The Controlled Preparation of a Carrier-Free Nanoparticulate Formulation Composed of Curcumin and Piperine Using High-Gravity Technology.

Carrier-free nanoparticulate formulations are an advantageous platform for the oral administration of insoluble drugs with the expectation of improving their bioavailability. However, the key limitation of exploiting carrier-free nanoparticulate formulations is the controlled preparation of drug nanoparticles on the basis of rational prescription design. In the following study, we used curcumin (Cur) and piperine (Pip) as model water-insoluble drugs and developed a new method for the controlled preparation of carrier-free drug nanoparticles via multidrug co-assembly in a high-gravity environment. Encouraged by the controlled regulation of the nucleation and crystal growth rate of high-gravity technology accomplished by a rotating packed bed, co-amorphous Cur-Pip co-assembled multidrug nanoparticles with a uniform particle size of 130 nm were successfully prepared, exhibiting significantly enhanced dissolution performance and in vitro cytotoxicity. Moreover, the hydrogen bonding interactions between Cur and Pip in nanoparticles provide them with excellent re-dispersibility and storage stability. Moreover, the oral bioavailability of Cur was dramatically enhanced as a result of the smaller particle size of the co-assembled nanoparticles and the effective metabolic inhibitory effect of Pip. The present study provides a controlled approach to preparing a carrier-free nanoparticulate formulation through a multidrug co-assembly process in the high-gravity field to improve the oral bioavailability of insoluble drugs.

The injury progression in acute blast-induced mTBI in rats reflected by DTI and immunohistochemical examination.

Diffusion tensor imaging (DTI) has emerged as a promising neuroimaging tool for detecting blast-induced mild traumatic brain injury (bmTBI). However, lack of refined acute-phase monitoring and reliable imaging biomarkers hindered its clinical application in early diagnosis of bmTBI, leading to potential long-term disability of patients. Here, we used DTI in a rat model of bmTBI generated by exposing to single lateral blast waves (151.16 and 349.75 kPa, lasting 47.48 ms) released in a confined bioshock tube (BST-I) to investigate whole-brain DTI changes in the acute-phase of bmTBI at 1, 3, 7 days after injury. Combined assessment of immunohistochemical analysis, transmission electron microscopy (TEM) and behavioral readouts allowed for linking DTI changes to synchronous cellular damages and identifying stable imaging biomarkers. The corpus callosum (CC) and brainstem were identified as predominantly affected regions, in which reduced fractional anisotropy (FA) was detected as early as the first day after injury, with a maximum decline occurring at 3 days after injury before returning to near normal levels by 7 days. Axial diffusivity (AD) values within the CC and brainstem also significantly reduced at 3 days after injury. In contrast, the radial diffusivity (RD) in the CC showed acute elevation, peaking at 3 days after injury before normalizing by the 7-day time point. Damages to nerve fibers, including demyelination and axonal degeneration, progressed in lines with changes in DTI parameters, supporting a real-time macroscopic reflection of microscopic neuronal fiber injury by DTI. The most sensitive biomarker was identified as a decrease in FA, AD and an increase in RD within the CC on the third day after injury, supporting the diagnostic utility of DTI in cases of bmTBI in the acute phase.

Nanodrug Delivery Systems in Antitumor Immunotherapy.

Cancer has become one of the most important factors threatening human health, and the global cancer burden has been increasing rapidly. Immunotherapy has become another clinical research hotspot after surgery, chemotherapy, and radiotherapy because of its high efficiency and tumor metastasis prevention. However, problems such as lower immune response rate and immune-related adverse reaction in the clinical application of immunotherapy need to be urgently solved. With the development of nanodrug delivery systems, various nanocarrier materials have been used in the research of antitumor immunotherapy with encouraging therapeutic results. In this review, we mainly summarized the combination of nanodrug delivery systems and immunotherapy from the following 4 aspects: (a) nanodrug delivery systems combined with cytokine therapy to improve cytokines delivery in vivo; (b) nanodrug delivery systems provided a suitable platform for the combination of immune checkpoint blockade therapy with other tumor treatments; (c) nanodrug delivery systems helped deliver antigens and adjuvants for tumor vaccines to enhance immune effects; and (d) nanodrug delivery systems improved tumor treatment efficiency and reduced toxicity for adoptive cell therapy. Nanomaterials chosen by researchers to construct nanodrug delivery systems and their function were also introduced in detail. Finally, we discussed the current challenges and future prospects in combining nanodrug delivery systems with immunotherapy.

