Andrographis paniculata from different parts and origins were analyzed by UPLC-PDA fingerprint to provide refererice for related preparation technology. Using the peak of andrographolide as reference, 27 common peaks were identified, and digitized UPLC-PDA fingerprints for 23 batches of andrographis paniculata were established in this research. Principal component analysis (PCA) was carried out after feature extraction. The contents of andrographolide, neoandrographolide, deoxyandrographolide, dehydroandrographolide were determined by external standard method. The Plackett-Burman design combined with pareto chart was used to analyze the factors influencing the robustness of the method. It was found that the medicinal part has a more remarkable influence on the quality of andrographis paniculata than the origin. The contents of the 4 lactones the differ greatly in the different parts of andrographis paniculata, and the pH of the mobile phase is an important factor that influenced the robustness of the method.
Andrographis ; chemistry ; Chromatography, Liquid ; methods ; Diterpenes ; analysis ; Drug Stability ; Glucosides ; analysis ; Principal Component Analysis ; Tetrahydronaphthalenes ; analysis

OBJECTIVETo study the role of osteocyte in bone remodeling due to mechanical loading in vitro.

METHODSMLO-Y4 osteocyte-like cells were exposed to fluid flow-induced shear stress(12dyn/cm(2))for 0, 1, 2, 4, 6, 12, and 24 hours. Osteocyte exposed to shear stress at different time points were used in co-culture system for 9 days, and then the cells were stained with tartrate-resistant acid phosphatase on the 9(th) day and the amount of positively stained osteoclasts were counted and compared. The expressions of osteoprotegerin (OPG) and receptor activator of nuclear factor-kappa (RANKL) were detected by semi-quantitative reverse transcription polymerase chain reaction (semi-quantitative RT-PCR).

RESULTSCompared with bone cells without stimulation with fluid flow-induced shear stress, the amount of osteocytes significantly decreased at all time points after the application of fluid flow-induced shear stress (all P<0.05). The OPG expression at mRNA levels was significantly up-regulated in the first 12 hours (P<0.001), the RANKL mRNA expression was significantly down-regulated in the first 4 hours (P<0.05), and the RANKL/OPG ratio significantly decreased within 12 hours (P<0.01). However, all these indicators showed no significant difference at 24 hours when compared with the pre-stimulation level.

CONCLUSIONOsteocytes may act as mechanosensors that are able to inhibit bone resorption after mechanical loading; however, such effect shows certain adaptation ability to shear stress as time goes.

Animals ; Cell Differentiation ; