Insight into the mechanism and formation process of bioinspired poly(amino acid)/polyphenol capsules engineered with fast pH switchable permeability.

Capsules have hollow cores and closed wall structures, and they have attracted considerable interest due to their wide applications and significance in life science. The engineering process of bioinspired capsules and related applications have earned heavy concerns. However, the mechanism of capsule formation is often ignored. Herein, based on polyornithine (POR) and tannic acid (TA), two facile strategies to engineer bioinspired capsules were proposed, and the formation mechanisms were deeply explored. We found that the oxidized state of TA had a profound influence not on the thickness or permeability of the formed capsule but on the mechanism and generation process. Compared to TA/POR capsules produced from TA without oxidization (TA/POR), capsules produced from TA with preoxidization (oTA/POR) had thicker shells with higher impermeability. The dominant construction mode in the shells of TA/POR capsules was electrostatic interactions but became Schiff base bonds in oTA/POR capsules. The permeability of oTA/POR displayed pH reversibility and strong pH dependence, with 100% permeability at lower pH and 100% impermeability at pH 7, completing loading/releasing kinetics in minutes at pH 4. We believe these findings contribute to knowledge of bioinspired capsules from engineering processes and formation mechanisms, extending their applications in various fields, such as in drug delivery, artificial cells, and sensors.

Efficient Delivery of Antibodies Intracellularly by Co-Assembly with Hexahistidine-Metal Assemblies (HmA).

PURPOSE: There has been a substantial global market for antibodies, which are based on extracellular targets. Binding intracellular targets by antibodies will bring new chances in antibody therapeutics and a huge market increase. We aim to evaluate the efficiency of a novel delivery system of His6-metal assembly (HmA) in delivering intracellular antibodies and biofunctions of delivered antibodies. METHODS: In this study, the physicochemical properties of HmA@Antibodies generated through co-assembling with antibodies and HmA were well characterized by dynamic light scatter. The cytotoxicity of HmA@Antibodies was investigated by Cell Counting Kit-8 (CCK-8). The endocytic kinetics and lysosome escape process of HmA@Antibodies were studied by flow cytometry and fluorescent staining imaging, respectively. Compared to the commercialized positive control, the intracellular delivery efficiency by HmA@Antibodies and biofunctions of delivered antibodies were evaluated by fluorescent imaging and CCK-8. RESULTS: Various antibodies (IgG, anti-ß-tubulin and anti-NPC) could co-assemble with HmA under a gentle condition, producing nano-sized (~150 nm) and positively charged (~+30 eV) HmA@Antibodies particles with narrow size distribution (PDI ~ 0.15). HmA displayed very low cytotoxicity to divers cells (DCs, HeLa, HCECs, and HRPE) even after 96 h for the feeding concentration ≤100 µg mL-1, and fast escape from endosomes. In the case of delivery IgG, the delivery efficiency into alive cells of HmA was better than a commercial protein delivery reagent (PULSin). For cases of the anti-ß-tubulin and anti-NPC, HmA showed comparable delivery efficiency to their positive controls, but HmA with ability to deliver these antibodies into alive cells was still superior to positive controls delivering antibodies into dead cells through punching holes. CONCLUSION: Our results indicate that this strategy is a feasible way to deliver various antibodies intracellularly while preserving their functions, which has great potential in various applications and treating many refractory diseases by intracellular antibody delivery.

Silanization of a Metal-Polyphenol Coating onto Diverse Substrates as a Strategy for Controllable Wettability with Enhanced Performance to Resist Acid Corrosion.

Wettability is a crucial characteristic of materials that plays a vital role in surface engineering. Surface modification is the key to changing the wettability of materials, and a simple and universal modification approach is being extensively pursued by researchers. Recently, metal-phenolic networks (MPNs) have been widely studied because they impart versatility and functionality in surface modification. However, an MPN is not stable for long periods, especially under acidic conditions, and is susceptible to pollution by invasive species. Spurred by the versatility of MPNs and various functionalities achieved by silanization, we introduce a general strategy to fabricate functionally stable coatings with controllable surface wettability by combining the two methods. The formation process of MPN and silane-MPN coatings was characterized by spectroscopic ellipsometry (SE), UV-visible-near-infrared (UV-vis-NIR) spectroscopy, X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), atomic force microscopy (AFM), water contact angle (WCA), etc. We found that the stability of the MPN was greatly enhanced after silanization, which is attributed to the cross-linking effect that occurs between silane and the MPN, namely, the cross-linking protection produced in this case. Additionally, the wettability of an MPN can be easily changed through our strategy. We trust that our strategy can further extend the applications of MPNs and points toward potential prospects in surface modification.

