BACKGROUND:Previous studies found that Semen Ziziphi Spinosae extract prolongs slow-wave sleep and promotes growth hormone secretion in mice by increasing the expression of brain tissue serotonin 1A receptor (5-HT1AR),which binds to serotonin,thus leading to bone growth. Serotonin 1A receptor acts as a protein,and its expression abundance is regulated by miRNAs. The authors hypothesized that Semen Ziziphi Spinosae extract may regulate the expression of 5-HT1AR through miRNAs and thus exert drug effects. OBJECTIVE:To observe the effect of Semen Ziziphi Spinosae extract on bone growth by regulating the miR-7b-3p/5-HT1AR pathway in mouse brain tissue. METHODS:(1) Kunming mice were divided into a normal control group,a drug administration group (gavage administration of Semen Ziziphi Spinosae extract 0.320 mg/g),a positive control group (gavage administration of jujuboside 0.013 mg/g),a Semen Ziziphi Spinosae extract+5-HT1AR selective inhibitor group (8 μg of P-MPPF,a 5-HTIAR selective inhibitor,was injected into the lateral ventricle every day for the last 3 days during the gavage administration of Semen Ziziphi Spinosae extract). After 25 days,the effects of Semen Ziziphi Spinosae extract on bone growth,serum growth hormone level and brain 5-HT1AR expression were observed. (2) Then the chip method was used to observe differentially expressed miRNAs in the brain tissues of growing mice and ordinary mice. PCR validation and dual luciferase reporter gene assay confirmed the regulatory relationship between the screened Mir-7B-3p and 5-HT1AR. (3) Mouse cerebral cortical cells were cultured and identified in vitro,and the effect of Semen Ziziphi Spinosae extract on 5-HT1AR expression in cerebral cortical cells was observed using western blot. (4) Kunming mice were divided into a normal control group,a medication group,a miR-7b-3p inhibitor group,a medication+5-HT1AR inhibitor group,and a positive control group. The expression of 5-HT1AR in brain tissues of mice and the binding activity of 5-HT and 5-HT1AR were observed. (5) Sprague-Dawley rats were divided into a normal control group,a medication group,a miR-7b-3p inhibitor group,a medication+miR-7b-3p mimics group and a positive control group. Changes in slow wave sleep in mice were observed.RESULTS AND CONCLUSION:(1) Semen Ziziphi Spinosae extract could promote the growth of body length and tibia,growth hormone secretion,and brain tissue 5-HT1AR level in mice. (2) The number of differentially expressed miRNAs screened by GeneChip was 16,of which 13 were up-regulated and 3 were down-regulated. Bioinformatics results predicted that down-regulation of miR-7b-3p could regulate 5-HT1AR expression,and dual-luciferase reporter gene experiments confirmed a direct regulatory relationship between the two. (3) Semen Ziziphi Spinosae extract and silencing of miR-7b-3p expression in cerebral cortical cells could cause high expression of 5-HT1AR. After silencing of miR-7b-3p,5-HT1AR expression,binding activity of serotonin and 5-HT1AR and growth hormone secretion in mouse brain tissue were all elevated. After overexpression of miR-7b-3p,5-HT1AR expression,binding activity of serotonin and 5-HT1AR and growth hormone secretion were all reduced. Accordingly,the slow-wave sleep period in mice was also prolonged or shortened. To conclude,Semen Ziziphi Spinosae extract can reduce the level of miR-7b-3p and increase the expression of 5-HT1AR in brain tissue to prolong slow wave sleep and promote the secretion of growth hormone,thereby playing a postive effect on bone growth. These findings provide a scientific basis for the use of Semen Ziziphi Spinosae extract as a potential measure to promote bone growth.