Structural characterization and immunomodulatory activity of a polysaccharide from Dioscotea opposita.

The purified polysaccharides fraction, DOP-2, was prepared from Dioscorea opposita Thunb (D. opposita). This study combined in vitro and in vivo experiments to comprehensively investigate the index changes in RAW264.7 cells and immunocompromised mice under DOP-2 intervention, aiming to elucidate the potential mechanisms of immunomodulatory effects of DOP-2. DOP-2 (10 âˆ¼ 500 µg/mL) significantly elevated the levels of NO, interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) factors secreted by RAW264.7 cells, and restored the body weight of immunosuppressed mice and improve the degree of injury to the immune organ index, resulting in significant immunomodulatory effects. Notably, DOP-2 promoted the production of short-chain fatty acids (SCFAs) in immunosuppressed mice and modulated the composition of their gut microflora. These findings highlight the potential benefits of DOP-2 therapy in improving immune function and gut health, and will provide a theoretical basis for the application of D. opposita polysaccharides as an immunomodulatory adjuvant.

Alleviation of endothelial dysfunction of Pheretima guillemi (Michaelsen)-derived protein DPf3 in ponatinib-induced thrombotic zebrafish and mechanisms explored through ox-LDL-induced HUVECs and TMT-based proteomics.

ETHNOPHARMACOLOGICAL RELEVANCE: Thrombus generation is one of the leading causes of death in human, and vascular endothelial dysfunction is a major contributor to thrombosis. Pheretima guillemi (Michaelsen), a traditional medicinal animal known as "Dilong", has been utilized to cure thrombotic disorders for many years. DPf3, a group of functional proteins extracted from P. guillemi, has been characterized and identified to possess antithrombotic bioactivity via in vitro and ex vivo experiments. AIM OF THE STUDY: This study is aimed to investigate the vascular-protection activity and related mechanism of antithrombotic protein DPf3 purified from Pheretima guillelmi systematically. MATERIALS AND METHODS: The antithrombotic activity and vascular endothelium protection effect of DPf3 was explored in vivo using ponatinib-induced vascular endothelial injury zebrafish thrombus model. Then, (hi) ox-LDL-induced HUVECs was applied to investigate the protection mechanism of DPf3 against the injury of vascular endothelium. In addition, TMT-based proteomics analysis was used to study the biomarkers, biological processes and signal pathways involved in the antithrombotic and vascular protective effects of DPf3 holistically. RESULTS: DPf3 exerted robust in vivo antithrombosis and vascular endothelial protection ability. DPf3 was identified to prevent HUVECs from damage by reducing ROS production, and to reduce monocyte adhesion by decreasing the protein content of adhesion factor VCAM 1. DPf3 was also observed to weaken the migration ability of injured cells and inhibit abnormal angiogenesis. The mechanism of DPf3's antithrombotic and vascular protective activity was mainly related to the regulation of lipid metabolism, energy metabolism, complement and coagulation system, ECM receptor interaction, MAPK signal pathway, etc. CONCLUSIONS: This study demonstrates that DPf3 has strong antithrombotic and endothelial protective effects. The endothelial protective ability and related mechanisms of DPf3 provide a scientific reference for the traditional use of earthworms in the treatment of thrombosis.

The Structural and Electronic Engineering of Molybdenum Disulfide Nanosheets as Carbon-Free Sulfur Hosts for Boosting Energy Density and Cycling Life of Lithium-Sulfur Batteries.

