Kidney Yang Deficiency Syndrome Exacerbates Aß25-35-Induced Pathological Changes, and Ginsenoside Re Ameliorates Synapse Lesions in Aß25-35- Injected Rats with Kidney Yang Deficiency Syndrome.

BACKGROUND: Traditional Chinese medicine (TCM) indicates that Alzheimer's disease (AD) is considered the consequence produced by Kidney Yang Deficiency Syndrome (KDS-Yang), which has similar clinical characteristics to glucocorticoid withdrawal syndrome. Ginsenoside Re (G-Re) has been found to ameliorate the symptoms and pathological impairments of AD. However, it's not clear whether G-Re could protect memory and synapse lesions against kidney deficiency dementia. METHODS: Subcutaneous injection of hydrocortisone for 14 days was used to produce KDS-Yang. On the 15th day, Aß25-35 peptide was injected into the intracerebroventricular (icv) of KDS-Yang rats. Spine density was analyzed by Golgi staining and the ultrastructural morphology of the synapse was detected using Transmission Electron Microscopy (TEM). Western blot was used to examine the expression of pS396, pS404, Tau-5, tGSK-3ß, pS9GSK-3ß, Syt, Syn I, GluA1, GluN2B, PSD93, PSD95, ß2-AR and pS346-b2-AR. RESULTS: Hyperphosphorylation of tau in Aß25-35-injected rats with KDS-Yang was stronger than in Aß25-35-injected rats at the sites of Ser396 and Ser404. G-Re improved spatial memory damage detected by Morris water-maze (MWM), enhanced spines density, the thickness of postsynaptic density (PSD) and increased the expression of Syt, Syn I, GluA1, GluN2B, PSD93 and PSD95. Moreover, GRe decreased the hyperphosphorylation of ß2-AR at serine 346 in Aß25-35-injected rats with KDS-Yang. CONCLUSION: KDS-Yang might exacerbate AD pathological lesions. Importantly, G-Re is a potential ingredient for protecting against memory and synapse deficits in kidney deficiency dementia.

High-quality Fagopyrum esculentum genome provides insights into the flavonoid accumulation among different tissues and self-incompatibility.

Common buckwheat (Fagopyrum esculentum) and Tartary buckwheat (Fagopyrum tataricum), the two most widely cultivated buckwheat species, differ greatly in flavonoid content and reproductive mode. Here, we report the first high-quality and chromosome-level genome assembly of common buckwheat with 1.2 Gb. Comparative genomic analysis revealed that common buckwheat underwent a burst of long terminal repeat retrotransposons insertion accompanied by numerous large chromosome rearrangements after divergence from Tartary buckwheat. Moreover, multiple gene families involved in stress tolerance and flavonoid biosynthesis such as multidrug and toxic compound extrusion (MATE) and chalcone synthase (CHS) underwent significant expansion in buckwheat, especially in common buckwheat. Integrated multi-omics analysis identified high expression of catechin biosynthesis-related genes in flower and seed in common buckwheat and high expression of rutin biosynthesis-related genes in seed in Tartary buckwheat as being important for the differences in flavonoid type and content between these buckwheat species. We also identified a candidate key rutin-degrading enzyme gene (Ft8.2377) that was highly expressed in Tartary buckwheat seed. In addition, we identified a haplotype-resolved candidate locus containing many genes reportedly associated with the development of flower and pollen, which was potentially related to self-incompatibility in common buckwheat. Our study provides important resources facilitating future functional genomics-related research of flavonoid biosynthesis and self-incompatibility in buckwheat.

