent trends in dose-escalation methods applied in phase 1 trials in the immunotherapeutics era are reviewed. Among 856 phase I trials from 2014 to 2019, a trend towards the increased use of model-based and model-assisted designs over time (OR = 1.24) was detected. However, only 8% of the studies used non-rule-based dose-escalation methods. Increasing familiarity with such dose-escalation methods will likely facilitate their uptake in clinical trials.

Efficacy of topical and systemic transplantation of mesenchymal stem cells in a rat model of diabetic ischemic wounds.

BACKGROUND: Mesenchymal stem cells (MSCs) exert positive effects in chronic wounds. However, critical parameters, such as the most effective administration routes, remain unclear. Accordingly, the purpose of this study was to compare the effects of topical and systemic transplantation MSCs on diabetic ischemic wound healing and explored the underlying mechanisms. METHOD: A diabetic ischemic wound model was created on the dorsal foot of type 2 diabetes mellitus (T2DM) rat. Bone marrow-derived mesenchymal stem cells (BM-MSCs) were administered via two routes: topical injection and intravenous (IV) infusion. Wound healing outcomes and blood glucose level were assessed dynamically. Meanwhile, blood flow recovery was evaluated in ischemic gastrocnemius muscles. The homing and transdifferentiation of mKate2-labeled BM-MSCs were assessed by fluorescence imaging and immunohistochemistry (IHC) analysis. RESULT: Both topical and systemic treatments had a positive effect on the diabetic ischemic wound showing a significant reduction in wound area at day 14. Histological results showed an increase in the length of epithelial edges, collagen content, microvessel density in the wound bed, and a higher expression of vascular endothelial growth factor (VEGF). Meanwhile, systemic administration can ameliorate hyperglycemia and improve the blood perfusion of the ischemic hindlimb. BM-MSCs administered systemically were found distributed in wounded tissue and transdifferentiated into endothelial cells. Furthermore, BM-MSCs stimulated angiogenesis at wound sites by downregulating phosphatase and tensin homolog (PTEN) and activation of AKT signaling pathway. CONCLUSIONS: The results demonstrated that both transplantation delivery method (topical and systemic) of BM-MSCs accelerated wound healing remarkably under pathological conditions. Nevertheless, systemic administration has the potential to ameliorate hyperglycemia and repair the damaged tissue.

The immunostimulatory effects of hydroxypropyltrimethyl ammonium chloride chitosan-carboxymethyl chitosan nanoparticles.

In this study, we generated chitosan nanoparticles by exploiting the electrostatic interactions between positively charged hydroxypropyltrimethyl ammonium chloride chitosan (HACC) and negatively charged carboxymethyl chitosan (CMC), and examined the effects of altering the molecular weight and carboxymethyl substitution sites of the chitosan molecules. Particle size, potential, and encapsulation efficiency of the various chitosan nanoparticles were examined; the particle size range was 162.40-332.80 nm, the charge range was 19.50-40.60 mV, and the encapsulation efficiency range was 48.4-70.7%. We then examined the immunostimulatory effects of the nanoparticle variants on dendritic cells (DCs); we found that the site of carboxymethyl substitution significantly affected the immunostimulatory effects of the nanoparticles. Two nanoparticle types, 200 kDa N,O-carboxymethyl chitosan-HACC (NO-CMC-HACC) and N-carboxymethyl chitosan-HACC (N-CMC-HACC), greatly promoted the expression of interleukin-6, tumor necrosis factor, and interleukin-1ß in DCs. Moreover, NO-CMC-HACC nanoparticles caused an increase in major histocompatibility complex-II (MHC-II), CD11c, CD80, and CD86 secretion in DCs, indicating that these nanoparticles promoted antigen presentation. We then examined chitosan nanoparticle uptake by DCs using laser confocal microscopy; we found that the NO-CMC-HACC nanoparticles were more readily absorbed by DCs compared to the N-CMC-HACC nanoparticles. Therefore, we concluded that 200 kDa NO-CMC-HACC nanoparticles exhibited strong potential as immunological adjuvants.