The compact sulfur cathodes with high sulfur content and high sulfur loading are crucial to promise high energy density of lithium-sulfur (Li-S) batteries. However, some daunting problems, such as low sulfur utilization efficiency, serious polysulfides shuttling, and poor rate performance, are usually accompanied during practical deployment. The sulfur hosts play key roles. Herein, the carbon-free sulfur host composed of vanadium-doped molybdenum disulfide (VMS) nanosheets is reported. Benefiting from the basal plane activation of molybdenum disulfide and structural advantage of VMS, high stacking density of sulfur cathode is allowed for high areal and volumetric capacities of the electrodes together with the effective suppression of polysulfides shuttling and the expedited redox kinetics of sulfur species during cycling. The resultant electrode with high sulfur content of 89 wt.% and high sulfur loading of 7.2 mg cm-2 achieves high gravimetric capacity of 900.9 mAh g-1 , the areal capacity of 6.48 mAh cm-2 , and volumetric capacity of 940 mAh cm-3 at 0.5 C. The electrochemical performance can rival with the state-of-the-art those in the reported Li-S batteries. This work provides methodology guidance for the development of the cathode materials to achieve high-energy-density and long-life Li-S batteries.

Protective Effect of Trillium tschonoskii Maxim Components Against Glutamate-Induced SH-SY5Y Cells Damage Through Regulating Apoptosis.

Context: Among the Tujia people, the root or rhizome of Trillium tschonoskii Maxim.in Bull.Acad (TTM) is considered a miraculous herb for headaches. Previous studies have shown ethyl acetate extract (TTM1) can protect SH-SY5Y cells against glutamate injury. Objective: This study clarified TTM1's mechanism against glutamate-induced cell damage, focusing on the regulation of apoptosis. The compounds were separated, identified, and performed molecular docking with pro-apoptotic proteins. Materials and Methods: SH-SY5Y cells were treated with glutamate (2 mM) for 12 hour, and the effect of TTM1 (2.5, 5, 10, and 20 µg/mL) was evaluated with MTT and LDH release assays, taking EGb761(40 µg/mL) as a control. Cell apoptosis was detected with Hoechst 33258 and Annexin V-FITC and measurements of intracellular calcium and caspase-3. The major components were separated and identified by LCMS-IT-TOF and NMR, then the proapoptotic activity of TTM1 was confirmed by molecular docking method. Results: TTM1 protected SH-SY5Y cells by resisting apoptosis, TTM1 (10 and 20 µg/mL) decreased apoptotic bodies and nuclear fragments, increased the proportion of normal cells to 68.38 ± 5.63% and 92.80 ± .88%, decreased VA cells to 4.30 ± .76% and 3.58 ± .45% and caspase-3 to .365 ± .034 and .344 ± .047 ng/mL.TTM1 (10 µg/mL) decreased intracellular free calcium to 2.77 ± .40. Polyphyllin VI and pennogenin 3-O-ß-chacotrioside were identified in TTM1 at 15.04% and 2.84%, and had potential anti-apoptosis activities. Discussion and Conclusions: Folk records of TTM for headache may be related to its anti-apoptosis of nerve cells. Identification and content determination of index components based on effective extract provides research paradigms for rare and endangered ethnic plants.

ETI signaling nodes are involved in resistance of Hawaii 7996 to Ralstonia solanacearum-induced bacterial wilt disease in tomato.

Bacterial wilt caused by the soil-borne pathogen Ralstonia solanacearum is a destructive disease of tomato. Tomato cultivar Hawaii 7996 is well-known for its stable resistance against R. solanacearum. However, the resistance mechanism of Hawaii 7996 has not yet been revealed. Here, we showed that Hawaii 7996 activated root cell death response and exhibited stronger defense gene induction than the susceptible cultivar Moneymaker after R. solanacearum GMI1000 infection. By employing virus-induced gene silencing (VIGS) and CRISPR/Cas9 technologies, we found that SlNRG1-silenced and SlADR1-silenced/knockout mutant tomato partially or completely lost resistance to bacterial wilt, indicating that helper NLRs SlADR1 and SlNRG1, the key nodes of effector-triggered immunity (ETI) pathways, are required for Hawaii 7996 resistance. In addition, while SlNDR1 was dispensable for the resistance of Hawaii 7996 to R. solanacearum, SlEDS1, SlSAG101a/b, and SlPAD4 were essential for the immune signaling pathways in Hawaii 7996. Overall, our results suggested that robust resistance of Hawaii 7996 to R. solanacearum relied on the involvement of multiple conserved key nodes of the ETI signaling pathways. This study sheds light on the molecular mechanisms underlying tomato resistance to R. solanacearum and will accelerate the breeding of tomatoes resilient to diseases.