Qing-Xin-Jie-Yu Granule inhibits ferroptosis and stabilizes atherosclerotic plaques by regulating the GPX4/xCT signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Qing-Xin-Jie-Yu Granule (QXJYG) is an integrated traditional Chinese medicine formula used to treat atherosclerotic (AS) cardiovascular diseases. A randomized controlled trial found that QXJYG reduced cardiovascular events and experiments also verified that QXJYG attenuated AS by remodeling the intestinal flora. AIM OF THE STUDY: To determine whether QXJYG would attenuate AS and plaque vulnerability by regulating ferroptosis in high-fat diet-induced atherosclerotic ApoE-/- mice and to investigate the effects of QXJYG on macrophage ferroptosis in RAS-selective lethal 3 (RSL3)-induced J744A.1 cells. METHODS: AS models in ApoE-/- mice and RSL3-induced ferroptosis in J744A.1 cells were established to measure the protective and anti-ferroptotic effects of QXJYG in vivo and in vitro. The glutathione peroxidase 4 (GPX4)/cystine glutamate reverse transporter (xCT) signal pathway was examined by immunohistochemistry and western blotting. RESULTS: QXJYG attenuated AS progression and plaque vulnerability. Characteristic morphological changes of ferroptosis in the QXJYG-treated animals were rare. Total iron was significantly lower in the QXJYG group than in the model group (P < 0.05); QXJYG suppressed the lipid peroxidation (LPO) levels (malondialdehyde), enhanced the antioxidant capacity (superoxide dismutase and glutathione), and reduced inflammatory factors (interleukin [IL]-6, IL-1ß, tumor necrosis factor-α) associated with ferroptosis. Expression of GPX4/xCT in aorta tissues was remarkably increased in the QXJYG group. QXJYG inhibited ferroptosis in J744A.1 macrophages disturbed using RSL3. The Fe2+, LPO, and reactive oxygen species levels were lower in the QXJYG group than in the RSL3 group (P < 0.05). The QXJYG group showed higher expression of the GPX4/xCT signal pathway. CONCLUSION: QXJYG inhibits ferroptosis in vulnerable AS plaques partially via the GPX4/xCT signaling pathway.

Characterizing thalamic and basal ganglia nuclei in medically intractable focal epilepsy by MR fingerprinting.

OBJECTIVES: Magnetic resonance fingerprinting (MRF) is a novel, quantitative, and noninvasive technique to measure brain tissue properties. We aim to use MRF for characterizing normal-appearing thalamic and basal ganglia nuclei in the epileptic brain. METHODS: A three-dimensional (3D) MRF protocol (1 mm3 isotropic resolution) was acquired from 48 patients with unilateral medically intractable focal epilepsy and 39 healthy controls (HCs). Whole-brain T1 and T2 maps (containing T1 and T2 relaxation times) were reconstructed for each subject. Ten subcortical nuclei in the thalamus and basal ganglia were segmented as regions of interest (ROIs), within which the mean T1 and T2 values, as well as their coefficient of variation (CV) were compared between the patients and HCs at the group level. Subgroup and correlation analyses were performed to examine the relationship between significant MRF measures and various clinical characteristics. Using significantly abnormal MRF measures from the group-level analyses, support vector machine (SVM) and logistic regression machine learning models were built and tested with 5-fold and 10-fold cross-validations, to separate patients from HCs, and to separate patients with left-sided and right-sided epilepsy, at the individual level. RESULTS: MRF revealed increased T1 mean value in the ipsilateral thalamus and nucleus accumbens; increased T1 CV in the bilateral thalamus, bilateral pallidum, and ipsilateral caudate; and increased T2 CV in the ipsilateral thalamus in patients compared to HCs (p < .05, false discovery rate [FDR] corrected). The SVM classifier produced 78.2% average accuracy to separate individual patients from HCs, with an area under the curve (AUC) of 0.83. The logistic regression classifier produced 67.4% average accuracy to separate patients with left-sided and right-sided epilepsy, with an AUC of 0.72. SIGNIFICANCE: MRF revealed bilateral tissue-property changes in the normal-appearing thalamus and basal ganglia, with ipsilateral predominance and thalamic preference, suggesting subcortical involvement/impairment in patients with medically intractable focal epilepsy. The individual-level performance of the MRF-based machine-learning models suggests potential opportunities for predicting lateralization.

Anti-hyperuricemic bioactivity of Alstonia scholaris and its bioactive triterpenoids in vivo and in vitro.