BACKGROUND:Previous studies found that Semen Ziziphi Spinosae extract prolongs slow-wave sleep and promotes growth hormone secretion in mice by increasing the expression of brain tissue serotonin 1A receptor (5-HT1AR),which binds to serotonin,thus leading to bone growth. Serotonin 1A receptor acts as a protein,and its expression abundance is regulated by miRNAs. The authors hypothesized that Semen Ziziphi Spinosae extract may regulate the expression of 5-HT1AR through miRNAs and thus exert drug effects. OBJECTIVE:To observe the effect of Semen Ziziphi Spinosae extract on bone growth by regulating the miR-7b-3p/5-HT1AR pathway in mouse brain tissue. METHODS:(1) Kunming mice were divided into a normal control group,a drug administration group (gavage administration of Semen Ziziphi Spinosae extract 0.320 mg/g),a positive control group (gavage administration of jujuboside 0.013 mg/g),a Semen Ziziphi Spinosae extract+5-HT1AR selective inhibitor group (8 μg of P-MPPF,a 5-HTIAR selective inhibitor,was injected into the lateral ventricle every day for the last 3 days during the gavage administration of Semen Ziziphi Spinosae extract). After 25 days,the effects of Semen Ziziphi Spinosae extract on bone growth,serum growth hormone level and brain 5-HT1AR expression were observed. (2) Then the chip method was used to observe differentially expressed miRNAs in the brain tissues of growing mice and ordinary mice. PCR validation and dual luciferase reporter gene assay confirmed the regulatory relationship between the screened Mir-7B-3p and 5-HT1AR. (3) Mouse cerebral cortical cells were cultured and identified in vitro,and the effect of Semen Ziziphi Spinosae extract on 5-HT1AR expression in cerebral cortical cells was observed using western blot. (4) Kunming mice were divided into a normal control group,a medication group,a miR-7b-3p inhibitor group,a medication+5-HT1AR inhibitor group,and a positive control group. The expression of 5-HT1AR in brain tissues of mice and the binding activity of 5-HT and 5-HT1AR were observed. (5) Sprague-Dawley rats were divided into a normal control group,a medication group,a miR-7b-3p inhibitor group,a medication+miR-7b-3p mimics group and a positive control group. Changes in slow wave sleep in mice were observed.RESULTS AND CONCLUSION:(1) Semen Ziziphi Spinosae extract could promote the growth of body length and tibia,growth hormone secretion,and brain tissue 5-HT1AR level in mice. (2) The number of differentially expressed miRNAs screened by GeneChip was 16,of which 13 were up-regulated and 3 were down-regulated. Bioinformatics results predicted that down-regulation of miR-7b-3p could regulate 5-HT1AR expression,and dual-luciferase reporter gene experiments confirmed a direct regulatory relationship between the two. (3) Semen Ziziphi Spinosae extract and silencing of miR-7b-3p expression in cerebral cortical cells could cause high expression of 5-HT1AR. After silencing of miR-7b-3p,5-HT1AR expression,binding activity of serotonin and 5-HT1AR and growth hormone secretion in mouse brain tissue were all elevated. After overexpression of miR-7b-3p,5-HT1AR expression,binding activity of serotonin and 5-HT1AR and growth hormone secretion were all reduced. Accordingly,the slow-wave sleep period in mice was also prolonged or shortened. To conclude,Semen Ziziphi Spinosae extract can reduce the level of miR-7b-3p and increase the expression of 5-HT1AR in brain tissue to prolong slow wave sleep and promote the secretion of growth hormone,thereby playing a postive effect on bone growth. These findings provide a scientific basis for the use of Semen Ziziphi Spinosae extract as a potential measure to promote bone growth.

As a type of
Humans ; Legionella/genetics* ; Legionella longbeachae/genetics* ; Legionellosis/drug therapy* ; Pneumonia/diagnosis* ; Retrospective Studies

Objective To establish an evaluation system of HPLC fingerprint of Wanbiqing Pills; To provide references for the quality control ofWanbiqing Pills.Methods Wanbiqing Pills were analyzed with HPLC method. Analysis was performed on Agilent C18 column (4.6 mm×250 mm, 5μm) with the mobile phase of acetonitrile- 0.1%H3PO4 water gradient elution at the flow rate of 1.0 mL/min. The detection wavelength was 258 nm. Sample injection was 20μL. Standard fingerprint ofWanbiqing Pills was established based on Similarity Evaluation System for Chromatographic Fingerprint of TCM (Version 2004 A). The similarity ofWanbiqing Pills from different batches was computed.Results A standard HPLC fingerprint procedure was developed forWanbiqing Pills with 20 common peaks. The similarity of 10 batches ofWanbiqing Pills was not lower than 0.98. Five peaks received chemical confirmation.Conclusion The HPLC method is stable and feasible and with good repeatability, which can be used for the quality control ofWanbiqing Pills.

OBJECTIVE:To establish a method for the contents determination of microelements in Chrysanthemum indicum from different production fields. METHODS:Graphite furnace atomic absorption (GFAAS) was used to determine the contents of Pb,Cd and As;HGAAS was used to determination the content of Hg;and FAAS was used to determine the contents of Cu,Fe, Mn,Ca,Mg and Zn. RESULTS:The linear range was 0-50 μg/L for Pb(r=0.999 9),0-10 μg/L for Cd(r=0.999 2),0-50 μg/L for As(r=0.999 0),0-20 μg/L for Hg(r=0.999 5),0-5 μg/L for Cu(r=0.999 3),0-15 μg/L for Fe(r=0.999 8),0-2 μg/L for Mn (r=0.999 9),0-50 μg/L for Ca(r=1.000 0),0-20 μg/L for Mg(r=0.999 9)and 0-2 μg/L for Zn(r=0.999 8);RSDs of precision, stability and reroducibility tests were lower than 3%;recoveries were 94.25%~97.43%(RSD=1.07%)、94.97%~99.46%(RSD=1.68%)、96.25%~99.46%(RSD=1.09%,n=6)、96.61%~99.91%(RSD=1.26%,n=6)、94.11%~98.41%(RSD=1.68%,n=6)、93.11%~99.59%(RSD=2.73%,n=6)、93.11%~99.48%(RSD=2.63%,n=6)、93.01%~99.85%(RSD=2.49%,n=6)、95.13%~99.75%(RSD=1.58%,n=6)、94.08%~97.37%(RSD=1.18%,n=6),respectively. CONCLUSIONS:The method is simple,stable and reproducible,and can be used for the contents determination of microelements in C. indicum from different pro-duction fields.

Objective To establish the HPLC fingerprint of Taoren Xikang pills and provide the method for the quality control and evaluation of the preparation. Methods Methanol extracts of ten batches of Taoren Xikang pills were analyzed with HPLC-UV method. Analysis was performed on C18 column (4.6 mm× 250 mm, 5 μm) with the mobile phase of acetonitrile-water gradient eluted at the flow rate of 1.0 mL/min. The detection wavelength was 203 nm. Results A standard HPLC fingerprint procedure was developed for Taoren Xikang pills with 26 common peaks. The similarity of 10 batches of Taoren Xikang pills was not lower than 0.9. Conclusion The method is accurate, repeatable and can be used for the quality control of Taoren Xikang pills.