[Effect of nanoparticles of different stiffness combined with menthol/curcumol on mechanical properties of bEnd.3 cells].

This study aimed to investigate the effects of nanoparticles PLGA-NPs and mesoporous silicon nanoparticles(MSNs) of different stiffness before and after combination with menthol or curcumol on the mechanical properties of bEnd.3 cells. The particle size distributions of PLGA-NPs and MSNs were measured by Malvern particle size analyzer, and the stiffness of the two nanoparticles was quantified by atomic force microscopy(AFM). The bEnd.3 cells were cultured in vitro, and the cell surface morphology, roughness, and Young's modulus were examined to characterize the roughness and stiffness of the cell surface. The changes in the mechanical properties of the cells were observed by AFM, and the structure and expression of cytoskeletal F-actin were observed by a laser-scanning confocal microscope. The results showed that both nanoparticles had good dispersion. The particle size of PLGA-NPs was(98.77±2.04) nm, the PDI was(0.140±0.030), and Young's modulus value was(104.717±8.475) MPa. The particle size of MSNs was(97.47±3.92) nm, the PDI was(0.380±0.016), and Young's modulus value was(306.019±8.822) MPa. The stiffness of PLGA-NPs was significantly lower than that of MSNs. After bEnd.3 cells were treated by PLGA-NPs and MSNs separately, the cells showed fine pores on the cell surface, increased roughness, decreased Young's modulus, blurred and broken F-actin bands, and reduced mean gray value. Compared with PLGA-NPs alone, PLGA-NPs combined with menthol or curcumol could allow deepened and densely distributed surface pores of bEnd.3 cells, increase roughness, reduce Young's modulus, aggravate F-actin band breakage, and diminish mean gray value. Compared with MSNs alone, MSNs combined with menthol could allow deepened and densely distributed surface pores of bEnd.3 cells, increase roughness, reduce Young's modulus, aggravate F-actin band breakage, and diminish mean gray value, while no significant difference was observed in combination with curcumol. Therefore, it is inferred that the aromatic components can increase the intracellular uptake and transport of nanoparticles by altering the biomechanical properties of bEnd.3 cells.

Differences in in vivo absorption of flavone glycosides, flavone aglycones and terpene lactones under different dosage forms and physiological conditions.

ETHNOPHARMACOLOGICAL RELEVANCE: Ginkgo biloba L. extract (GBE) oral preparations have been used for many years in the prevention and treatment of cardiovascular and cerebrovascular diseases, and the main active ingredients are flavonoids and terpene lactones. Among them, the oral absorption of the prototype components of flavonoid glycosides into the blood needs to be further clarified, and the differences in the oral absorption of different components in GBE by different dosage forms and physiological conditions are not clear yet. AIM OF THE STUDY: To clarify the oral absorption of the prototype flavonoid glycosides in vivo, and to further explore the differences in the oral absorption of various active compounds under different oral dosage forms and dietary conditions. MATERIALS AND METHODS: Firstly, the target compounds were selected based on the characteristic chromatogram of GBE and literature. Then, the content differences of three different oral GBE preparations were studied, and their pharmacokinetics (PK) were compared. Finally, the PK differences of the preparations with better oral absorption under different dietary conditions were studied. RESULTS: Five flavonoid glycosides, three aglycones and four terpene lactones were selected as the research objects. The content determination results of GBE tablets, guttate pills and tinctures showed that the content of several components especially flavonoid glycosides in the tincture was higher than that of the other two preparations. After oral administration of these three preparations, the PK study showed different results from previous studies. The PK behavior of flavonoid glycosides was also determined at the same time as flavonoid glycosides and terpene lactones. and the bioavailability of flavonoid glycosides in the tincture was higher than that of the other two preparations. PK results of fasting and non-fasting showed that taking GBE tincture on an empty stomach increased the absorption of various compounds, especially flavonoid glycosides. However, due to the existence of food residues in the gastrointestinal tract, the oral bioavailability of flavonoid glycosides was significantly improved. CONCLUSIONS: This study discussed the differences in the content and oral absorption of active compounds in different oral preparations of GBE, clarified the in vivo absorption of flavonoid glycosides prototype, as well as the influence of diet on the PK of active compounds, which has certain guiding significance for the clinical application of GBE oral preparations.