Transcriptome Sequencing reveals the expressed profiles of mRNA and ncRNAs and regulate network via ceRNA mediated molecular mechanism of lung adenocarcinoma bone metastasis in Xuanwei.

BACKGROUND: The most ordinary subtype of lung cancer is lung adenocarcinoma (LuAC), which is characterized by strong metastatic ability. And LuAC rates in Xuanwei leads to the poor prognosis and high death rate. In this study, we systematically explored the molecular mechanism of LuAC bone metastasis in Xuanwei by transcriptome sequencing. METHODS: RNA Sequencing was conducted to explore the noncoding RNAs (ncRNAs) expression profiles in primary LuAC and LuAC bone metastasis. We identified differentially expressed mRNAs (DEmRNAs), miRNAs (DEmiRNAs), lncRNAs (DElncRNAs) and circRNAs (DEcircRNAs). Bioinformatics analyses the possible relationships and functions of the LuAC bone metastasis-related competing endogenous RNA (ceRNA). And qRT-PCR was performed to evaluate the expression of these differently expressed genes in serum. RESULTS: A total of 2,141 DEmRNAs, 43 DEmiRNAs, 136 DElncRNAs and 706 DEcircRNAs were identified in the Xuanwei patients with primary LuAC vs. LuAC bone metastasis, respectively. The circRNA/miRNA/mRNA and lncRNA/miRNA/mRNA networks of LuAC in Xuanwei with bone metastasis were built, and the gene expression mechanisms regulated by ncRNAs were unveiled via the ceRNA regulatory networks. We observe that lncRNA (ADAMTS9-AS2, TEX41, DLEU2, LINC00152)-miR-223-3p-SCARB1 and hsa_circ_0000053-miR-196a-5p/miR-196b-5p-HOXA5 ceRNA networks might play an important role in bone metastasis of Xuanwei LuAC. CONCLUSIONS: We comprehensively identified ceRNA regulatory networks of LuAC in Xuanwei with bone metastasis as well as revealed the contribution of different ncRNAs expression profiles. Our data demonstrate the association between mRNAs and ncRNAs in the metastasis mechanism of LuAC in Xuanwei with bone metastasis.

Review: Advances in preparation of chitooligosaccharides with heterogeneous sequences and their bioactivity.

Chitooligosaccharides has attracted increasing attention due to their diverse bioactivities and potential application. Previous studies on the bioactivity of chitooligosaccharides were mostly carried out using a mixture. The structure-function relationship of chitooligosaccharides is not clear. Recently, it is confirmed that chitooligosaccharides with different degrees of polymerization play different roles in many bioactivities. However, heterogeneous chitooligosaccharides with a single degree of polymerization is still a mixture of many uncertain sequences and it is difficult to determine which structure is responsible for biological effects. Therefore, an interesting and challenging field of studying chitooligosaccharides with heterogeneous sequences has emerged. Herein, we reviewed the current methods for preparing heterogeneous chitooligosaccharides, including chemical synthesis, separation techniques and enzymatic methods. Advances in the bioactivities of chitooligosaccharides with heterogeneous sequences are also reviewed.

Immunostimulatory effect of N-2-hydroxypropyltrimethyl ammonium chloride chitosan-sulfate chitosan complex nanoparticles on dendritic cells.