Effects of Urtica dioica extract on malignant biological behaviors of breast cancer cells and its possible mechanism / 中国肿瘤生物治疗杂志

@#[摘 要] 目的：探讨大荨麻提取物对乳腺癌细胞增殖、凋亡和细胞周期的影响，并初步探讨其可能的作用机制。方法：用不同质量浓度的大荨麻提取物（0、1、2、4、8、16、32、64 mg/ml）处理乳腺癌细胞MCF-7和MDA-MB-231 24 h，MTT法检测细胞增殖活力，选择中位抑制浓度附近的浓度（5和10 mg/ml）作为给药浓度分别处理MCF-7和MDA-MB-231细胞24 h后，平板克隆形成实验和流式细胞术分别检测大荨麻提取物对乳腺癌细胞增殖、周期和凋亡的影响，WB法检测对细胞周期和凋亡相关蛋白以及PI3K/AKT信号通路相关蛋白表达的影响。在MCF-7细胞用5 mg/ml大荨麻提取物处理的同时转染过表达AKT质粒（大荨麻+AKT组），转染空载质粒为对照组（大荨麻+vec组），WB法检测过表达效率，比较过表达AKT对细胞增殖、周期和凋亡的影响。结果：各大荨麻提取物处理组MCF-7和MDA-MB-231细胞增殖活力均显著低于对照组（P<0.05或P<0.01）；与对照组比较，5或10 mg/ml大荨麻处理组乳腺癌细胞的克隆形成数显著减少，G0/G1期细胞占比和凋亡率显著增加（P<0.05或P<0.01），P21、BAX蛋白表达显著升高而Cyclin D1、CDK4、Bcl2蛋白以及p-PI3K、p-AKT蛋白表达显著降低（P<0.05或P<0.01）。大荨麻+AKT组p-AKT和AKT蛋白表达显著高于大荨麻+vec组，克隆数、S期和G2/M期细胞占比均高于大荨麻+vec组（P<0.05或P<0.01），G0/G1期细胞占比和凋亡率低于大荨麻+vec组（P<0.05或P<0.01）。结论：大荨麻提取物可以抑制乳腺癌细胞增殖、促进凋亡且阻滞细胞在G0/G1期，其作用机制可能与抑制PI3K/AKT信号通路相关。

One-Pot Generating Subunit Vaccine with High Encapsulating Efficiency and Fast Lysosome Escape for Potent Cellular Immune Response.

Utilizing nanoparticles to deliver subunit vaccine is considered to be a promising strategy to improve immune response. However, currently reported systems suffered from one or more points, for example, delicate design on molecular structures and elaborate synthesis process, low antigen and/or adjuvant encapsulation efficiency, involvement of toxic materials, and denaturing of bioactivity of antigen and/or adjuvant. To address these issues, here, for the first time, we developed a one-pot method to produce a subunit vaccine by using hexa-histidine metal assembly (HmA) to codeliver tumor-associated antigens (GP100, a peptide KTWGQYWQV) and adjuvant (CpG). The generation of subunit vaccines was detailedly characterized by various techniques, including dynamic scatter, scanning electron microscopy, transmission electron microscopy, UV-visible spectroscopy, agarose gel electrophoresis, etc. HmA displayed high efficiency on encapsulating both subunits (GP100 and CpG) under mild conditions, and the generated subunit vaccine showed a pH-dependent release profile of loaded subunits. In the cellular tests, these subunit vaccines behaved with a quick endocytosis into immune cells and a fast endo/lysosomes escape, inducing maturation of antigen presentative cells and stimulating a potent cellular immune response. These results suggested that HmA is a robust platform for fabricating subunit vaccine, with immense potential for the immunotherapy of various diseases.