Progressive alteration of dynamic functional connectivity patterns in subcortical ischemic vascular cognitive impairment patients.

Alterations in the temporal evolution of brain states in the process of cognitive impairment aggravation due to subcortical ischemic vascular disease (SIVD) is not understood. The dynamic functional connectivity was investigated to identify the abnormal temporal properties of brain states associated with cognitive impairment caused by SIVD. Eighteen patients with subcortical ischemic vascular cognitive impairment with no dementia (SIVCIND), 19 dementia patients (SIVaD) and 26 normal controls were enrolled. We found that the occupancy rate and mean lifetime of brain states were associated with cognitive performance. SIVCIND had a higher occupancy rate and longer mean lifetime in weakly connected states than normal controls. SIVaD had similar but more extensive changes in the temporal properties of brain states. In addition, switching from weakly connected states to more strongly connected states was more difficult in SIVCIND and SIVaD patients than in normal controls, especially in SIVaD patients. The results revealed that not only the transition to but also maintenance in strongly connected states became increasingly difficult when SIVD-related cognitive impairment progressed into a more severe stage.

Covalent Organic Frameworks for Separator Modification of Lithium-Sulfur Batteries.

Lithium-sulfur (Li-S) batteries are regarded as one of the promising energy storage systems. However, rapid capacity attenuation caused by shuttle effect of soluble polysulfides is major challenge in practical application. The separator modification is regarded as one countermeasure besides the construction of sulfur host materials. Covalent organic frameworks (COFs) are one type of outstanding candidates for suppressing shuttle effect of polysulfides. Herein, recent advances of COFs in the application as commercial separator modifiers are summarized. COFs serve as ionic sieves, the importance of porous size and surface environments in inhibiting soluble polysulfides shuttling and promoting lithium ions conduction is highlighted. The superiority of charge-neutral COFs, ionic COFs, and the composites of COFs with conductive materials for improving reversible capacity and cycling stability is demonstrated. Some new strategies for the design of COF-based separator modifiers are proposed to achieving high energy density. The review provides new perspectives for future development of high-performance Li-S batteries.

Chemical and pharmacological difference between honey-fried licorice and fried licorice.

ETHNOPHARMACOLOGICAL RELEVANCE: According to textual research of books from ancient times till now, there are three main preparation methods of "fried licorice", including frying licorice without excipients (F), frying licorice after dipping with water (W), and frying licorice with honey (H). However, with the development over many successive generations, honey frying has gradually become the main processing form of licorice, whereas the fried licorice is nowadays rarely used. AIM OF THE STUDY: The objectives of this study were to clarify the differences of the three forms of "fried licorice" in chemical composition and pharmacological activities, and to screen quality markers for differently processed licorice. It is expected to provide a scientific basis for the rational choice of "fried licorice" as medicine. MATERIALS AND METHODS: Non-target metabolomic analysis based on UHPLC-QE-Orbitrap-MS was conducted to compare the chemical differences between the differently processed licorice material. Pharmacodynamically, the differences in immunomodulatory activity (including intestinal flora experiment), anti-inflammatory activity, and hepatoprotective activity of the differently processed licorice were evaluated. Furthermore, multivariate statistical analysis was performed to screen potential quality markers of honey-fried licorice. The serum concentration of selected markers was determined by UHPLC-QqQ-MS. RESULTS: Metabolomic analysis showed no difference in the chemical composition of F and W, whereas the chemical composition of H was significantly different from that of F and R. The immunomodulatory activity, anti-acute inflammatory effect, and hepatoprotective effect of licorice were significantly improved after frying with honey; No significant differences were observed between F and H in term of immunomodulatory activity and anti-acute inflammatory effect, whereas, H is better than F in terms of liver-protective activity. The intestinal flora experiment confirmed that H does have immunomodulatory activity, while F may induce an increased abundance of certain pathogenic bacteria in the intestine. Multivariate statistical analysis suggests that the content of liquiritin (2), liquiritigenin (3), isoliquiritin (5), isoliquiritigenin (6) and glycyrrhizic acid (7) plusing glycyrrhetinic acid (8) in H group is closely correlated with its improved effects. CONCLUSIONS: This study provides a scientific rational for the selection of "fried licorice" processing methods. In addition, it provides a scientific basis for the selection of quality markers of differently processed licorice.

Effects of acute high-altitude exposure on working memory: A functional near-infrared spectroscopy study.

INTRODUCTION: Inadequate oxygen availability may lead to impairment of neurocognitive functions. The aim of the present study was to investigate the effect of acute high-altitude exposure on the cerebral hemodynamic response and working memory. METHODS: The same subjects performed working memory exercises with forward and backward digit span tasks both under normal oxygen conditions and in large simulated hypobaric hypoxia chambers, and a series of physiological parameters were evaluated. Functional near-infrared spectroscopy was used to measure cerebral blood flow changes in the dorsolateral prefrontal cortex (DLPFC) during the tasks. RESULTS: Compared with normoxic conditions, under hypoxic conditions, the heart rate and blood pressure increased, blood oxygen saturation decreased significantly, and the forward task had similar accuracy and response time, while the backward task had lower accuracy and longer response time. Neuroimaging analysis showed increased activation in the DLPFC during the forward task and deactivation during the backward task under hypobaric hypoxia conditions. CONCLUSION: Acute high-altitude exposure leads to physiological adaptations. The abnormal hemodynamic responses of the DLPFC to hypoxia at low pressure reveal the disruption of neurocognitive function by acute high-altitude exposure, which compromises complex cognitive functions, and provides a promising application for functional near infrared spectroscopy in the exploration of neural mechanisms in the brain during high-altitude exposure.

Differentiation of three Asparagus species by UHPLC-MS/MS based molecular networking identification and chemical profile analysis.

Asparagi Radix (AR), a traditional Chinese medicine, is the dried roots of Asparagus cochinchinensis (Lour.) Merr. Modern pharmacological studies have shown that AR has various excellent bioactivities, such as antioxidative, antitumor, antibacterial, anti-inflammatory, and hypoglycemic effects. However, the quality control method of AR is incomplete and there are various AR adulterants in markets due to their similar morphological characters. Here, holistic and practical quality evaluation methods were developed to chemically distinguish three common Asparagus species in markets, including Asparagus cochinchinensis (Lour.) Merr., Asparagus officinalis L., and Asparagus lycopodineus (Baker) F.T.Wang & Tang. The chemical constituents of three species were rapidly tentatively annotated using a combination of ultra-high pressure liquid chromatography-linear ion trap-orbitrap high resolution mass spectrometry (UHPLC-LTQ-Orbitrap-MS) and molecular networking (MN). Fifty-six steroidal saponins were annotated, including common and characteristic chemical constituents of the three Asparagus species. Besides, to establish holistic and practical methods to differentiate three Asparagus species, an HPLC-ELSD (evaporative light scattering detector) was applied for fingerprint analysis and content determination of the sum of protoneodioscin and protodioscin of twenty samples. Each Asparagus species showed characteristic chemical profile and AR showed much higher level of the sum of protoneodioscin and protodioscin than that in the others. The above analyses showed that the three Asparagus species mainly contain steroidal saponins and the developed HPLC-ELSD profile of saponin can be used to differentiate them. In conclusion, this study reveals the different chemical constituents of three Asparagus species and provides relatively feasible quality evaluation methods for them which are essential for the rational utilization of these Asparagus species.

