[Modern research progress in external application of traditional Chinese medicine to acupoints].

Traditional Chinese medicine(TCM) has a long history and abundant experience in external therapy, which marks human wisdom. In the early history of human, people found that fumigation, coating, and sticking of some tree branches and herb stems can help alleviate scabies and remove parasites in productive labor, which indicates the emergence of external therapy. Pathogen usually enters the body through the surface, so external therapy can be used to treat the disease. External therapy is among the major characteristic of surgery of TCM. As one of the external therapies in TCM, external application to acupoints smooths the zang-fu organs through meridians and collaterals, thereby harmonizing yin and yang. This therapy emerged in the early society, formed the Spring and Autumn Period and the Warring States Period, improved in the Song and Ming dynasties, and matured in the Qing dynasty. With the efforts of experts in history, it has had a mature theory. According to modern research, it can avoid the first-pass effect of liver and the gastrointestinal irritation and improve the bioavailability of Chinese medicine. Based on the effect of Chinese medicine and the theory of meridian and collateral, it can stimulate the acupoints, exert regulatory effect on acupoints, and give full play to the efficacy of TCM and the interaction of the two. Thereby, it can regulate qi and blood and balance yin and yang, thus being widely used in the treatment of diseases. In this paper, the use of external application to acupoints, the effect on skin immunity, the regulation of neuro-inflammatory mechanism, the relationship between acupoint application and human circulation network, and the development of its dosage form were summarized through literature review. On this basis, this study is expected to lay a foundation for further research.

[Preparation and intestinal absorption of Panax notoginseng saponins chitosan nanoparticles].

In this experiment, Panax notoginseng saponins chitosan nanoparticles(PNS-NPs) were prepared by self-assembly and their appearance, particle size, encapsulation efficiency, drug loading, polydispersity index(PDI), Zeta potential, and microstructure were characterized. The prepared PNS-NPs were intact in structure, with an average particle size of(209±0.258) nm, encapsulation efficiency of 42.34%±0.28%, a drug loading of 37.63%±0.85%, and a Zeta potential of(39.8±3.122) mV. The intestinal absorption of PNS-NPs in rats was further studied. The established HPLC method of PNS was employed to investigate the effects of pH, perfusion rate, and different drugs(PNS raw materials, Xuesaitong Capsules, and PNS-NPs). The absorption rate constant(K_a) and apparent permeability coefficient(P_(app)) in the duodenum, jejunum, ileum, and colon were calculated and analyzed. As illustrated by the results, the intestinal absorption of PNS-NPs was increased in the perfusion solution at pH 6.8(P<0.05), and perfusion rate had no significant effect on the K_a and P_(app) of PNS-NPs. The intestinal absorption of PNS-NPs was significantly different from that of PNS raw materials and Xuesaitong Capsules(P<0.05), and the intestinal absorption of PNS-NPs was significantly improved.

Coarse-Grained Molecular Dynamic and Experimental Studies on Self-Assembly Behavior of Nonionic F127/HS15 Mixed Micellar Systems.

The self-assembly of a nonionic triblock copolymer (F127) and a nonionic surfactant (HS15) has been investigated due to favorable changes in properties in their mixtures. The effect of the mixing ratio on the self-assembly process and on the structural stability of the mixtures was studied by coarse-grained molecular dynamic simulation (CGMD) and experimental measurements (transmission electron microscopy, dynamic light scattering measurement, drug loading stability analysis, and fluorescence spectroscopy measurement). The CGMD provided the information on self-assembly behavior. The microstructure and micellar stability are affected by different proportions of F127/HS15. Pure HS15 molecules (system I) can rapidly form stable aggregates driven by strong hydrophobic force, including two steps: the formation of seed clusters and the fusion of them. At low F127 ratio (system II), the self-assembly process is dynamic unstable, and a volatile "coil/cluster-like" aggregate is formed under the single "binding" effect. As the ratio of added F127 increase, such as system III, stable "lotus-seedpod-like" aggregates form under the double effects of "binding plus wrapping". Its dynamic equilibrium can be achieved rapidly. The experimental results approved the assumption of "different mixing ratio with different structural stability" and even different loading stability of F127/HS15 systems for drugs with different log P, such as PUE and DTX, which means different loading area for them in the micellar systems at different mixing ratios because of less hydrophobic microdomains with the increase of F127 molecules.

Design, synthesis and biological evaluation of coumarin-based N-hydroxycinnamamide derivatives as novel histone deacetylase inhibitors with anticancer activities.

