Smart Responsive Quercetin-Conjugated Glycol Chitosan Prodrug Micelles for Treatment of Inflammatory Bowel Diseases.

The incidence and progression of inflammatory bowel disease are closely related to oxidative stress caused by excessive production of reactive oxygen species (ROS). To develop an efficacious and safe nanotherapy against inflammatory bowel diseases (IBD), we designed a novel pH/ROS dual-responsive prodrug micelle GC-B-Que as an inflammatory-targeted drug, which was comprised by active quercetin (Que) covalently linked to biocompatible glycol chitosan (GC) by aryl boronic ester as a responsive linker. The optimized micelles exhibited well-controlled physiochemical properties and stability in a physiological environment. Time-dependent NMR spectra traced the changes in the polymer structure in the presence of H2O2, confirming the release of the drug. The in vitro drug release studies indicated a low release rate (<20 wt %) in physiological conditions, but nearly complete release (>95 wt % after 72 h incubation) in a pH 5.8 medium containing 10 µM H2O2, exhibiting a pH/ROS dual-responsive property and sustained release behavior. Importantly, the negligible drug release in a simulated gastric environment in 1 h allowed us to perform intragastric administration, which has potential to achieve the oral delivery by mature enteric-coating modification in future. Further in vivo activities and biodistribution experiments found that the GC-B-Que micelles tended to accumulate in intestinal inflammation sites and showed better therapeutic efficacy than the free drugs (quercetin and mesalazine) in a colitis mice model. Typical inflammatory cytokines including TNF-α, IL-6, and iNOS were significantly suppressed by GC-B-Que micelle treatment. Our work promoted inflammatory-targeted delivery and intestinal drug accumulation for active single drug quercetin and improved the therapeutic effect of IBD. The current study also provided an alternative strategy for designing a smart responsive nanocarrier for a catechol-based drug to better achieve the target drug delivery.

Inducing Optimal Antitumor Immune Response through Coadministering iRGD with Pirarubicin Loaded Nanostructured Lipid Carriers for Breast Cancer Therapy.

Chemotherapeutic agents trigger antitumor immune response through inducing immunogenic tumor cell death. However, severe toxicity to immune system and insufficient immunogenic cell death hinder chemotherapy from arousing efficient antitumor immunity in vivo. In this study, the cytotoxic drug, pirarubicin (THP), was entrapped into nanostructured lipid carriers (NLC); THP-NLC significantly reduced the toxicity of THP to immune system and improved immune status of breast cancer bearing mice. When THP-NLC was coinjected with iRGD (a tumor-penetrating peptide), drug accumulation in tumors was greatly elevated, which led to significant control of tumor growth and increase of immunogenic tumor cell death. Subsequently, the cytotoxic T lymphocytes (CD3+ and CD8+ cells) infiltration and cytokine (IFN-γ and INF-α) secretion in tumors were heavily increased. The efficient T-cell dependent control of tumors in the late stage and the lower side effects contributed to the longest whole survival of THP-NLC + iRGD treated mice. Therefore, the coadministration of THP-NLC with iRGD resulted in increased tumor cell direct-killing death and enhanced antitumor immune response. Our results illustrated that THP could serve as an immunogenic cell death inducer and the proposed drug delivery strategy might impact cancer immunotherapy by arousing increased immunogenic tumor cell death.

Bu-Zhong-Yi-Qi Granule Enhances Colonic Tight Junction Integrity via TLR4/NF-κB/MLCK Signaling Pathway in Ulcerative Colitis Rats.

BACKGROUND: Bu-zhong-yi-qi granule (BZYQ), a sort of Chinese herbal medicine, has exhibited therapeutic effects on ulcerative colitis (UC). However, the mechanism of BZYQ has not been fully clarified. This study was aimed at investigating the effects of BZYQ on UC rats model and at exploring its potential mechanism. METHODS: The UC rats were established by enema of trinitrobenzene sulfonic acid (TNBS). The therapeutic effects of BZYQ treatment were evaluated by disease activity index (DAI), colon macroscopic damage index (CMDI) scores, and histological observation. The mRNA levels of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and interleukin-10 (IL-10) were measured by quantitative real time-polymerase chain reaction (qPCR). The expression of tight junction (TJ) proteins, occludin and claudin-1, in the colon was determined by Western blot and immunofluorescence. The expression of toll-like receptors 4 (TLR4), nuclear factor-kappa B (NF-κB), p-NF-κB, myosin light chain kinase (MLCK), MLC, and p-MLC levels in colon was determined by Western blot or qPCR. RESULTS: The results showed that BZYQ could attenuate DAI, CMDI, and histological inflammation. TJ proteins expression was decreased in UC rats, but treatment with BZYQ restored the expression of occludin and claudin-1. In addition, BZYQ administration ameliorated UC-associated increase in the production of TNF-α, IL-1ß, and the expression of TLR4, NF-κB, p-NF-κB, MLCK, MLC, and p-MLC, while BZYQ administration increased the production of IL-10. CONCLUSIONS: The therapeutic effect of BZYQ on UC is at least partially through regulation of the secretion of some inflammatory cytokines and improvement of TJ integrity via TLR4/NF-κB/MLCK pathway.