[Radix hedysari extract promotes peripheral nerve regeneration].

Treatment of peripheral nerve injury is a major challenge in clinical practice. With advances in molecular biology and development of microsurgical techniques and tissue engineering, peripheral nerve repair procedures have been greatly improved. In the last 10 decades, most treatments for peripheral nerve injury in animal models have achieved histological and functional recovery, the treatments in humans, however, produce insufficient recovery, especially for proximal nerve injury. Increasing attention has been paid to the Traditional Chinese Medicine (TCM) for promoting peripheral nerve regeneration, since these remedies often display effective clinical outcome, minor side effects and effectiveness for multiple targets. Although TCM has complex ingredients and the specific pharmacological mechanisms for their effectiveness are still unclear, an effective clinical outcome is welcomed by many clinicians. In the past 20 years, we have made a series of detailed studies including the toxicity tests, pharmacodynamic tests, pharmacological experiments etc, about a new traditional formula which mainly contains the Radix hedysari, Epimedium etc. RESULTS have shown that this formula is safe to be used in both animals and humans with no toxicity and adverse effect, and systemic administration of this formula could enhance the peripheral nerve regeneration.

Qualitative and quantitative analysis of anthraquinones in rhubarbs by high performance liquid chromatography with diode array detector and mass spectrometry.

Rhubarb is well known in traditional Chinese medicines (TCMs) mainly due to its effective purgative activity. Anthraquinones, including anthraquinone derivatives and their glycosides, are thought to be the major active components in rhubarb. To improve the quality control method of rhubarb, we studied on the extraction method, and did qualitative and quantitative analysis of widely used rhubarbs, Rheum tanguticum Maxim. ex Balf. and Rheum palmatum L., by HPLC-photodiode array detection (HPLC-DAD) and HPLC-mass spectrum (HPLC-MS) on a Waters SymmetryShield RP18 column (250 mm × 4.6 mm i.d., 5 µm). Amount of five anthraquinones was viewed as the evaluating standard. A standardized characteristic fingerprint of rhubarb was provided. From the quantitative analysis, the rationality was demonstrated for ancestors to use these two species of rhubarb equally. Under modern extraction methods, the amount of five anthraquinones in Rheum tanguticum Maxim. ex Balf. is higher than that in Rheum palmatum L. Among various extraction methods, ultrasonication with 70% methanol for 30 min is a promising one. For HPLC analysis, mobile phase consisted of methanol and 0.1% phosphoric acid in water with a gradient program, the detection wavelength at 280nm for fingerprinting analysis and 254 nm for quantitative analysis are good choices.

Influence of different distal nerve degeneration period on peripheral nerve collateral sprouts regeneration.

The effects of the distal nerve degeneration on the regeneration of the collateral sprouts from the proximal nerve stump have been examined. The delayed cross-suture anastomosis technique was used in which the tibial nerve was denervated for 0-8 weeks before cross-suture of the freshly axotomized common peroneal and chronically denervated TIB nerve stumps. There was a remarkable decreasing of the regenerated myelinated axons number after the distal nerve suffered 8 weeks deterioration, suggesting that short-term denervation did not affect the collateral sprouts regeneration but more prolonged denervation profoundly reduced collateral sprouts regenerated in the distal nerve stump.

Lumbricus extract promotes the regeneration of injured peripheral nerve in rats.