A series of novel coumarin-based N-hydroxycinnamamide derivatives were designed and synthesized as histone deacetylase (HDAC) inhibitors. Most of the synthesized compounds showed potent HDAC inhibitory activity and significant antiproliferative activity against human cancer cell lines MCF-7, HepG2, HeLa and HCT-116. Among them, compound 14f displayed the most potent HDAC inhibition, especially against HDAC1 with IC50 value of 0.19 µM, which was better than that of SAHA (IC50 = 0.23 µM). It also showed the strongest antiproliferative activity towards HeLa cells and more than 26-fold selectivity for HDAC1 compared with HDAC6. Molecular docking studies revealed the possible binding modes of compound 14f into the two isoforms and provided a reasonable explanation for the selectivity. In addition, compound 14f could inhibit colony formation, upregulate the acetylation level of histone H3, and induce apoptosis and cell cycle arrest at G2/M phase in HeLa cells. Taken together, these results highlighted that compound 14f might be a promising HDAC inhibitor for cancer therapy.

Ocular delivery of cyanidin-3-glycoside in liposomes and its prevention of selenite-induced oxidative stress.

CONTEXT: Cataracts have become the leading cause of blindness around the world, which is mainly mediated by oxidative stress. OBJECTIVE: N-trimethyl chitosan (TMC)-coated liposomes of cyanidin-3-glycoside (C3G) (C3G-TCL) were prepared to attenuate oxidative stress induced by selenite sodium in rats. MATERIALS AND METHODS: C3G-TCL were prepared by reverse-phase evaporation method and then coated with self-synthesized TMC. The physicochemical properties were determined. A gamma-scintigraphy study was employed to evaluate the precorneal elimination of the radioactive preparations. The transcorneal visualization for fluorescence-labeled samples was determined by confocal laser scanning microscopy (CLSM). The in vivo anti-oxidative study using C3G-TCL was carried out in rats with selenite-induced cataracts by topical administration. RESULTS: The sphere-like morphological characterization of the vesicles was confirmed by TEM, with a size of 158.3 ± 2.8 nm and a zeta potential of 31.7 mV. The encapsulation efficiency was (53.7 ± 0.2) % as measured by ultrafiltration. C3G-TCL showed a 3.3-fold increment in precorneal residence time when compared with that of the (99m)Tc-solution. A TMC coating enhanced the transepithelial transport of liposomes to a depth of 40-µm in the cornea. Moreover, C3G-TCL could significantly elevate the activity of superoxide dismutase and catalase in lens and also show a considerable reversal of reduced glutathione activity. The lipid peroxidation in lens was strongly prevented when compared with that of groups treated with uncoated C3G-loaded liposomes. DISCUSSION AND CONCLUSION: The coating material TMC for liposomes helps improve the anti-oxidative effect of C3G in vivo through prolonged residence time on the cornea and improved permeability in the corneal epithelium.

[Intestinal absorption of pulchinenosides from Pulsatilla chinensis in rats].

HPLC-ELSD was applied to explore the absorption mechanism of pulchinenosides (B3, BD, B7, B10, B11) in rats. The experimental results showed that the absorption rate constant, Ka value (B3, BD) and Permeability coefficient, Peff value (B3, B7) displayed significant difference (P <0.05) in various intestinal segments, The Ka value and Peff value of PRS was different from each other with the highest absorption in duodenum (duodenum > jejunum > colon > ileum); The PRS displayed excessive satuation as the concentration increased over 0.05-2.5 g · L(-1). There were no obvious linear correlations between Peff values and concentrations in duodenum (0.6007 ≤ R2 ≤ 0.7727); Ka and Peff value declined when the PRS was perfused with P-glycoprotein promoter digoxin, on the other hand, inclined when perfused with P-glycoprotein inhibitor verapamil with significant difference among PRS B3, BD, B7, B11 (P <0.05). All the above results demonstrated that B3, BD, B7 were greatly influenced by absorption sites, duodenum was the main absorption site; PRS didn't entirely transported in a concentration dependent manner, and the transporter-protein involved the transportation, so the intestinal absorption of the five pulchinenosides was not entirely passive diffusion; and PRS might be the substrates of P-glycoprotein.

Panax Notoginseng Saponins Regulate Angiogenic Cytokines Through the PI3K/AKT/mTOR Signaling Pathway to Promote Fracture Healing in Ovariectomized Rats.