In this study, we synthesized negatively charged chitosan sulfate and positively charged hydroxypropyltrimethyl ammonium chloride chitosan (HACC), and then prepared chitosan derivatives with positive and negative ions as nanoparticles (NPs) by ovalbumin encapsulation using the polyelectrolyte method. NPs with different substitution sites and molecular weights (MW) were prepared by varying conditions. We then determined the zeta potential average, diameter, encapsulation effect, and their immunostimulatory effects on dendritic cells (DCs). The results showed that chitosan-derivative NPs ranged in size from 153.33 to 320.90 nm; all NPs were positive, with charges ranging from 17.10 to 39.30 mV and the encapsulation rates of 65 %-75 %. Three NPs greatly promoted the expression and secretion of interleukin-6 (IL-6), tumor necrosis factor (TNF-α), and interleukin-1ß (IL-1ß) in DC cells: C2,3,6 chitosan sulfate-HACC (C2,3,6-HACC; 200 kDa), C3,6 chitosan sulfate-HACC (C3,6-HACC; 200 kDa) and C6 chitosan sulfate-HACC (C6-HACC; 50 kDa). We also found that 200-kDa C2,3,6-HACC and 50-kDa C6-HACC NPs greatly increased secretion of the major histocompatibility complex-II (MHC-II), CD40, CD80, and CD86, indicating that these NPs promote effective antigen presentation, further increasing immunity effects. Finally, we applied laser confocal photography and determined that NPs entered the cell to promote the regulation of cellular immune activity; this discovery lays a foundation for further research on their mechanism of their action. Therefore, C2,3,6-HACC and C6-HACC NPs have the potential as immunological adjuvants.

A 3-min UPLC-MS/MS method for the simultaneous determination of plasma catecholamines and their metabolites: Method verification and diagnostic efficiency.

OBJECTIVE: To verify a rapid and sensitive ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for the quantification of catecholamines and their metabolites, and to validate its efficiency for the diagnosis of phaeochromocytomas and paragangliomas (PPGLs). METHODS: Plasma samples were pretreated with solid-phase extraction, followed by a 3-min UPLC-MS/MS analysis to quantify epinephrine (E), norepinephrine (NE), dopamine (DA), metanephrine (MN), normetanephrine (NMN) and 3-methoxytyramine (3-MT), simultaneously. The UPLC-MS/MS method was comprehensively verified and its diagnostic efficiency on PPGLs was tested using 7 PPGLs and 408 non-PPGLs patient plasma samples. RESULTS: Using the developed method, the limit of detections (LODs) of the 6 analytes ranged from 0.0002 nmol/L (MN) to 0.0250 nmol/L (NE), while the lower limit of measuring intervals (LLMIs) ranged from 0.05 nmol/L (E, MN and NMN) to 0.10 nmol/L (NE and DA). The reportable ranges were 0.05-30.00 nmol/L for E, MN and NMN, 0.10-30.00 nmol/L for NE and DA, 1.00-300.00 pg/mL for 3-MT. No significant matrix effect was detected after correcting using internal standard. Besides, intra-day and inter-day precision were also within acceptance criteria with coefficient of variations (CVs) ≤ 15% and recoveries ranged from 95% to 115% for all the 6 analytes. The carryover effect was lower than 10%. Its diagnostic efficiency for PPGLs was significantly increased, the areas under the receiver operating characteristic (ROC) curves were increased from 68.7% to 89.1% (using E, NE and DA) to 75.2%-99.9% (using MN, NMN and 3-MT). CONCLUSION: This study verified a rapid UPLC-MS/MS method for the determination of catecholamines and their metabolites in human plasma. It showed high diagnostic efficiency and will serve as an important tool to avoid the risk for missing patients with PPGLs.

Polyion Complex Micelles for Protein Delivery Benefit from Flexible Hydrophobic Spacers in the Binding Group.

Although a range of polymer-protein polyion complex (PIC) micelle systems have been developed in the literature, relatively little attention has been paid to the influence of polymer structure on the assembly, or to the mechanism of disassembly. In this work, Förster resonance energy transfer is used in combination with light sheet fluorescence microscopy and isothermal calorimetry to monitor the formation and stability of PIC micelles with various carboxylic-acid-based binding blocks in MCF-7 cancer spheroid models. All micelles are stable in the presence of free protein, but are unstable in solutions with an ionic strength >200 mm and prone to disassembly at reduced pH. Introducing carbon spacers between the backbone and the binding carboxylic acid results in improved PIC micelle stability at physiological pH, but also increases the pKa of the binding moiety, resulting in improved protein release upon cell uptake. These results give important insights into how to tune PIC micelle stability for controlled protein release in biological environments.

