OBJECTIVE To investigate the effect of berberine on differentiation of rat bone marrow mesenchymal stem cells (MSCs) to adipocytes and its mechanism. METHODS Rat MSCs were isolated and cultured, adipocytic differentiation was induced with adipogenesis-inducing medium (AIM). Cells were assigned into 6 groups:normal control, AIM group, AIM+berberine 0.1, 0.3, 1 and 3 μmol·L-1 groups, respectively. Morphology characteristics of mesenchymal stem cells were observed under an inverted microscope and adipocyte levels were analyzed by oil O staining. Alkaline phosphatase (ALP) activity was detected using p-nitrophenyl phosphate as a substrate. The cell survival was determined by MTT assay. Expressions of peroxisome proliferator activated receptor γ (PPARγ), fatty acid binding protein (aP2) and CCAAT enhancer-binding protein α (C/EBPα) mRNA were detected by semiquantitative RT-PCR. RESULTS Compared with normal control group, MSCs adipogenic differentiation, PPARγ, aP2 and C/EBPα mRNA expression significantly increased in AIM group (P<0.01), ALP activity in AIM group significantly decreased (P<0.01). Compared with AIM group, berberine inhibited MSCs adipogenic differentiation (P<0.01) and berberine 0.1, 0.3, 1 and 3 μmol·L-1 increased ALP activity by 26%, 54%, 81% and 122%, respectively. Berberine 3 μmol·L-1 significantly downregulated PPARγ expression (0.91±0.10 vs 1.34±0.06) (P<0.01), aP2 (1.05±0.10 vs 1.53±0.09) (P<0.01) and C/EBPα mRNA (1.24±0.06 vs 1.54±0.09) (P<0.01). Berberine had no effect on proliferation of MSCs. CONCLUSION Berberine inhibits differentiation of MSCs into adipocytes, which might be closely related to the downregulation of PPARγ, aP2 and C/EBPα mRNA.

The paper presents a flexible and simple direct analysis in real-time ( DART) device without grid electrode for mass spectrometer injection. It contained inert carrier gas, ionizer, heater and temperature-controller etc. Excluding the grid electrode and then reducing the structure units, the device could be easy to build up in low cost and flexible to connect with a variety of mass spectrometers. The experimental conditions like the kind of carrier gas, flow rate and temperature were investigated for the device. By using argon as carrier gas, flow rate as 7. 5 L/min, and temperature of heat tape as 300 ℃, the device was used to analyze benzene alcohol, linoleic acid, dichlorvos emulsion, mosquito coils, citrus peel, and sample ( propranolol hydrochloride) on thin-layer plate combined with mass spectrometer. The results were accurate and the device was stable and reliable.

The ionic liquid butylamine 2,5-dihydroxybenzoate (DHB-BuN) was chosen as the matrix for improving repeatability of oligosaccharides quantitation, and imaging oligosaccharides in soybean and leaf by matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS).Using the ionic liquid matrix methanol solution (20%, V/V) to cover sample slice, and setting MS acquisition parameters including polarity as positive and laser power as 70%, the MS imaging experiment was performed to analyze oligosaccharides and their distribution in soyben and soybean leaf.As a result, the MALDI-MS analysis of 3 kinds of oligosaccharides including sucrose, raffinose and stachyose was performed using the ionic liquid matrix and the reproducibility RSDs were less than 3% for in-spot and less than 4% for spot-to-spot.The standard curves linear correlation coefficients (R2) were all no less than 0.996 between MS peak intensity and oligosaccharides concentraions in the concentration rage of 0.062-1.00 mg/mL, which showed quantitative power in MALDI-MS analysis.The matrix was used for MALDI-MS imaging analysis of soybean chips and soybean leaf surface disaccharide, trisaccharide and tetrasaccharide to obtain the soybean mass spectrum and spatial resolution of 150 μm of the three oligosaccharides imaging.Three kinds of oligosaccharides in the soybean roughly balanced distribution, and the oligosaccharides distribution in the leave was mostly at its tip part.The contents of the 3 kinds of oligosaccharides could be estimated by their MS peak intensity and above standard curve.The result suggested that the ionic liquid DHB-BuN matrix could be well applied in MALDI-MS imaging analysis of oligosaccharides in different sample.