ETHNOPHARMACOLOGY RELEVANCE: One folk use of Alstonia scholaris (L.) R. Br. in "Dai" ethno-medicine system is to treat gouty arthritis, which might be caused by hyperuricemia, but anti-hyperuricemic investigation of A. scholaris were rarely reported. AIM OF THE STUDY: To verify anti-hyperuricemic property of A. scholaris, and explore its bioactive compounds in vivo and in vitro. MATERIALS AND METHODS: The anti-hyperuricemic bioactivity of the non-alkaloids fraction and compounds were evaluated with potassium oxonate (PO) induced hyperuricemia mice model in vivo, and monosodium urate (MSU) induced human renal tubular epithelial cells (HK-2) was selected to test in vitro, respectively, with benzobromarone as the positive control. 11 triterpenoids were isolated by phytochemical methods and their structures were elucidated by spectroscopic analysis and ECD calculation. RESULTS: The non-alkaloids fraction of A. scholaris decreased the serum uric acid (UA) level in mice model significantly at the doses of 100 mg/kg and 200 mg/kg, and then nine ursane- and two oleanane-triterpenoids including four new compounds (1-3 and 10) were isolated from the bioactive fraction, in which compounds 1, 4, 5, 6 and 10 exhibited better anti-hyperuricemic tendency in vitro by promoting the excretion of UA in MSU-induced HK-2 cell model at a concentration of 5 µM. Furthermore, compounds 1 and 4 were proved to reduce the serum UA level in mice significantly at 5 mg/kg in vivo. CONCLUSIONS: The results supported the traditional use of A. scholaris in treating gouty arthritis, and also provided new bioactive triterpenoids for further chemical and pharmacological investigation.

Two new dammarane-type triterpenoids from the green walnut husks of Juglans mandshurica Maxim.

Two new dammarane-type triterpenoids, dammar-3α,12(R),20(S)-triol-12,32(R);20,32-diepoxy-25-methy-25-en-tridecacyclic ether (1) and (23E)-12ß,20(R),25(S),26-tetrahydroxydammar-23-en-3-one (2) were isolated from the green walnut husks of Juglans mandshurica Maxim together with six known compounds. Their structures were elucidated through extensive spectroscopic analyses and by comparison with the literature, and the cytotoxic activities of these compounds were evaluated.

Efficacy of Sijunzi decoction on limb weakness in spleen Qi deficiency model rats through adenosine monophosphate-activated protein kinase/unc-51 like autophagy activating kinase 1 signaling.

OBJECTIVE: To investigate the efficacy of Sijunzi decoction () on limb weakness in a rat model of spleen Qi deficiency (SQD), and to study its effect on mitophagy in skeletal muscle through adenosine monophosphate-activated protein kinase (AMPK) / unc-51 like autophagy activating kinase 1 (ULK1) signaling. METHODS: SQD model rats were produced by fasting combined with forced swimming method for 15 d. After model assessment, rats were randomly divided into four groups of 10 [low/middle/high (L/M/H) Sijunzi decoction dose groups and a normal saline (S) group]. Limb holding power (HP) and body mass (BM) were measured after 2 weeks of treatment. Following euthanasia, quadriceps femoris were dissected and myofiber and mitochondrial morphology were observed by transmission electron microscopy (TEM). Mitochondrial membrane potential (MMP), adenosine triphosphatase (ATP) and reactive oxygen species (ROS) levels were determined using colorimetric methods, and immunoblot analysis of Microtubule-associated protein light chain 3 (LC3) and Sequestosome 1 (p62) was performed to monitor mitophagy and AMPK/ULK1 signaling. RESULTS: Compared with control (C) group rats, in the S group, HP was reduced, the myofiber Z line was disordered, mitochondria were scattered, and numerous vacuoles and mitophagy were observed. MMP and ATP levels were reduced, ROS levels were elevated, and LC3B expression, and p-AMPKα (Thr172)/AMPKα, p-ULK1 (Ser555)/ULK1, and p-Raptor (Ser792)/Raptor ratios were increased, while p62 expression and p-mTOR (Ser2448)/mTOR and p-ULK1 (Ser757)/ULK1 ratios were decreased. After treatment, compared with the S group, HP was improved in M and H groups but not in the L group. Mitophagy was reduced in M, H and L groups but the Z line was disordered and vacuolization remained in the L group. ATP levels were elevated in M, H and L groups, and MMPs were elevat-ed in M and H groups but not in the L group. ROS levels were decreased in M, H and L groups, as were LC3B expression and p-Raptor (Ser792)/Raptor ratios, while p62 expression and p-mTOR (Ser2448)/mTOR and p-ULK1 (Ser757)/ULK1 ratios were increased in M and H groups but not in the L group. p-AMPKα (Thr172)/AMPKα and p-ULK1 (Ser555)/ULK1 ratios were decreased in M, H and L groups. CONCLUSIONS: Sijunzi decoction improved HP, possibly by inhibiting mitophagy via suppression of AMPK/ULK1 signaling. This restored mitochondrial morphology and improved oxidative phosphorylation, which contributed to recovery of limb weakness in SQD model rats.