Pectic polysaccharides from Radix Sophorae Tonkinensis exhibit significant antioxidant effects.

Two pectic polysaccharides (WRSP-A2b and WRSP-A3a) have been obtained from Radix Sophorae Tonkinensis and comparatively investigated in terms of their physical properties and antioxidant activities. Monosaccharide composition, FT-IR, NMR and enzymatic analyses indicate that both WRSP-A2b (13.6 kDa) and WRSP-A3a (44.6 kDa) consist of homogalacturonan (HG), rhamnogalacturonan I (RG-I) and rhamnogalacturonan II (RG-II) domains, with mass ratios of 0.9:1.8:1 and 2.3:2.9:1, respectively. The RG-I domains were further purified and characterized. Results show that WRSP-A2b contains a highly branched RG-I domain, primarily substituted with α-(1→5)-linked arabinans, whereas WRSP-A3a contains a small branched RG-I domain mainly composed of ß-(1→4)-linked galactan side chains. WRSP-A3a exhibits stronger antioxidant activity in scavenging different radicals than WRSP-A2b, a finding that may be due to its higher content of GalA residues and HG domains. Our results provide useful information for screening natural polysaccharide-based antioxidants from Radix Sophorae Tonkinensis.

An efficient protocol for the preparation of linear arabino-oligosaccharides.

In this study, we report the development of an efficient protocol in the preparation of linear arabino-oligosaccharides derived from sugar beet arabinan. By optimizing hydrolytic conditions and separation on tandem Bio-Gel P4 columns, we obtained arabino-oligosaccharides with various degrees of polymerization (DP) from 2 to 15. All of these α-1,5-linked arabino-oligosaccharides are highly pure (>95%) as determined by HPAEC, MALDI-TOF mass spectrometry and 13C NMR spectroscopy. Due to their purity, these oligosaccharides can be used as standards to identify other oligosaccharides and as substrates to characterize new arabinan-specific enzymes, as well as for the development of new functional oligosaccharides.

Reactive oxygen species-responsive amino acid-based polymeric nanovehicles for tumor-selective anticancer drug delivery.

Stimuli-triggered drug delivery systems have been recognized as a crucial strategy to achieve on-demand drug release at the tumor for improving therapeutic efficacy. In this work, novel biocompatible and biodegradable reactive oxygen species (ROS)-responsive amino acid- based polymeric micelles were developed for tumor-specific drug release triggered by high ROS levels in cancer cells, which were composed of amphiphilic poly(aspartic acid) (PASP) derivatives (PASP-BSer) with phenylborate serine (BSer) side groups as the ROS-responsive unit. A series of PASP-BSer conjugates with different degree of substitution (DS) were synthesized, and their self-assembly and H2O2-responsive behaviors were investigated to optimize the structure of PASP-BSer. In vitro drug loading and release studies confirmed that the optimized PASP-BSer micelles could effectively encapsulate the model anticancer drug doxorubicin (Dox) and exhibit desirable H2O2-triggered release behaviors. More importantly, Dox-loaded PASP-BSer micelles showed high selective cytotoxicity against A549 cancer cells than L929 normal cells. Accordingly, PASP-BSer micelles have significant potential as on-demand drug carriers for anticancer therapy.

Tumors and Their Microenvironment Dual-Targeting Chemotherapy with Local Immune Adjuvant Therapy for Effective Antitumor Immunity against Breast Cancer.

Chemotherapy turns tumor cells into "tumor vaccines" by immunogenic cell death (ICD). However, it remains a challenge to exploit chemotherapy-induced "tumor vaccines" for solid cancer immunotherapy due to the inefficient effector T cells activation and tumor microenvironment immunosuppression. Here, a matrix metalloprotease 2 responsive liposome (PEG-FA-Lip) composed of cleavable PEG chains covering the folate (FA)-modified liposome is developed to deliver ICD inducer doxorubicin. In breast cancer-bearing mice, PEG-FA-Lip targets both 4T1 breast cancer cells and M2-tumor associated macrophages (M2-TAMs) via FA-receptor mediated endocytosis, resulting in abundant "tumor vaccines" and efficient elimination of M2-TAMs. The combination of local cytosine-phosphate-guanine (CpG) therapy facilitates PEG-FA-Lip induced "tumor vaccines" to effectively arouse systematic effector T cells immune response through promoting dendritic cell maturation and immunostimulatory cytokines secretion. The simultaneous elimination of M2-TAMs ensures the activated effector T cells exert antitumor immunity within tumor via decreasing immunosuppressive cytokines secretion and tumor infiltration of Treg cells. After receiving the combined treatment, 30.1% of breast cancer-bearing mice (initial tumor volume > 100 mm3) achieves the goal of tumor eradication. Remarkably, this combination therapy greatly inhibits lung metastasis and controls the growth of already metastasized breast cancers (initial tumor volume > 100 mm3).