BACKGROUND: Earthworms regenerate amputated parts of their body if the nervous system is intact. Lumbricus is one traditional Chinese medicine (TCM), which has been used in China to promote nerve function for hundreds of years. AIM OF THE STUDY: To investigate the beneficial effect of lumbricus extract on peripheral nerve regeneration in rats. MATERIALS AND METHODS: Nerve function was surgically impaired in Sprague-Dawley (SD) rats by clamping of the left sciatic nerve. The sham-operated group (surgery but no sciatic nerve clamping), control group, and treatment group were treated with 2 ml 0.9% NaCl, 0.9% NaCl, and lumbricus extract (1g/ml), respectively. Treatments were administered once daily after the operation for 6 weeks. During this period, motor function was monitored by walking track analysis, conduction function of injured sciatic nerve was monitored by electrophysiology, and regeneration of myelinated nerve was assessed by immunohistochemistry. RESULTS: (1) For nerve function index value, treatment group is higher than control group. (2) For conduction velocity of injured sciatic nerve, treatment group is higher than control group at week 3 and 6. (3) For the number of regenerated myelinated nerve fibers, treatment group is higher than control group at week 2 and 6. CONCLUSIONS: Lumbricus extract appears to enhance sciatic nerve regeneration and function recovery following injury, suggesting the clinical potential of lumbricus extract on the treatment of peripheral nerve injury in humans.

Effects of Hedysari polysaccharides on regeneration and function recovery following peripheral nerve injury in rats.

It has been demonstrated that aqueous extract of Radix Hedysari Prescription and modified Radix Hedysari Prescription could improve the regeneration of injured peripheral nerve. Radix Hedysari is a main component in these two formulas. We hypothesized that Hedysari polysaccharides (HPS), a main active ingredient, could also enhance peripheral nerve regeneration after nerve injury in adult animals. In the present study, we examined the effects of HPS on sciatic nerve regeneration for 6 weeks following clamping in rats (administrated orally of 2 ml HPS liquid daily, 0.25 g/ml). The results showed that HPS was able to enhance sciatic function index (SFI) value, tibial function index (TFI) value, peroneal nerve function index (PFI) value, conduction velocity, and the number of regenerated myelinated nerve fibers, suggesting the potential clinical application of HPS for the treatment of peripheral nerve injury in humans.

[Traditional Chinese medicine and formulas of improving peripheral nerve regeneration].

Peripheral nerve impairment is a common complication in surgery, which repair relates directly to the recovery of motor function and sensory function. Clinical researchers always do nerve sutrure using microsurgical technique and adjuvant treatment to improve peripheral nerve regeneration. Western medicine used usually of adjuvant drugs, such as neurotrophic factors, are limited by their defects in clinical application. Traditional Chinese medicine classifies peripheral nerve impair as paralysis and arthromyodynia, considers that it is the result of defects of meridian and vessels, QI and blood, bones and muscles. So, drugs used usually are QI invigorating herbs, blood circulation promoting herbs for unblocking collaterals, and nourishing herbs, including astragali, hedysari, ginkgo leaf, angelica, danshen root, paeoniae radix, epimedium, chuanxiong, and common basic formulas, such as Buyang Huanwu decoction, Huangqi Guizhi Wuwu decoction, Huoxue Kangyuan decoction, compound radix hedysari, etc. To be ready for further study and development, we review the traditional Chinese medicine and formulas in this article.

[Primary study on effect of various components of modified formula radix hedysari on peripheral nerve regeneration].