Human umbilical cord mesenchymal stem cell-derived small extracellular vesicles ameliorate collagen-induced arthritis via immunomodulatory T lymphocytes.

BACKGROUND: Rheumatoid arthritis (RA) is an autoimmune disease for which there are currently no effective therapies. Although mesenchymal stem cells (MSCs) can prevent arthritis through immunomodulatory mechanisms, there are several associated risks. Alternatively, MSC-derived small extracellular vesicles (sEVs) can mimic the effects of MSCs, while reducing the risk of adverse events. However, few studies have examined sEVs in the context of RA. Here, we evaluate the immunomodulatory effects of human umbilical cord MSC (hUCMSC)-derived sEVs on T lymphocytes in a collagen-induced arthritis (CIA) rat model to elucidate the possible mechanism of sEVs in RA treatment. We then compare these mechanisms to those of MSCs and methotrexate (MTX). METHODS: The arthritis index and synovial pathology were assessed. T lymphocyte proliferation and apoptosis, Th17 and Treg proportions, and interleukin (IL)-17, IL-10, and transforming growth factor (TGF)-ß expression were detected using flow cytometry. Retinoic acid receptor-related orphan receptor gamma t (RORγt) and forkhead box P3 (FOXP3), which are master transcriptional regulators of Th17 and Treg differentiation, were also assessed using immunohistochemistry and reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: sEV treatment ameliorated arthritis and inhibited synovial hyperplasia in a dose-dependent manner. These effects were mediated by inhibiting T lymphocyte proliferation and promoting their apoptosis, while decreasing Th17 cell proportion and increasing that of Treg cells in the spleen, resulting in decreased serum IL-17, and enhanced IL-10 and TGF-ß expression. Transcriptionally, sEVs decreased RORγt and increased FOXP3 expression in the spleen, and decreased RORγt and FOXP3 expression in the joints. In some aspects sEVs were more effective than MSCs and MTX in treating CIA. CONCLUSIONS: hUCMSC-derived sEVs ameliorate CIA via immunomodulatory T lymphocytes, and might serve as a new therapy for RA.

Genome-Wide Identification of CsATGs in Tea Plant and the Involvement of CsATG8e in Nitrogen Utilization.

Nitrogen (N) is a macroelement with an indispensable role in the growth and development of plants, and tea plant (Camellia sinensis) is an evergreen perennial woody species with young shoots for harvest. During senescence or upon N stress, autophagy has been shown to be induced in leaves, involving a variety of autophagy-related genes (ATGs), which have not been characterized in tea plant yet. In this study, a genome-wide survey in tea plant genome identified a total of 80 Camellia Sinensis autophagy-related genes, CsATGs. The expression of CsATG8s in the tea plant showed an obvious increase from S1 (stage 1) to S4 (stage 4), especially for CsATG8e. The expression levels of AtATGs (Arabidopsis thaliana) and genes involved in N transport and assimilation were greatly improved in CsATG8e-overexpressed Arabidopsis. Compared with wild type, the overexpression plants showed earlier bolting, an increase in amino N content, as well as a decrease in biomass and the levels of N, phosphorus and potassium. However, the N level was found significantly higher in APER (aerial part excluding rosette) in the overexpression plants relative to wild type. All these results demonstrated a convincing function of CsATG8e in N remobilization and plant development, indicating CsATG8e as a potential gene for modifying plant nutrient utilization.

The Effect of Vitamin D Supplementation on Attention-Deficit/Hyperactivity Disorder: A Systematic Review and Meta-Analysis of Randomized Controlled Trials.

Objective: A systematic review and meta-analysis of randomized controlled trials (RCTs) were conducted to assess the benefits and harms of vitamin D supplementation for attention-deficit/hyperactivity disorder (ADHD) patients. Methods: We followed the standard methodological procedures of the Cochrane Handbook for Systematic Reviews of Intervention. PubMed, Embase, the Cochrane Central Register of Controlled Trials, Science and Conference Proceedings Citation Index-Social Science and Humanities (Web of Science), ClincalTrials.gov, and World Health Organization's International Clinical Trials Registry Platform were searched for RCTs in January 2019. Independently, two authors (J.G., T.X.) extracted data, assessed the risk of bias, combined the data, and graded evidence quality using the Grading of Recommendations Assessment, Development, and Evaluation approach. Our primary outcomes were assessed through rating scales of ADHD severity. Secondary outcomes measured were the possible adverse effects of vitamin D supplementation and vitamin D status after supplementation for ADHD. Results: We included four RCTs with 256 children addressing vitamin D supplementation as adjunctive therapy to methylphenidate on ADHD symptoms. Vitamin D supplementation demonstrated a small but statistically significant improvement in ADHD total scores, inattention scores, hyperactivity scores, and behavior scores. The improvement was likely limited due to the low to very low quality of evidence in the literature. There was no statistically significant improvement in oppositional scores. Reported adverse events in the vitamin D group were mild and not significantly different from the control group. Vitamin D supplementation increased serum vitamin D levels and the ratio of patients with sufficient vitamin D levels. Conclusions: Vitamin D supplementation as adjunctive therapy to methylphenidate appeared to reduce ADHD symptoms without serious adverse events, associated with improved vitamin D status. However, considering the generally low strength of evidence, well-designed RCTs are needed to determine the efficacy and safety of vitamin D supplementation for both children and adults with ADHD, especially in the setting of a combination of vitamin D and other ADHD treatments.

KIFC1 is essential for acrosome formation and nuclear shaping during spermiogenesis in the lobster Procambarus clarkii.

In order to study the function of kinesin-14 motor protein KIFC1 during spermatogenesis of Procambarus clarkii, the full length of kifc1 was cloned from testes cDNA using Rapid-Amplification of cDNA Ends (RACE). The deduced KIFC1 protein sequence showed the highest similarity between Procambarus clarkii and Eriocheir senensis (similarity rate as 64%). According to the results of in situ hybridization (ISH), the kifc1 mRNA was gathered in the acrosome location above nucleus in the mid- and late-stage spermatids. Immunofluorescence results were partly consistent with the ISH in middle spermatids, while in the late spermatids the KIFC1 was distributed around the nucleus which had large deformation and formed four to six nuclear arms. In the mature sperm, KIFC1 and microtubules were distributed around the sperm, playing a role in maintaining the sperm morphology and normal function. Overexpression of P. clarkii kifc1 in GC1 cells for 24 hours resulted in disorganization of microtubules which changed the cell morphology from circular and spherical into fusiform. In addition, the overexpression also resulted in triple centrosomes during mitosis which eventually led to cell apoptosis. RNAi experiments showed that decreased KIFC1 protein levels resulted in total inhibition of spermatogenesis, with only mature sperm found in the RNAi-testis, implying an indispensable role of KIFC1 during P. clarkii spermiogenesis.

[Effects of Rhubarbs from different regions on blood lipid and antioxidation of hyperlipidemia rats].

OBJECTIVE: To comparatively study the effects of Rhubarbs from different regions on blood lipid and antioxi dation of hyperlipidemia rats. METHODS: Male rats were randomly divided into 9 groups ( n = 8) and fed with high-fat diet to replicate the hyperlipidemia model. Meanwhile, Rheum tanguticum was administrated intragastrically at two doses (3.0 g/kg and 1.0 g/kg), once a day for continuous 28 days. The effects of Rheum tanguticum planted in Gannan (RT-GN), Rheum tanguticum planted in Xinin (RT-XN) and Rheum plmatum planted in Lixian (RP-LX) were evaluated through detecting the parameters of blood lipids, blood viscosity and antioxidant system. RESULTS: T-GN, RT-XN and RP-LX in the range of 1.0-3.0 g/kg could decrease the blood levels of total cholesterol (TC), triglyceride (TG), low density lipoprotein (LDL) and malonaldehyde (MDA) in blood. Besides, they could reduce blood viscosity, increase high density lipoprotein (HDL) level and upregulate the activity of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px). Interestingly, their effects on blood viscosity was obviously in a dose dependent manner. In addition, the effects of RT-GN on LDL, MDA and blood viscosity were not significantly different from those of RT-XN and better than those of RP-LX. CONCLUSION: The RT has better hypolipidemic effects than the RP, but RT-GN and RT-XN are not different from the above effects.

Repellent and Contact Toxicity of Alpinia officinarum Rhizome Extract against Lasioderma serricorne Adults.