Golgi Apparatus-Targeted Chondroitin-Modified Nanomicelles Suppress Hepatic Stellate Cell Activation for the Management of Liver Fibrosis.

Liver fibrosis is a serious liver disease associated with high morbidity and mortality. The activation of hepatic stellate cells (HSCs) and the overproduction of extracellular matrix proteins are key features during disease progression. In this work, chondroitin sulfate nanomicelles (CSmicelles) were developed as a delivery system targeting HSCs for the treatment of liver fibrosis. CS-deoxycholic acid conjugates (CS-DOCA) were synthesized via amide bond formation. Next, retinoic acid (RA) and doxorubicin (DOX) were encapsulated into CSmicells to afford a DOX+RA-CSmicelles codelivery system. CSmicelles were selectively taken up in activated HSCs and hepatoma (HepG2) cells other than in normal hepatocytes (LO2), the internalization of which was proven to be mediated by CD44 receptors. Interestingly, DOX+RA-CSmicelles preferentially accumulated in the Golgi apparatus, destroyed the Golgi structure, and ultimately downregulated collagen I production. Following tail-vein injection, DOX+RA-CSmicelles were delivered to the cirrhotic liver and showed synergistic antifibrosis effects in the CCl4-induced fibrotic rat model. Further, immunofluorescence staining of dissected liver tissues revealed CD44-specific delivery of CS derivatives to activated HSCs. Together, our results demonstrate the great potential of CS based carrier systems for the targeted treatment of chronic liver diseases.

Novel Low-Toxic Derivative of Celastrol Maintains Protective Effect against Acute Renal Injury.

This study aimed to novelly design and synthesize an amide derivative as a potential substitute of celastrol (CLT). We constituted the compound celastrol-glucosamine (CLG) by conjugating 1-(2-aminoethoxy)-2-glucosamine to celastrol (CLT) and confirmed its chemical structure by 1H NMR, 13C NMR, and LC-MS/MS. Then, the potential efficacy of the CLG was investigated on renal ischemia-reperfusion injury animal models. The results demonstrated that the decorated compound CLG could completely reverse the disease progression as same as CLT. Furthermore, the toxicity of CLG was also fully evaluated in rat blood, liver, kidney, heart, spleen, lung, and reproductive system. Compared to the performance of CLT on normal organs, CLG could remarkably maintain high safety and significantly reduce the side effects. Taken together, the CLG could keep the same efficacy as CLT while processing lower toxicity in vivo.

Effects of Fengliao-Changweikang in Diarrhea-predominant Irritable Bowel Syndrome Rats and Its Mechanism Involving Colonic Motility.

BACKGROUND/AIMS: This study was designed to investigate the effect of Fengliao-Changweikang (FLCWK) in diarrhea-predominant irritable bowel syndrome (IBS-D) rats and explore its underlying mechanisms. METHODS: IBS-D model rats were induced by neonatal maternal separation (NMS) combined with restraint stress (RS). In in vivo experiments, the model rats were randomly divided into 5 groups: NMS + RS, FLCWK (low dose, middle dose, and high dose), and pinaverium bromide. The normal control (no handling) rats were classified as the NH group. The therapeutic effect of FLCWK was evaluated by fecal characteristics, electromyographic response and abdominal withdrawal reflex scores. In in vitro experiments, the model rats were randomly divided into 2 groups: NMS + RS, FLCWK (middle dose), and no handling rats were used as the NH group. The differences in basic tension and ACh-induced tension of isolated colonic longitudinal smooth muscle strips (CLSMs) among the 3 groups were observed. In addition, different inhibitors (nifedipine, TMB-8, L-NAME, methylene blue, and 4-AP) were pretreated to explore the underlying mechanisms. RESULTS: In in vivo experiments, fecal characteristics, electromyographic response, and abdominal withdrawal reflex scores significantly improved in the FLCWK group, compared with the NMS + RS group. In in vitro experiments, the basic tension and ACh-induced tension of CLSMs in IBS-D rats were significantly inhibited by FLCWK. After pre-treatment with different inhibitors, the ACh-induced tension of CLSMs in each group showed no significant difference. CONCLUSIONS: FLCWK manifested curative effect in IBS-D rats by inhibiting colonic contraction. The underlying mechanisms may be related to regulatory pathway of nitric oxide/cGMP/Ca²âº and specific potassium channels.