BACKGROUND: It has been reported that a combination of estrogen and progestin has a protective synergistic effect on osteoporosis with only little side effects.OBJECTIVE: This study was designed to investigate the effect of a combination of norethisterone and ethinyl estradiol (EE) on bone mass in ovariectomized rats.DESIGN: This study was a randomized controlled experiment.SETTING:It was conducted at the Department of Pharmacology of Guangdong Medical University.MATERIALS: Twenty-four specific pathogen free (SPF) unmated SD rats were selected, aging 4 and half months and weighing 230±15 g.METHODS: The experiment was conducted in the Department of Pharmacology of Guangdong Medical College from May to November 2002.These rats were randomly divided into 3 groups: pseudo-operation group, ovariectomy group and compound norethisterone group, each containing 8 rats. For the former two groups, ethanol solution (volume fraction=0.056), at a dose of 5 mL/(kg.d), was administered by gavage. While for compound norethisterone group, 60μg/(kg·d) norethisterone and 3.5μg/(kg·d) EE were given by gavage (according to the dosage for human, which was 20-35 μg EE combined with norethisterone). Duration of treatment was 90 days for all the animals. Then their tibias were removed. Employing a fullyautomatic imaging analysis system, osteoclasts and the relevant dynamic and static parameters reflecting secondary trabeculaes formation region in proximal tibias were measured. Respectively, the humeral samples were removed and employing the palsma emission spectrograph of full-spectrum direct reading, calcium content and hydroxyproline content in bone samples were measured. Meanwhile, urine calcium and hydroxyproline concentrations were examined as well.MAIN OUTCOME MEASURES: ①The trabecular area (Th. Ar), trabecular thickness (TbkTh), trabecular number (Tb.N) and trabecular separation (Tb. Sp) and the changes in static parareters of perimeters of osteoclasts were investigated. Variance in percent labeled perimeter (L. Pm ％), mineral apposition rate (MAR) and bone formation rate (BFR/BV) were also calculated. ②Changes in serum alkaline phosphatase (AKP), calcium and hydroxyproline contents in bone and urine were all measured.RESULTS: All the 24 rats entered the analysis procedure. Compared to pseudo-operation group, for the ovariectomy group, Tb. Ar and Tb.N decreased, Tb. Sp increased and osteoclast perimeter significantly increased (P＜0.01). Addtionally, the bone formation markers increased apparently with an increase in L. Pm ％ and MAR (P＜0.05) and a significant increase in BFR/BV (P＜0.01). Compared with the ovariectomy group, for the compound norethisterone group,the bone mass and the Tb.N increased, marked by an increase of 82％ in Tb. Ar and an increase of 83％ in Tb.N (P＜0.05), and the Tb.Sp decreased, marked by a decrease of 51％ (P＜0.05). Meanwhile, there was a decrease of 52.5％ in osteoblast perimeter (P＜0.01), an increase in organic bone matrix and a decrease in urine hydroxyproline (P＜0.05).CONCLUSION: A combination of estrogen and progestin has a protective synergistic effect on ovariectomized rats with osteoporosis, and it is capable of increasing the organic bone matrix without significant inhibitory effects on bone formation. The experimental dosage of the compound was calculated according to the clinical dosage, 20-35 μg estrogen combined with a progestin, which will yield optimal protective effects on bone sometimes.