Circular RNA circMTO1 Suppresses RCC Cancer Cell Progression via miR9/LMX1A Axis.

Renal cell carcinoma is one of the most common kidney cancer, which accounts almost 90% of the adult renal malignancies worldwide. In recent years, a new class of endogenous noncoding RNAs, circular RNAs, exert important roles in cell function and certain types of pathological responses, especially in cancers, generally by acting as a microRNA sponge. Circular RNAs could act as sponge to regulate the microRNA and the target genes. However, the knowledge about circular RNAs in renal cell carcinoma remains unclear so far. In the research, we selected a highly expressed novel circular RNAs named circMTO1 in renal cell carcinomas. We investigated the roles of circMTO1 and found that circMTO1 overexpression could suppress cell proliferation and metastases in both A497 and 786-O renal cancer cells, while silencing of circMTO1 could promote the progression in SN12C and OS-RC-2 renal cancer cells. The study showed that circMTO1 acted as miR9 and miR223 sponge and inhibited their levels. Furthermore, silencing of circMTO1 in renal cell carcinoma could downregulate LMX1A, the target of miR-9, resulting in the promotion of renal cell carcinoma cell proliferation and invasion. In addition, LMX1A expression suppression induced by transfection of miR9 mimics confirmed that miR9 exerted its function in renal cell carcinoma by regulating LMX1A expression. What's more, miR9 inhibitor and LMX1A overexpression could block the tumor-promoting effect of circMTO1 silencing. In conclusion, circMTO1 suppresses renal cell carcinoma progression by circMTO1/miR9/ LMX1A, indicating that circMTO1 may be a potential target in renal cell carcinoma therapy.

The bioactivity of new chitin oligosaccharide dithiocarbamate derivatives evaluated against nematode disease (Meloidogyne incognita).

Plant-parasitic nematodes cause substantial crop losses annually; however, current nematicides are environmentally unfriendly and highly toxic to nontarget organisms. The development of green efficient nematicides from multifunctional natural bioactive substances such as chitin oligosaccharide (COS) is promising. In this paper, COS dithiocarbamate derivatives (COSDTC, COSDTA, COSDTB) were synthesized to increase nematicidal activity (against Meloidogyne incognita), and their structures were characterized by FTIR, NMR, TGA/DTG and elemental analysis. Furthermore, the nematicidal activities, egg hatching inhibitory activities, plant growth adjustment abilities, cytotoxicity and phytotoxicity of the derivatives were evaluated. The primary mechanism was assessed by heavy metal ion absorption and GSH-binding assays. The results showed COS dithiocarbamate derivatives could possess multiple efficacies, including high nematicidal activities and egg hatching inhibitory activities, plant growth regulating effects, low cell toxicities and phytotoxicities. Additionally, it was inferred that nematicidal activity may be correlated with GSH-binding activity but not heavy metal ion complexation. COS modification has immense potential for controlling plant-parasitic nematodes.

Advances in chitosan-based nanoparticles for oncotherapy.

Chitosan has attracted considerable attention as an anti-tumor drug carrier material in recent years, which is due to its biocompatibility and biodegradability, as well as the simple and mild preparing techniques of drug-loaded nanoparticles. Chitosan-based nanoparticles can deliver various anti-tumor agents to specific tumor tissues by passive and active targeting mechanisms, including traditional chemotherapeutic agents, DNA or siRNA, proteins, photosensitizers and so on. In this review, we summarized the factors affecting the anti-tumor efficacy of chitosan-based nanoparticles, to aid exploring the function-structure relationship. The recent studies on chitosan-based nanoparticles for oncotherapy were highlighted, including their structures, properties and pharmacological effects. Finally, we offered our perspectives on the challenges and future development of this area.