The repellent and contact toxicities of Alpinia officinarum rhizome extract on Lasioderma serricorne adults, and its ability to protect stored wheat flour from L. serricorne adults infestation were investigated. The A. officinarum extract exhibited strong repellent and contact toxicities against L. serricorne adults. The toxicities enhanced significantly with the increasing treatment time and treatment dose. The mean percentage repellency value reached 91.3% at class V at the dose of 0.20 µL/cm2 after 48 h of exposure. The corrected mortality reached over 80.0% at the dose of 0.16 µL/cm2 after 48 h of exposure. The A. officinarum extract could significantly reduce L. serricorne infestation level against stored wheat flour. Particularly, the insect infestation was nil in wheat flour packaged with kraft paper bags coated with the A. officinarum extract at the dose of above 0.05 µL/cm2. The naturally occurring A. officinarum extract could be useful for integrated management of L. serricorne.

A novel algal biofilm membrane photobioreactor for attached microalgae growth and nutrients removal from secondary effluent.

In this study, a novel algal biofilm membrane photobioreactor (BMPBR) equipped with solid carriers and submerged membrane module was developed for attached growth of Chlorella vulgaris and secondary effluent treatment. The volumetric microalgae production achieved in BMPBR was 0.072 g L(-1) d(-1), which was 1.44-fold larger than that in suspended growth membrane photobioreactor (MPBR). Furthermore, 72.4% of the total produced algal biomass was immobilized as algal biofilm in BMPBR. Advanced nutrients removal from secondary effluent was achieved both in BMPBR and MPBR, with average reduction of about 85% for PO4(3-)-P in the stable stage. Additionally, BMPBR showed better nitrogen removal performance than MPBR due to its higher algal biomass productivity. Moreover, with the filtration effect of the submerged membrane module in the reactor, suspended microalgae could be completely isolated from the effluent and a low average SS concentration of 0.28 mg L(-1) was achieved in the effluent of BMPBR.

Crucial role of heme oxygenase-1 in the sensitivity of acute myeloid leukemia cell line Kasumi-1 to ursolic acid.

Ursolic acid (UA), which has been used extensively as an antileukemic agent in traditional Chinese medicine, is safely edible if originating from food. We found that the apoptotic rate of acute myeloid leukemia (AML) subtype M2 (AML-M2) cell line Kasumi-1 treated by UA was higher than those of other leukemia cell lines, but was not as high as that treated by arabinofuranosyl cytidine (Ara-C), suggesting that UA is an important chemotherapeutic agent to treat AML-M2. Heme oxygenase-1 (HO-1) is a key enzyme exerting potent cytoprotection, cell proliferation, and drug resistance. HO-1 in Kasumi-1 cells was upregulated by being treated with low-dose rather than high-dose UA. Inhibition of HO-1 by zinc protoporphyrin (ZnPP) IX sensitized Kasumi-1 cells to UA, and the apoptotic rate was close to that induced by Ara-C (P<0.01). The sensitizing effect of ZnPP was associated with caspase activation, bcl-2 downregulation, and PARP activation. After silencing HO-1 by siRNA transfection with lentivirus, the cells' proliferation induced by UA was increased as it was by Ara-C. Furthermore, combining ZnPP with UA prolonged the survival of mice bearing the AML subtype M2 tumor with smaller volume of tumor and size of spleen. The results showed that the Kasumi-1 cell line was the most sensitive to UA, but the apoptotic effect was inferior to that treated by Ara-C because of HO-1 upregulation. AML-M2 can feasibly be treated by target-inhibiting HO-1 that enhances the antileukemia effects of UA in vitro and in vivo.

[Protoplasts isolation, purification and plant regeneration of Pinellia cordata].

The main factors which affected the isolation, purification and cultivation of Pinellia cordata protoplasts from leaves were studied. The results indicated that the optimum enzyme solution for P. cordata leaves was 13% CPW + 1.0% Cellulose +0.1% Pectolase, at pH 6.0, temperature (25-28 degrees C ) for 4 h. The sucrose density gradient centrifugation was adopted to purificate the protoplasts collected, when 25% sucrose was used as mediator, centrifugating at 500 rpm for 10 min. When the protoplasts were shallow liquid and liquid-solid double layer cultured on the medium of MS + 0.5 mg x L(-1) 6-BA + 0.25 mg x L(-1) NAA + 13% mannitol at the density of 2.5 x 104 protoplasts/mL, or fed and nursed cultured at the density of 100-500 protoplasts/mL, cell division could be observed for 3 days; granular calli appeared for 30 days. Calli was proliferated on the medium of MS + 0.5 mg x L(-1) 6-BA + 0.25 mg x L(-1) NAA solidified by 0.55% agar, and differentiated and regenerated after 5-6 months. Plant generation of P. cordata is successfully established.