Osteoporotic fractures seriously affect the quality of life of the elderly. Panax notoginseng saponins (PNS) have the potential function of preventing osteoporosis. The Phosphatidylinositol 3-kinase (PI3K)/protein kinase (AKT)/mammalian target of rapamycin (mTOR) pathway is involved in the regulation of osteoporosis and has been proven to be related to VEGF secretion and angiogenesis. Therefore, this study aimed to explore the effects of PNS on ovariectomized rats with osteoporotic fracture through the PI3K/AKT/mTOR pathway and angiogenesis-related factors. Female Sprague-Dawley rats were randomly divided into normal control, fracture model, ovariectomized fracture model, low-dose PNS (100 mg/kg/d), and high-dose PNS (200 mg/kg/d). The ovariectomized rat fracture model was established. In low and high dose groups, PNS was administered intraperitoneally. The vascularization of fracture ends was detected in vitro by micro-CT on the 7th, 14th, and 21st day after modeling, and the area and number of blood vessels in the unit field of vision of the callus healing plane were seen by hematoxylin-eosin staining. The expression levels of PI3K, AKT1, mTOR, hypoxia inducible factor-1; VEGF: vascular endothelial growth factor (HIF-1), VEGF, Ang-1, VEGFR2, and angiopoietin like 2 Gene (ANGPTL2) were determined using Western blotting. In the PNS treatment group, the area of cortical bone increased, the area of callus decreased, and the number and area of blood vessels increased significantly when compared with the ovariectomized fracture model group. PNS regulates the PI3K/AKT/mTOR signaling pathway and promotes the expression of vascular-related cytokines (VEGF, Ang-1, VEGFR2, and ANGPTL2) in osteoporotic fractures. PNS may regulate the expression of vascular-related factors through the PI3K/AKT/mTOR pathway and promote the healing of osteoporotic fractures in ovariectomized rats.

Functional properties and molecular docking of different nanoparticles with ROS-sensitive phenylboronylated chitosan as the carrier.

OBJECTIVE: To prepare chitosan-loaded nanoparticles (NPs) that enhance the oral bioavailability of puerarin (Pur) and render it responsive to reactive oxygen species (ROS). SIGNIFICANCE: This research makes substantial progress towards the theory of intelligent drug delivery, offering a new reference for combining Pur with other natural medicinal active ingredients. METHODS: The acylation reaction between chitosan and ROS-sensitive 3-carboxyphenylboronic acid (PBA) was used to synthesise ROS-sensitive phenylboronylated chitosan (PBACS). Subsequently, PBACS-PBA-Pur-NPs and PBACS-TPP-Pur-NPs were prepared via ion gelation after the addition of PBA and sodium tripolyphosphate(TPP), respectively. The physicochemical and functional properties of both NPs were compared, and their differences were preliminarily studied through molecular docking. RESULTS: Reactive oxygen species-sensitive PBACS was successfully synthesised. Of the two NPs prepared, PBACS-TPP-Pur-NPs had a size of 127.2 ± 0.80 nm, polydispersity index (PDI) of 0.129 ± 0.0008, and an encapsulation rate of 95.75 ± 0.387 %, whereas PBACS-PBA-Pur-NPs had a size of 149.8 ± 0.1414 nm, PDI of 0.389 ± 0.0012, and an encapsulation rate of 91.77 ± 0.279 %. The micromorphology of the PBACS-TPP-Pur-NPs exhibited better physical properties. However, PBACS-PBA-Pur-NPs demonstrated a faster in vitro release and more significant in vitro anti-inflammatory effects. Pharmacokinetically, the AUC0-24, Tmax, and Cmax of PBACS-PBA-Pur-NPs were 3.485, 2.117, and 3.339 times higher, respectively, than those of Pur. The AUC0-24, Tmax, and Cmax of PBACS-TPP-Pur-NPs were 2.41, 1.33, and 2.03 times higher, respectively, than those of Pur. Molecular simulation revealed that the binding energy of PBACS-PBA-Pur -NPs was approximately -4.34 kcal/mol and that of PBACS-TPP-Pur-NPs was even lower, approximately -5.93 kcal/mol, suggesting that the NPs prepared with TPP are more densely packed than those designed with PBA, resulting in slower and reduced drug release. CONCLUSION: The NPs constructed in this study effectively reduced inflammatory factors at the disease site, providing a theoretical and experimental basis for the application of nano drugs in inflammatory disease models. In addition, the molecular docking study of the two NPs offered insights into the relationship between the release and structure of subsequent nano drugs.

Mechanism of oxymatrine in the treatment of cryptosporidiosis through TNF/NF-κB signaling pathway based on network pharmacology and experimental validation.