Immunostimulatory Effects of Chitooligosaccharides on RAW 264.7 Mouse Macrophages via Regulation of the MAPK and PI3K/Akt Signaling Pathways.

Chitooligosaccharides (COS), the hydrolyzed products of chitin and chitosan, can be obtained by various methods. In this study, water-soluble COS were prepared from α- and ß-chitosan by microwave-assisted degradation and their immunostimulatory effects were investigated in RAW 264.7 macrophages. The results indicated that α-COS were more active than ß-COS in promoting the production of nitric oxide (NO) and cytokines, such as tumor necrosis factor-α (TNF-α) and interleukin 6 (IL-6). Quantitative real-time reverse transcription polymerase chain reaction and Western blotting indicated that COS also enhanced the expression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and TNF-α. Further analyses demonstrated that COS induced the phosphorylation of extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), p38, p85 and Akt, and the nuclear translocation of p65, indicating that they are able to activate the mitogen-activated protein kinases (MAPKs) and phosphoinositide 3-kinases (PI3K)/Akt signaling pathways dependent on nuclear factor (NF)-κB activation. In conclusion, COS activate RAW 264.7 cells via the MAPK and PI3K/Akt signaling pathways and are potential novel immune potentiators.

Vascular targeted chitosan-derived nanoparticles as docetaxel carriers for gastric cancer therapy.

A gastric cancer angiogenesis marker peptide, GX1, is promising to be a desirable ligand for anti-angiogenesis targeted drug of gastric cancer treatment. In this study, GX1 was utilized to fabricate a multifunctional vascular targeting docetaxel (DCT)-loaded nanoparticle with N-deoxycholic acid glycol chitosan (DGC) as the carrier and GX1-PEG-deoxycholic acid (GPD) conjugate as the targeting ligand. The mean size of obtained GX1-DGC-DCT was 150.9 nm with a narrow size distribution and their shape was spherical with smooth surface texture. The in vitro drug release test revealed a sustained release manner and an acid pH could accelerate the release compared with the neutral pH. Furthermore, GX1-DGC-DCT showed stronger cytotoxicity against co-cultured gastric cancer cells and human umbilical vein endothelial cells (co-HUVEC) than DCT within 100 µM. In addition, GX1 efficiently enhanced the cellular uptake of nanoparticles in co-HUVEC cells as confirmed by confocal fluorescence scanning microscopy. Moreover, in vivo delivery of GX1-DGC-DCT was demonstrated to inhibit tumor growth in SGC791 tumor-bearing mice with tumor inhibition rate (TIR) of 67.05% and no weight loss of mice was observed. The anti-tumor effects were further confirmed by H&E and TUNEL analysis. Therefore, this new drug delivery system represents a potential strategy for gastric cancer therapy.

Hydroxypropyltrimethyl ammonium chloride chitosan activates RAW 264.7 macrophages through the MAPK and JAK-STAT signaling pathways.

Hydroxypropyltrimethyl ammonium chloride chitosan (HACC) is a water-soluble derivative of chitosan. To investigate the immunostimulatory effects of HACC, quaternized chitosans with different molecular weights were prepared and their effects on RAW 264.7 macrophages were compared. The results showed that HACC promoted nitric oxide (NO) production in a molecular weight- and dose-dependent manner. Lower molecular weight HACC was more active in promoting NO production. Furthermore, flow cytometry analysis showed that HACC significantly promoted the production of interleukin-6 and tumor necrosis factor-α. These results were further demonstrated by quantitive real-time reverse transcription polymerase chain reaction and western blot analysis. Moreover, western blotting revealed that HACC induced the phosphorylation of extracellular signal-regulated kinase, c-Jun N-terminal kinase, p38, and signal transducer and activator of transcription (STAT) proteins. In conclusion, HACC activated RAW 264.7 cells through the mitogen-activated protein kinases and Janus kinase/STAT pathways.