WWC proteins mediate LATS1/2 activation by Hippo kinases and imply a tumor suppression strategy.

YAP and TAZ (YAP/TAZ), two major effectors of the Hippo signaling pathway, are frequently activated in human cancers. The activity of YAP/TAZ is strictly repressed upon phosphorylation by LATS1/2 tumor suppressors. However, it is unclear how LATS1/2 are precisely regulated by upstream factors such as Hippo kinases MST1/2. Here, we show that WWC proteins (WWC1/2/3) directly interact with LATS1/2 and SAV1, and SAV1, in turn, brings in MST1/2 to phosphorylate and activate LATS1/2. Hence, WWC1/2/3 play an organizer role in a signaling module that mediates LATS1/2 activation by MST1/2. Moreover, we have defined a minimum protein interaction interface on WWC1/2/3 that is sufficient to activate LATS1/2 in a robust and specific manner. The corresponding minigene, dubbed as SuperHippo, can effectively suppress tumorigenesis in multiple tumor models. Our study has uncovered a molecular mechanism underlying LATS1/2 regulation and provides a strategy for treating diverse malignancies related to Hippo pathway dysregulation.

Synergistic mechanism for the bioactivity fortification of licorice by honey.

ETHNOPHARMACOLOGICAL RELEVANCE: Honey-processed licorice has been used since ancient times. It was recorded that honey-processing has the effect of improving the immunomodulatory efficacy of licorice, which has been confirmed by modern pharmacological studies. However, it is still unknown why honey-processing can enhance the immunomodulatory activity of licorice. Our previous research demonstrated that honey has natural deep eutectic solvent (NADES) characteristics. In this study, we investigated the synergistic effect of honey on licorice to elucidate the possible potentiation of honey-frying on licorice. MATERIALS AND METHODS: Immunological experiments were conducted to investigate whether the honey-processing could enhance the immunomodulatory efficacy of licorice in vivo. Then, the synergistic mechanism of honey and licorice was explored based on cell bioactivity tests, metabolomics analysis, bioavailability test, and Fourier transform-infrared (FT-IR) spectra. RESULTS: Pharmacological experiment verified that honey-processing enhanced the immunomodulatory efficacy of licorice. Moreover, honey increased the total flavonoid and polysaccharide contents in licorice decoction, improved the thermal stability and oral bioavailability of certain pharmacologically active constituents, and augmented their overall immunostimulatory functions. Similar effects of honey were also observed with a honey analogue GFSH, a NADES made of glucose, fructose, and sucrose with certain amount of water. The above effects might be due to multiple molecular interactions between active compounds and sugar molecules of honey. CONCLUSION: These findings indicate that the biological activities of medicinal plants might be fortified by honey due to the synergism between licorice and honey. At the meantime, these findings provide theoretical and empirical basis for potential novel applications of honey or other NADESs at augmenting the health-promoting effects of medicinal plants.

A Ralstonia solanacearum effector targets TGA transcription factors to subvert salicylic acid signaling.

The bacterial pathogen Ralstonia solanacearum causes wilt disease on Arabidopsis thaliana and tomato (Solanum lycopersicum). This pathogen uses type III effectors to inhibit the plant immune system; however, how individual effectors interfere with plant immune responses, including transcriptional reprograming, remain elusive. Here, we show that the type III effector RipAB targets Arabidopsis TGACG SEQUENCE-SPECIFIC BINDING PROTEIN (TGA) transcription factors, the central regulators of plant immune gene regulation, via physical interaction in the nucleus to dampen immune responses. RipAB was required for R. solanacearum virulence on wild-type tomato and Arabidopsis but not Arabidopsis tga1 tga4 and tga2 tga5 tga6 mutants. Stable expression of RipAB in Arabidopsis suppressed the pathogen-associated molecular pattern-triggered reactive oxygen species (ROS) burst and immune gene induction as well as salicylic acid (SA) regulons including RBOHD and RBOHF, responsible for ROS production, all of which were phenocopied by the tga1 tga4 and tga2 tga5 tga6 mutants. We found that TGAs directly activate RBOHD and RBOHF expression and that RipAB inhibits this through interfering with the recruitment of RNA polymerase II. These results suggest that TGAs are the bona fide and major virulence targets of RipAB, which disrupts SA signaling by inhibiting TGA activity to achieve successful infection.