A novel dexamethasone-loaded liposome alleviates rheumatoid arthritis in rats.

In this study, an inflammation-targeted delivery system based on a liposomal carrier was developed to deliver hydrophobic dexamethasone against arthritis. Using two FDA-approved excipients for intravenous injection, dexamethasone loaded liposome (Dex-Lip) was prepared by a thin-film hydration method. The biodistribution of 1,1'-dioctadecyl-3,3,3',3'-tetramethylindodicarbocyanine-loaded liposomes (DiD-Lips) were performed in rats with adjuvant-induced arthritis and demonstrated specific targeting efficacy in the disease site. DiD-Lips showed prolonged retention time in the inflammatory joint tissues compared with free DiD. Dex-Lips effectively suppressed the joint swelling in arthritis rats and significantly down-regulated serum pro-inflammatory cytokines including tumor necrosis factor-α and interleukin-1ß when compared to free dexamethasone. Furthermore, Dex-Lips had no significantly impact on the body weight, alleviated the hyperglycemia and improved haematological profiles of rheumatoid arthritis rats during the treatment process. Taken together, a safe liposomal delivery system was developed to achieve inflammation targeted therapy against arthritis.

Effects of Hemp seed soft capsule on colonic ion transport in rats.

AIM: To investigate the effect of Hemp seed soft capsule (HSCC) on colonic ion transport and its related mechanisms in constipation rats. METHODS: Sprague-Dawley male rats were randomly divided into three groups: normal group, constipation group and HSSC group. Rats in the constipation and HSSC groups were administrated loperamide 3 mg/kg per day orally for 12 d to induce the constipation model. Then, the HSSC group was given HSSC 0.126 g/kg per day by gavage for 7 d. The normal and constipation groups were treated with distilled water. After the treatment, the fecal wet weight and water content were measured. The basal short-circuit current (Isc) and resistance were measured by an Ussing Chamber. Besides the in vivo drug delivery experiment above, an in vitro drug application experiment was also conducted. The accumulative concentrations of HSSC (0.1 mg/mL, 0.5 mg/mL, 1.0 mg/mL, 2.5 mg/mL, 5.0 mg/mL, 10.0 mg/mL and 25.0 mg/mL) were added to the normal isolated colonic mucosa and the Isc was recorded. Further, after the application of either ion (Cl- or HCO3-) substitution, ion channel-related inhibitor (N-phenylanthranilic acid, glybenclamide, 4,4-diisothiocyano-2,2-stilbenedisulfonic acid or bumetanide) or neural pathway inhibitor [tetrodotoxin (TTX), atropine, or hexamethonium], the Isc induced by HSSC was also measured. RESULTS: In the constipation group, the fecal wet weight and the water content were decreased in comparison with the normal group (P < 0.01). After the treatment with HSSC, the fecal wet weight and the water content in the HSSC group were increased, compared with the constipation group (P < 0.01). In the constipation group, the basal Isc was decreased and resistance was increased, in comparison with the normal group (P < 0.01). After the treatment with HSSC, the basal Isc was increased (P < 0.05) and resistance was decreased (P < 0.01) in the HSSC group compared with the constipation group. In the in vitro experiment, beginning with the concentration of 1.0 mg/mL, differences in Isc were found between the experimental mucosa (with HSSC added) and control mucosa. The Isc of experimental mucosa was higher than that of control mucosa under the same concentration (1.0 mg/mL, P < 0.05; 2.5-25 mg/mL, P < 0.01). After the Cl- or HCO3- removal and pretreated with different inhibitors (cAMP-dependent and Ca2+-dependent Cl- channels, Na+-K+-2Cl- cotransporter (NKCC), Na+-HCO3- cotransporter or Cl-/HCO3- exchanger inhibitor), there were differences between experimental mucosa and control mucosa; the Isc of experimental mucosa was lower than that of control mucosa under the same concentration (P < 0.05). Meanwhile, after pretreatment with neural pathway inhibitor (TTX, atropine, or hexamethonium), there were no differences between experimental mucosa and control mucosa under the same concentration (P > 0.05). CONCLUSION: HSSC ameliorates constipation by increasing colonic secretion, which is mediated via the coaction of cAMP-dependent and Ca2+-dependent Cl- channels, NKCC, Na+-HCO3- cotransporter or Cl-/HCO3- exchanger.