Cryptosporidiosis is a worldwide zoonotic disease. Oxymatrine, an alkaloid extracted and isolated from the plant bitter ginseng, has been reported to have therapeutic effects on cryptosporidiosis. However, the underlying mechanism of its action remains unclear. In this study, we utilized network pharmacology and experimental validation to investigate the mechanism of oxymatrine in the treatment of cryptosporidiosis. First, the potential targets of drugs and diseases were predicted by TCMSP, Gene Cards, and other databases. Following the intersection of drug-disease targets, the DAVID database was used to implement the enrichment analysis of GO functions and KEGG pathways, and then the network diagram of "intersected target-KEGG" relationship was constructed. Autodock 4.2.6 software was used to carry out the molecular docking of core targets to drug components. Based on the establishment of a mouse model of cryptosporidiosis, the validity of the targets in the TNF/NF-κB signaling pathway was confirmed using Western blot analysis and Quantitative Rea-ltime-PCR. A total of 41 intersectional targets of oxymatrine and Cryptosporidium were generated from the results, and five core targets were screened out by network analysis, including RELA, AKT1, ESR1, TNF, and CASP3. The enrichment analysis showed that oxymatrine could regulate multiple gene targets, mediate TNF, Apoptpsis, IL-17, NF-κB and other signaling pathways. Molecular docking experiments revealed that oxymatrine was tightly bound to core targets with stable conformation. Furthermore, we found through animal experiments that oxymatrine could regulate the mRNA and protein expression of IL-6, NF-κB, and TNF-α in the intestinal tissues of post-infected mice through the TNF/NF-κB signaling pathway. Therefore, it can be concluded that oxymatrine can regulate the inflammatory factors TNF-α, NF-κB, and IL-6 through the TNF/NF-κB signaling pathway for the treatment of cryptosporidiosis. This prediction has also been validated by network pharmacology and animal experiments.

[Rat intestinal absorption trait of peimine and peiminine in Thunberg fritillary bulb extract].

To study the in situ intestinal absorption kinetics and compatibility influence of peimine and peiminine in rats, the absorption of peimine and peiminine in small intestine (duodenum, jejunum and ileum) and colon of rats was investigated using in situ single-pass perfusion method and the drug content was measured by HPLC-ELSD. Perfusion rate, pH, concentration of drug, gender and bile duct ligation can significantly affect the absorption of peimine and peiminine, the Ka, and Papp values in the condition of pH 6.8 and pH 7.4 had significant difference (P<0.01), as drug concentration irlcreased, the absorption parameters of peimine and peiminine decreased, Ka and Papp between low concentrations and middle concentrations was significant difference (P<0.01). Verapamil can not affect Ka and Papp of peimine and peiminine which are in the extract (P> 0.05). Bitter almonds and licorice can significantly reduce the absorption of peimine and peiminine with the usual dose (P<0.01), extracted separately and together had no significant difference on Ka and Papp (P> 0.05). Experimental results show that the absorption features of peimine and peiminine are basically the same, both of them could be absorbed at all segments of the intestine in rats and had no special absorption window, and with significant differences between male and female individuals. The absorption of peimine and peiminine complies with the active transport and facilitated diffusion in the general intestinal segments. Bitter almond and licorice can reduce the intestinal absorption rate ofpeimine and peiminine.

[Protective effect of radix sophorae flavescentis mixture on intestinal mucosa in mice infected with Cryptosporidium parvum].

OBJECTIVE: To investigate the protective effect of radix sophorae flavescentis (RSF) mixture on intestinal mucosa in mice infected with Cryptosporidium parvum. METHODS: Thirty BALB/c male mice were randomly divided into control group, infection group and RSF mixture treatment group. Mice of the posterior two groups were inoculated intragastrically with 1 x 10(5) C. parvum oocysts, immunosuppressed with dexamethasone (5 microg/ml) and gentamycin sulfate (40 microg/ml) in drinking water. At the 8th day post-infection, mice in RSF mixture treatment group were treated with 0.2 ml dose of RSF mixture twice a week (three-day intervals) for three weeks. The mice in infection group and RSF mixture treatment group were monitored for oocyst shedding in fecal pellets every two days after treatment. At 28 days after infection, experimental mice were sacrificed, jejunal tissue was removed for preparation of paraffin-embedded sections. The changes of CD3+, CD4+, CD8+ T lymphocytes and IgA plasmocytes in intestinal mucosa were determined by immunohistochemistry. In addition, jejunum of infected mice and treated mice were collected, and ultrastructural changes were observed under electron microscopy. RESULTS: Compared with infection group, the level of oocyst shedding was obviously lower and the time of the oocyst discharging was significantly shorter in RSF mixture treatment group. The proportion of CD3+, CD4+ T lymphocyte and CD4+/CD8+ T cell ratio in infection group (49.7% +/- 2.4%, 25.7% +/- 2.2%, 1.1 +/- 0.3) were significantly lower than that of treatment group (62.4% +/- 1.4%, 37.5% +/- 3.1%, 1.5 +/- 0.3) and control group (66.5% +/- 1.9%, 40.1% +/- 1.8%, 1.5 +/- 0.2) (P < 0.01). CD8+ T lymphocytes showed no significant difference in each group (P > 0.05). The number of IgA plasmocytes in treatment group (52.7 +/- 3.5) was significantly higher than that of control group (8.3 +/- 2.3) and infection group (33.7 +/- 2.6) (P < 0.01). After administration for three weeks, the damaged C. parvum parasites were seldom seen in mouse jejunum, and lysosomes appeared in large number, RSF mixture treatment improved mitochondrial structure and repaired microvilli. In infection group, mitochondria ridges were significantly broken and microvilli surrounding C. parvum oocysts were shed, resulting in the appearance of crater-like lesions on the surface, the oocyst wall and host cell membrane fused together. CONCLUSION: RSF mixture is effective against Cryptosporidium parvum. The damage of intestinal mucosa in infected mice can be repaired after treatment.