Studies of Mucosal Irritation and Cellular Uptake Mechanisms of Xingnaojing Nanoemulsion

Abstract Xingnaojing (XNJ) injection was used to treat pneumonia and stroke in clinic in China, but with poor patient compliance. Xingnaojing nanoemulsion for intranasal delivery was developed to improve it. This article tried to evaluate the mucosal irritation of Xingnaojing nanoemulsion and investigate cellular uptake mechanism of its encapsulated lipophilic drugs. The toad palate model and rat nasal mucosa model were used to study the nasal ciliotoxicity and nasal mucosal irritation of nanoemulsion to evaluate its safety intranasally. The cellular uptake mechanism was studied by Calu-3 cell model. Coumarin 6 was encapsulated in nanoemulsion and the endocytic pathways were studied by cellular uptake experiments after being treated with different inhibitors. In toad palate model, the cilia movement of Xingnaojing nanoemulsion group last for 467.40 ± 39.02 min, which was obviously longer than deoxycholate group (90.60 ± 15.40 min). Studies on rats showed that the damage caused by nanemulsion is capable of being recovered. Nanoemulsion uptake was reduced obviously when cells were treated with wortmannin, and it also decreased about 13% when the temperature reduced from 37ºC to 4ºC. Mucosal irritation caused by nanoemulsion is low and the damage is recoverable. The cellular uptake of Xingnaojing nanoemulsion is energy-dependent, and macropinocytosis was the most important pathway for cellular uptake.

[Puerarin-Chuanxiong oil submicron emulsion combining medicine and adjuvant].

This study was mainly based on the compatibility of Puerariae Lobatae Radix and Chuanxiong Rhizoma to prepare submicron emulsion and evaluated its physical and pharmaceutical properties. Firstly, pseudo-ternary phase diagrams were drawn by dripping method which took Chuanxiong oil as the oil phase and the area of microemulsion region as the index. On this basis, suitable emulsifier and co-emulsifier were screened for the preparation of Chuanxiong oil submicron emulsion. Then, the formula realizing the largest oil loading was selected. Finally, puerarin substituted part of emulsifier and co-emulsifier to lower their content, so as to form puerarin-Chuanxiong oil submicron emulsion featuring the combination of medicine and adjuvant. Its particle size, zeta potential, centrifugal stability and storage stability were determined, and the in vitro drug release behavior was investigated by dialysis bag method, based on which the quality of the as-prepared submicron emulsion was evaluated comprehensively. The proposed method was proved feasible for the preparation of Chuanxiong oil submicron emulsion, which adopted polyoxyethylene castor oil(EL-40) as the emulsifier and was free from co-emulsifier. The formula of the maximum oil loading was found as Chuanxiong oil∶EL-40∶water 3∶7∶90. Further, puera-rin successfully replaced up to 10% of the emulsifier in submicron emulsion. Eventually, the optimal drug-loading formula was determined as puerarin∶Chuanxiong oil∶EL-40∶water 7∶30∶63∶900. The quality evaluation results of the as-prepared submicron emulsion demonstrated that the average emulsion droplet size was 333.9 nm, the PDI 0.26, and the zeta potential-10.12 mV. The submicron emulsion had a good centrifugal stability and did not present any instable phenomena such as delamination and precipitation during its standing still for 50 days. The evaluation of in vitro drug release behavior indicated that the submicron emulsion was capable of releasing the drug completely. The puerarin-chuanxiong oil submicron emulsion prepared in this study possessed a stable quality and to some extent increased the solubility of puerarin along with a sustained-release effect. This study provided ideas for the clinical application of puerarin.