Rapid analysis of Fructus forsythiae essential oil by ionic liquids-assisted microwave distillation coupled with headspace single-drop microextraction followed by gas chromatography-mass spectrometry.

A rapid, green and effective miniaturized sample preparation and analytical technique, i.e. ionic liquids-assisted microwave distillation coupled with headspace single-drop microextraction (ILAMD-HS-SDME) followed by gas chromatography-mass spectrometry (GC-MS) was developed for the analysis of essential oil (EO) in Fructus forsythiae. In this work, ionic liquids (ILs) were not only used as the absorption medium of microwave irradiation but also as the destruction agent of plant cell walls. 1-Ethyl-3-methylimidazolium acetate ([C2mim]OAc) was chosen as the optimal ILs. Moreover, n-heptadecane (2.0 µL) was selected as the appropriate suspended solvent for the extraction and concentration of EO. Extraction conditions of the proposed method were optimized using the relative peak area of EO constituents as the index, and the optimal operational parameters were obtained as follows: irradiation power (300 W), sample mass (0.7 g), mass ratio of ILs to sample (2.4), temperature (78°C) and time (3.4 min). In comparison to previous reports, the proposed method was faster and required smaller sample amount but could equally monitor all EO constituents with no significant differences.

[Effects of early neurodevelopmental treatment on motor and cognitive development of critically ill premature infants].

OBJECTIVE: To study the effects of neurodevelopmental treatment (NDT) on motor and cognitive development of critically ill premature infants. METHODS: There were 203 infants, who were less than 32 week gestational age (GA), with very-low-birth weight, were included in the study. The infants were assigned to NDT group (n = 96) or control group (n = 107) according to the parent's decision. The infants in NDT group received NDT intervention once per week from corrected age (CA) 1 month to 3 months, and 3 to 5 times per week in the following 9 months. NDT intervention included elongation, establishing alignment, optimizing base of support, facilitation and inhibition, stimulation for activating muscle activity. Both groups received family intervention, such as massage and exercise based on early education. Bayley Scales of Infant Development-II (BSID-II) score was assessed at 3,6,9,12 months CA for all the infants. RESULTS: NDT intervention achieved significant effects on motor and cognitive development. Mental development index (MDI) and psychomotor development index (PDI) of BSID-II were significant higher in NDT group (P < 0.05). Cerebral palsy was finally diagnosed in 15 children in NDT group (16.67%) and 12 (12.12%) in control group at 12 months CA, there was no significant difference between the two groups (P > 0.05). CONCLUSION: NDT intervention can improve motor and cognitive development in critically ill premature infants within 12 months CA.

[Study on two strains of mouse developing dampness-heat models infected by Dengue virus].

OBJECTIVE: To establish two strains of mouse developing dampness-heat syndrome models infected by Dengue virus and to compare the difference in infection, so as to choose a suitable mouse strain for modeling. METHODS: According to the modeling methods of the seasonal febrile disease of the dampness-heat syndrome in Chinese medicine, BALB/C and C57BL/6 mice were respectively treated with complex factors as the high glucose and high fat forage + high-temperature chamber + Dengue virus. At the same time a normal control group, the virus infection group (modeled by Dengue virus infection), and the dampness-heat group (modeling by pure dampness heat circumstance) were set up. Changes of the body temperature, platelet counts, virus in the separate serum, pathological changes of the liver, and serological indicators were observed to compare the modeling difference. RESULTS: After modeling low-grade fever appeared in mice in the high-temperature chamber. Compared with the normal control group, the platelet count decreased in mice of the BALB/C model group. AST increased in both BALB/C and C57BL/6 mice and the virus infection group. TC and TG increased in BALB/C model group and the dampness-heat group, with statistical significance (P<0.05). Various degrees of pathological changes were shown in the liver tissue of each group, with the most severe one in the BALB/C model group. The serum virus titers were detected with Real-time PCR after modeling. The virus load was 2.9 x 10(4) - 5.5 x 10(4) copies/mL. No significant difference was found among these groups. CONCLUSIONS: The mouse model of dampness-heat syndrome infected by Dengue virus was primarily established. When compared the infection between BALB/C mice and C57BL/6 mice, BALB/C mice were more suitable for modeling.