Quercetin modulates the liver metabolic profile in a chronic unpredictable mild stress rat model based on metabolomics technology.

Depression is the most prevalent psychiatric disease, and its pathogenesis is still unclear. Currently, studies on the pathogenesis of depression are mainly focused on the brain. The liver can modulate brain function via the liver-brain axis, indicating that the liver plays an important role in the development of depression. This study aims to explore the protective effect of quercetin against chronic unpredictable mild stress (CUMS)-induced metabolic changes and the corresponding mechanisms in the rat liver based on untargeted metabolomics technology. In this study, 96 male rats were divided into six groups: control, different doses of quercetin (10 mg per kg bw or 50 mg per kg bw), CUMS, and CUMS + different doses of quercetin. After 8 weeks of CUMS modeling, the liver samples were collected for metabolomics analysis. A total of 17 altered metabolites were identified, including D-glutamic acid, S-adenosylmethionine, lithocholylglycine, L-homocystine, prostaglandin PGE2, leukotriene E4, cholic acid, 5-methyltetrahydrofolic acid, taurochenodeoxycholic acid, S-adenosylhomocysteine, deoxycholic acid, folic acid, L-methionine, leukotriene C5, estriol-17-glucuronide, PE, and PC, indicating that methionine metabolism, bile acid metabolism, and phosphatidylcholine biosynthesis are the major pathways involved in CUMS-induced hepatic metabolic disorders. Hepatic methylation damage may play a role in the pathophysiology of depression, as evidenced by the first discovery of the abnormality of hepatic methionine metabolism. Abnormal changes in hepatic bile acids may provide stronger evidence for depression pathogenesis involving the microbiota-gut-brain axis, suggesting that the liver is involved in depression development and may be a treatment target. The quercetin treatment alleviated the CUMS-induced liver metabolism disorder, suggesting that quercetin may protect against depression by regulating liver metabolism.

Untargeted metabolomics reveals the preventive effect of quercetin on nephrotoxicity induced by four organophosphorus pesticide mixtures.

This research aimed to explore the protective effect of quercetin against nephrotoxicity induced by four organophosphate pesticide mixtures (PM) using untargeted metabolomics technology in rat kidneys. Sixty male Wistar rats were randomly divided into six groups: control, low-dose quercetin treated (10 mg/kg bw), high-dose quercetin treated (50 mg/kg bw), PM-treated, and two dosages of quercetin + PM-treated. Metabolomics results showed that 17 differential metabolites were identified in the PM-treated group, and pathway analysis revealed that renal metabolic disorders include purine metabolism, glycerophospholipid metabolism, and vitamin B6 metabolism. When high-dose quercetin and PM-treated were administered to rats concurrently, the intensities of differential metabolites were substantially restored (p < 0.01), suggesting that quercetin can improve renal metabolic disorders caused by organophosphate pesticides (OPs). Mechanistically, quercetin could regulate the purine metabolism disorder and endoplasmic reticulum stress (ERS)-mediated autophagy induced by OPs by inhibiting XOD activity. Moreover, quercetin inhibits PLA2 activity to regulate glycerophospholipid metabolism and it could also exert antioxidant and anti-inflammatory effects to correct vitamin B6 metabolism in rat kidneys. Taken together, the high dose of quercetin (50 mg/kg. bw) has a certain protective effect on OPs-induced nephrotoxicity in rats, which provides a theoretical basis for quercetin against nephrotoxicity caused by OPs.