Aim To investigate the preventive effects of misoprostol on osteoporosis in aged ovariectomized(OVX)rats.Methods Female,10-month-old SD rats were ovariectomized(OVX)and,2 months later,were treated with misoprostol or controls for 2 months.The static and dynamic parameters in trabecular bone of the forth lumbar vertebrae(LV4)were examined with histomorphometrical analyses;the fifth lumbar vertebrae(LV5)was used to perform the compression test.Results Compared with the data from the sham-operated rats,the percent trabecular area and elastic modulus significantly decreased in OVX rats.Correspondingly,the bone break load and break stress decreased of post OVX was compared with those of sham-operated rats.Misoprostol increased the percent trabecular area by 21.6% compared with OVX rats,but it couldn't meet the statistical significance.Misoprostol enhanced the break load and elastic modulus compared with OVX rats.Conclusion Misoprostol can improve biomechanics of bone in ovariectomized osteoporosis rats.

AIM To determine whether norethisterone in combination with ethinylestradiol can completely prevent bone loss in ovariectomized rats. METHODS Twenty-four Sprague-Dawley female rats at the age of 4.5 months were sham-operated and treated orally with vehicle, or ovariectomized (OVX) and treated orally with either vehicle or combined norethisterone (norethisterone at 60 ?g?kg -1?d -1 and ethinylestradiol at 3.5 ?g?kg -1?d -1) for 90 days. Double in vivo fluorochrome labeling was administrated. The undecalcified longitudinal proximal tibial metaphyseal sections were used for the bone histomorphometric analysis. The humerus and urine calcium contents were assayed by the method of atomic absorption spectrometry. The humerus and urine hydroxyproline contents were assayed by colorimetry. Blood serum alkaline phosphatase (ALP) were tested by BECKMAN auto bio-chemistry analyzer. RESULTS After 90 days post OVX the cancellous bone mass and showed high bone turnover indices. Bone hydroxyproline contents were lost markedly. The ALP activity increased. The urine hydroxyproline contents increased significantly. The combined norethisterone treated group prevented bone lost when compared with OVX. The combined norethisterone were shown to inhibit osteoclasts surface and decrese bone turnover rate. The combined norethisterone can increase hydroxyproline contents and reduce urine hydroxyproline contents significantly from OVX group. CONCLUSION Combining norethisterone can prevent OVX-induced cancellous bone loss in rats.

AIM To determine whether epimedium pubescens flavonoids(EF) in combination with diethylstilbestrol(DES) can completely prevent bone loss in ovariectomized rats. METHODS Sixty Sprague-Dawley female rats at age of 4 months were divided into six groups. Ten rats were sacrificed at age of 4.5 months before operation. The other rats were sham-operated and treated orally with vehicle or ovariectomized (OVX) and treated orally with either vehicle, or diethylstilbestrol(DES) at 22.5 ?g?kg -1 ?d -1 , or EF at 300 mg?kg -1 ?d -1 , or EF 300 mg?kg -1 ?d -1 in combination with DES 22.5 ?g?kg -1 ?d -1 for 90 days. Double in vivo fluorochrome labeling was administrated. The undecalcified longitudinal proximal tibial metaphyseal sections were used for [FQ(9*2。46,X-WZ]-the bone histomorphometric analysis. RESULTS DES prevented OVX-induced bone loss and decreased body weight gain and total serum cholesterol,but it increased uterine luminal epithelial thickness.Combination of EF with DES completely prevented OVX-induced bone loss and did not increase uterine luminal epithelial thickness.The effect of the combination in preventing bone loss was more significant than that of DES. CONCLUSION EF in combination with DES showed synergistic effect on prevention of osteoporosis induced by ovariectomy.-