OBJECTIVE: To investigate the effects of icariin and mixed prescription of icariin, radix hedysari polysaccharide, and liquid extracted from earthworm on peripheral nerve regeneration. METHODS: Twenty male SD rats weighing (200 +/- 10) g were selected and randomized into four groups (n=5 per group): sham operated group (group A), model group (group B), icariin group (group C), and mixed liquid group (group D). In group A, the left sciatic nerves of the rats were only exposed, and treated at fixed time from the following day with the NS (2 mL/d). In groups B, C, D, the models were made by clamping sciatic nerve and treated with NS, icariin and mixed liquid, respectively (2 mL/d). The general state of animals was observed after the treatment daily. The nerve function index, motor nerve conductive velocity and the amorphous and number of myelinated sciatic nerve fibers were measured at 21 days. RESULTS: Animals in various groups were all in good state. After 21 days, the weights of rats in groups A, B, C and D were (366.9 +/- 14.0), (370.1 +/- 16.3), (373.3 +/- 19.6) and (374.0 +/- 11.4) g, respectively, and there was no significant difference among these groups (P > 0.05). For sciatic function index, there was no significant difference between group A and group D (P > 0.05), between group B and group C (P > 0.05), while there was significant difference between group B and group D (P < 0.05). For tibial function index, there was significant difference between group A and groups B, C, D (P < 0.05), there was no significant difference between group B and groups C, D (P > 0.05). For peroneal function index, there was no significant difference between group A and groups C, D (P > 0.05), between group B and groups C, D (P > 0.05). The sciatic motor nerve conductive velocities of group A, B, C and D were (45.0 +/- 2.9), (8.0 +/- 2.6), (13.4 +/- 6.8), and (19.6 +/- 9.3) m/s, respectively, there was no significant difference between group B and group C (P > 0.05), and there was significant difference between group A and groups B, C, D and between group B and group D (P < 0.05). The size of individual myelinated sciatic nerve fibers of regenerated nerves in groups B, C, and D was significantly smaller than that in group A. Comparing with group A, the number of myelinated sciatic nerve fibers in groups B, C, and D was 93.3% +/- 35.6%, 90.6% +/- 37.1%, and 115.4% +/- 40.6%, respectively, but there was no significant difference among four groups (P > 0.05). CONCLUSION: Icariin and mixed prescription are safe. The improving peripheral nerve regeneration effect of mixed prescription is more obvious than that of icariin, indicating the comprehensive study of modified formula radix hedysari is necessary to find the effective part or mixture of effective compounds with fixed percentage.

Pro-apoptotic and microtubule-disassembly effects of ardisiacrispin (A+B), triterpenoid saponins from Ardisia crenata on human hepatoma Bel-7402 cells.

Ardisiacrispin (A+B) is a mixture of ardisiacrispins A and B, derived from Ardisia crenata with a fixed proportion (2:1). The present study was conducted to investigate its anticancer activity on human cancer cells and its underlying mechanism of action. The (IC50)s of ardisiacrispin (A+B) on proliferation of several human cancer cell lines were in the range of 0.9-6.5 microg/ml by sulphorhodamine B-based colorimetric assay, in which Bel-7402 was the most sensitive cell line. Moreover, ardisiacrispin (A+B) induced dose-dependent apoptosis in Bel-7402 cells at doses of 1-10 microg/ml by flow cytometry, and resulted in the changes of the mitochondrial membrane depolarization, membrane permeability enhancement, and nuclear condensation in a dose-dependent manner through high-content screening analysis. Furthermore, ardisiacrispin (A+B) could disassemble microtubule in Bel-7402 cells; the fluorescence intensity of microtubules decreased at the concentration of 5-20 microg/ml. These findings suggest that ardisiacrispin (A+B) could inhibit the proliferation of Bel-7402 cells by inducing apoptosis and disassembling microtubule.

Induction of apoptosis accompanying with G(1) phase arrest and microtubule disassembly in human hepatoma cells by jaspolide B, a new isomalabaricane-type triterpene.

Jaspolide B is a novel isomalabaricane-type triterpene isolated from sponge Jaspis sp. We investigated its effects on human hepatoma cells in this study. After 48 h of incubation, jaspolide B inhibited the growth of Bel-7402 and HepG2 cells with IC(50) values of 29.1 and 29.5 µM, respectively. Incubation with 0.5 µM of jaspolide B caused time-dependent induction of apoptosis in up to 66.8% of Bel-7402 cells for 48 h, and the induction of apoptosis was confirmed by the enhancement of mitochondrial masses and cell membrane permeability, and nuclear condensation in Bel-7402 and HepG2 cells. Moreover, jaspolide B arrested cell cycle progression at G(1) phase of human hepatoma cells in a dose- and time-dependent manner. In addition, treatment of the compound caused dose-dependent disassembly of microtubule cytoskeleton in Bel-7402 cells at indicated concentrations, and this effect being similar but weaker than that of colchicine, a well-known microtubule-disassembly agent. We conclude that the anti-cancer effect of jaspolide B relates to the apoptosis induction, cell cycle arrest and microtubule disassembly, and these multiple mechanisms of jaspolide B, especially the induction of apoptosis, open interesting perspectives for further exploration of the isomalabaricane-type terpenes and compounds of marine sponge origin as potential anticancer agents.