Quasi-opsonin conjugated lipase-sensitive micelles activate macrophages against facultative intracellular bacterial infection.

Drug resistance caused by facultative intracellular bacteria such as Salmonella typhimurium (S. typhimurium) is still a tough challenge. Bacteria phagocytosed by macrophages have evolved a variety of mechanisms to defend against host attack, and the poor entry of antibiotics into infected macrophages is conducive to the survival of intracellular bacteria. In this report, we prepared a quasi-opsonized chloramphenicol (Chl)-loaded micellar system (B-mLBP-M/Chl) assembled by a bacterial lipase-sensitive polymer with a conjugate of lipopolysaccharide-binding protein (LBP) analog and biotin (B) as a ligand, which could eliminate drug-resistant S. typhimurium with quasi-opsonization via 3 steps: (i) target and release antibiotics on bacteria lipase, (ii) opsonize S. typhimurium to be digested by the macrophage, and (iii) activate the macrophage for fighting. The B-mLBP-M/Chl could target bacterial LPS through mLBP by simulating the N-terminal sequence of native LBP, exhibiting a high ability to target the localized infection site in mice. It could also activate the phagocytosis of macrophages via coupled biotin, cooperating with antibiotics and effectively improving the survival of mice with little pathological damage to tissues. Moreover, compared with native opsonin, B-mLBP does not cause an excessive inflammatory response and could recover homeostasis after exerting the quasi-opsonization by regulating the levels of pro-inflammatory cytokines and anti-inflammatory cytokines. With a universal target site for Gram-negative bacteria and macrophage activation, this B-mLBP-M/Chl could be applied to other bacterial infections in the future. In particular, this analog may also serve as a useful template to design safe artificial opsonin, which could be a ligand for drug delivery systems or prodrugs.

[Effect of Toxoplasma gondii infection on the embryonic neural stem cells in rats].

OBJECTIVE: To investigate the effect of Toxoplasma gondii infection on the proliferation, differentiation and migration of the embryonic neural stem cells (NSCs) in early pregnancy of rat. METHODS: Twelve pregnant Sprague-Dawley rats were randomly divided into control and infection groups. Rats in the infection group were each inoculated intraperitoneally with 1 x 10(5) T. gondii RH strain tachyzoites at day 1 (E1 day). Same amount of physiological saline was intraperitoneally injected for rats in control group. At E5 day, blood samples were taken from caudal vein and Giemsa staining of blood cells was performed to find T. gondii. At E9, E10 and E11 day, two rats in each group per time point were sacrificed and reverse transcription PCR (RT-PCR) was performed to detect B1 gene expression of T. gondii in amniotic fluid to confirm T. gondii infection. NSCs were cultured in vitro. The proliferation level was detected by methyl thiazolyl tetrazolium (MTT) assay. After differentiation culture of NSCs, the immunofluorescence assay was conducted to detect the expression of nestin, microtubule-associated protein 2 (MAP2) and glial fibrillary acidic protein (GFAP) to calculate the ratio of NSCs which differentiated to neurons and astrocytes. The embryonic nerve tissues at E9, E10 and E11 day in each group were taken to make frozen sections. The immunofluorescence assay was carried out to detect the expression of neuronal cell adhesion molecule (NCAM) in the nerve tissues at different developmental stages. RESULTS: Both the results of blood smears and RT-PCR confirmed that the pregnant rats and embryos were all infected by T. gondii in infection group. The morphology of the cultured NSCs under microscope was consistent with the characteristics of the normal NSCs. In addition, the NSC biomarker nestin protein was stained positive. The MTT assay showed that the proliferation level was lower in infection group than that of the control, and statistical differences were found between the two groups at day 3 and 4 after passages (P < 0.05). The immunofluorescence staining of MAP2 and GFAP showed that the percentage of neuron differentiation was 15.15% (55/363) in control group and 8.73% (31/355) in infection group, respectively, with a statistical difference (P < 0.05), and the percentage of astrocyte differentiation was 53.35% (199/374) and 67.48% 249/369), respectively (P > 0.05). In both groups, NCAM protein was found expressed at E9, E10 and E11 day in embryo nerve tissues. The fluorescence became stronger with time. The expression level in control group was significantly higher than that in infection group (P < 0.01). CONCLUSION: T. gondii infection at early gestation may inhibit the proliferation, differentiation and migration of neural stem cells in rats.

Effect of pectoralis minor relaxation on the prognosis of rotator cuff injury under arthroscopy.