AIM: To study the effects of epimedium pubescens flavonoids (EF) on the skeleton in ovariectomized rats. METHODS: Forty 4.5-month-old Sprague-Dawley female rats were randomly divided into 4 groups. Rats in sham group were sham-operated and treated by daily oral gavage with vehicle. Rats in other three groups were bilaterally ovariectomized (OVX) and treated by either daily oral gavage with vehicle, or diethylstilbestrol (DES) at 22.5 ?g?kg~ -1?d~ -1, or EF at 300 mg?kg~ -1?d~ -1 for 90 days. Bone histomorphometric analysis of the proximal tibial metaphysis (PTM), fifth lumbar vertebrae (LV5) and tibial shaft (Tx) was performed in undecalcified sections. The left femur was collected to determine bone weight, contents of calcium (Ca) , phosphorus (P ) and hydroxyproline. The uterine weight and the uterine luminal epithelial thickness (ULET) were determined. RESULTS: A significant increase in contents of Ca and P of femur was found in EF group. A tendency of increase was found in %Tb.Ar of PTM, but no significant change was found in bone histomorphometric parameters of LV5 and Tx in EF group. EF had no effect on uterine weight and ULET. CONCLUSION: EF can prevent OVX-induced bone mineral loss of femur, but does not prevent bone loss of PTM and LV5.

AIM: To study the correlative changes of bone mineral contents and histomorphometry of lumbar vertebrae in ovariectomized rats by analysis of T-type correlation degree. METHODS: Forty 10.5-month-old virgin female Sprague-Dawley rats were randomly divided into five groups: (1) Basal: at 10.5 mon of age; (2) sham-1:sham-operated at 13 mon of age; (3) OVX-1: ovariectomized at 13 mon of age; (4) sham-2: sham-operated at 16 mon of age; (5) OVX-2:ovariectomized at 16 mon of age. After ovariectomy, all rats were treated orally with NS at 5 ml?kg -1?d -1. At the end-point of study, the undecalcified longitudinal fourth lumbar vertebra (LV4) sections were cut and stained with Goldner's Trichrome for bone histomorphometric analyses. The fifth lumbar vertebra (LV5) was dried with temperature and digested with acid for testing of bone mineral content. Then the effects of bone mineral contents on bone histomorphometry were assessed by analyzing the T-type correlation degree in the Grey system. RESULTS: All degrees of correlation between bone mineral contents and static histomorphometric parameters (trabecular bone volume) (BV/TV), trabecular thickness (Tb.Th) were positive, but for dynamic histomorphometric parameter (BFR/BV), the correlation degrees were negative. The effect of contents of calcium (Ca) and phosphorus (P) on histomorphometric parameters of lumbar vertebrae was much greater than that of the other bone mineral contents. CONCLUSIONS: Analysis of T-type correlation degree can evaluate the correlative changes of bone mineral contents and histomorphometry of lumbar vertebrae in ovariectomized objectively and fairly.

Objective To investigate the morphological changes of the head,body and tail of the hippocampus in patients with Alzheimer's disease (AD).Methods Totally 30 AD (AD group),30 mild cognition impairment (MCI) patients (MCI group) and 30 normal old persons (normal control group) underwent brain MR scan.Based on the MR images,the head,body and tail of the hippocampus were segmented and the volume was measured.The difference of the volume in total,head,body and tail of hippocampus in AD group,MCI group and normal control group were compared.The correlation between hippocampal volume and the scores of neurological assessment scale was analyzed.Results The left total,head,body and tail volume were greater than those of the right side in 3 groups (all P＜0.05).For the comparison of total volume of left and right sides among the 3 groups,AD group was less than normal control group (both P＜0.01) and MCI group (both P＜0.05),MCI group was less than normal control group (both P＜0.05).Compared with the normal control group,the head,body and tail volume of hippocampus of both sides were decreased in AD group (all P＜0.05) and the volume of left head,body and right head of hippocampus in MCI group were decreased (all P＜0.05).Both sides hippocampus's head and body volume of AD group were less than those of MCI group (both P＜0.05).The scores of minimum mental state examination (MMSE) were positively correlated with the total and the head,body,tail volume of bilateral hippocampus.Except volume of left tail,there were negative correlations between the scores of activities of daily living (ADL),clinical dementia rating (CDR) and the volume of total and head,body,tail of bilateral hippocampus,which were positively correlated with Montreal cognitive assessment (MoCA) score.Conclusion The volume of bilateral hippocampus was significantly reduced in AD patients.The head of hippocampal volume was significantly reduced and the body and tail was not obvious.