BACKGROUND: Rotator cuff injury is the main cause of shoulder joint pain. Rotator cuff tear is a serious stage of rotator cuff injury. The purpose of this study was to observe the effect of pectoralis minor relaxation on the prognosis of rotator cuff injury under arthroscopy. METHODS: The clinical data of patients with rotator cuff injury admitted to our department from March 2019 to August 2020 were retrospectively analyzed. These patients were divided into a conventional arthroscopy surgery group and a pectoralis minor relaxation group. The American Shoulder and Elbow Surgeons' Form (ASES) and University of California at Los Angeles Shoulder Scores (UCLASS) were used to assess shoulder joint function during the postoperative and follow-up periods, and the visual analogue scale (VAS) was used to assess shoulder joint pain. RESULTS: Shoulder joint function of the two groups of patients was significantly improved at 1 week postoperatively compared with that before surgery (P<0.05). The shoulder joint function of the pectoralis minor relaxation group was better than that of the conventional arthroscopy surgery group (P<0.05). Also, the shoulder joint function of patients in pectoralis minor relaxation group was better than that in conventional arthroscopy surgery group at 6 weeks, 12 weeks and 6 months after surgery (P<0.05). The UCLASS evaluation results were consistent with the ASES results. At 1 week after surgery, the pain of the two groups of patients was significantly less than before the operation, but the degree of pain relief was greater at 6 weeks postoperatively. Starting at 6 weeks after surgery, the shoulder joint pain of the pectoralis minor relaxation group was less than that of conventional arthroscopy surgery group (P<0.05). The healing of the incision was observed after the operation. No incision infection or exudation was found in the two groups of patients. CONCLUSIONS: The addition of pectoralis minor relaxation on the basis of conventional arthroscopic surgery is beneficial to the further recovery of shoulder joint function and pain reduction in patients with rotator cuff injury.

Metabolomics analysis of the effects of quercetin on Cd-induced hepatotoxicityin rats.

Cadmium (Cd) is known to cause damage to the liver. In this study, metabolomics technology was used to investigate the effect of quercetin (QE) on Cd-induced hepatotoxicity. A total of 60 male SD rats were randomly divided into the following six groups: control group (C), low and high-dose QE group (Q1: 10 mg/kg·bw, Q2: 50 mg/kg·bw), Cd group (D), low and high-dose QE and Cd combined intervention group (DQ1, DQ2). The rats were given Cd chloride (CdCl2) at a concentration of 40 mg/L through free drinking water. After 12 weeks of treatment, liver samples of rats were collected for metabonlomic analysis. A total of 12 metabolites were identified, the intensities of PC (18:0/14:1(9Z)) and arachidonate acid were decreased in the Cd-treated group (p < 0.01), whereas the intensities of chenodeoxyglycocholic acid, cholic acid, taurochenodesoxycholic acid, glycocholic acid, prostaglandin D2, 15-deoxy-d-12,14-PGJ2, oxidized glutathione, cholesterol, protoporphyrin IX, bilirubin were increased significantly in the Cd-treated group compared with group C (p < 0.01). When rats were given high doses of QE and Cd at the same time, the intensity of the above metabolites was significantly restored in group DQ2. Results suggest that the protective effect of QE on Cd-induced liver injury is associated with antioxidant activity of QE, as well as QE can regulates hepatic bile acid metabolism by affecting FXR and BSEP, and regulates AA metabolism by inhibiting Cd-induced activities of COX-2 and PLA2.

Nose-to-brain delivery of drug nanocrystals by using Ca2+ responsive deacetylated gellan gum based in situ-nanogel.

The objective of this study is to use a carbohydrate polymer deacetylated gellan gum (DGG) as matrix to design nanocrystals based intranasal in situ gel (IG) for nose-to -brain delivery of drug. The harmine nanocrystals (HAR-NC) as model drug were prepared by coupling homogenization and spray-drying technology. The HAR-NC was redispersed in the (DGG) solutions and formed the ionic-triggered harmine nanocrystals based in situ gel (HAR-NC-IG). The crystal state of HAR remained unchanged during the homogenization and spray-drying. And the HAR-NC-IG with 0.5% DGG exhibited excellent in situ-gelation ability, water retention property and in vitro release behavior. The bioavailability in brain of intranasal HAR-NC-IG were 25-fold higher than that of oral HAR-NC, which could be attributed to nanosizing effect of HAR-NC and bioadhesive property of DGG triggered by nasal fluid. And the HAR-NC-IG could significantly inhibit the expression of acetylcholinesterase (AchE) and increase the content of acetylcholin (ACh) in brain compared with those of reference formulations (p < 0.01). The DGG based nanocrystals-in situ gel was a promising carrier for nose-to-brain delivery of poorly soluble drug, which could prolong the residence time and improve the bioavailability of poorly soluble drugs in brain.

Downregulation of DNMT3a expression by RNAi and its effect on NF-κBs expression of thymic epithelial cells.

OBJECTIVE: To understand the characteristics of DNA methyltransferase 3a (DNMT3a) in thymoma associated Myasthenia Gravis reveal its transcriptional regulator network as while as analyze the effect of DNMT3a on Rel/ nuclear factor-kappaB family (RelA/RelB) and its downstream autoimmune regulatory factor (Aire). METHODS: Tissues of 30 patients with thymoma, with or without myasthenia gravis (MG), were collected and the DNMT3a protein expression were evaluated through immunohistochemistry. We performed mRNA expression profiling microarray detection and analysis, and integrated the analysis by constructing protein-protein interaction networks and the integration with other database. We identified molecular difference between low and high DNMT3a in the thymoma by heatmap. We also performed PCR validation in thymoma tissues. The DNMT3a-shRNA plasmid was transfected into TEC cells, and these cells were treated with 5-aza-2-deoxycytidine, a blocker of DNMT3a. After the down-regulation of DNMT3a in TEC cells, the transcript and protein levels of RelA, RelB, Aire, and CHRNA3 were evaluated by western blotting. In addition, changes in gene expression profiles were screened through microarray technology. We performed differential gene analysis in the thymoma cohort by heatmap with R (v.4.3.0) software. RESULTS: In 30 matched tissue specimens, the expression of DNMT3a protein in thymoma with MG was lower than that in thymoma. Through mRNA expression profiling analysis, we constructed a co-expression network of DNMT3a and found direct interaction between IKZF1 and DNMT3a, and this co-expression relationship was overlappted with Cistrome DB database. We found up-regulation of 149 mRNAs and repression of 177 mRNAs in thymoma with MG compared with thymoma. Gene ontology and pathway analysis show the involvement of a multitude of genes in the mis-regulation of MG-related pathways. RNA interference significantly reduced the level of mRNA of DNMT3a, which proved that plasmid DNMT3a was effective. In comparison to the control group, the levels of DNMT3a, Aire, and CHRNA3 mRNA and protein in TEC cells transfected with DNMT3a-shRNA interference plasmid were significantly decreased, while the expression level of RelA and RelA/RelB was significantly increased. CONCLUSIONS: Our study reveals the DNMT3a-NF-κB pathway has a major effect on MG, and can be used as a marker for diagnosis as well as a target for MG treatment.

Changes in expression of serum chemokine CXCL13 and IL-6 after hip replacement, and the relationship with lower limb vein thrombus.

Changes in the expression of serum chemokine CXC ligand 13 (CXCL13) and interleukin-6 (IL-6), and the relationship with lower limb vein thrombus were explored. A total of 128 patients undergoing hip replacement in The First Affiliated Hospital of Guizhou University of Traditional Chinese Medicine from May 2017 to June 2019 were selected, and the patients suffering from lower limb vein thrombus were enrolled as group A and other patients not suffering from it were enrolled as group B. Enzyme-linked immuno-sorbent assay was employed to determine the levels of serum chemokine CXCL13 and IL-6, and receiver operating characteristic curves of serum chemokine CXCL13 and IL-6 levels in diagnosing restenosis after surgery were drawn. Pearson's correlation coefficient was adopted to analyze the correlation between serum chemokine CXCL13 and IL-6, and the logistic regression analysis to analyze the risk factors affecting hip replacement in patients. The levels of serum CXCL13 and IL-6 in group A were significantly higher than those in group B (both P<0.001). The specificity and sensitivity of serum CXCL13 level in diagnosis of lower limb vein thrombus after hip replacement were 61.76 and 80.00%, respectively, and those of serum IL-6 level in diagnosis were 70.59 and 66.67%, respectively. Serum CXCL13 level was positively correlated with serum IL-6 level (P<0.001), and age, body mass index (BMI), CXCL13 level and IL-6 level of the patients were independent risk factors affecting the efficacy of hip replacement. Serum CXCL13 level and serum IL-6 level can be used as biological indexes for prediction of early lower limb vein thrombus after hip replacement, and logistic regression analysis revealed that the age of the patients, BMI, diabetes history, hyperlipidemia history, hypertension history, CXCL13 level and IL-6 level are independent risk factors affecting the efficacy of hip replacement.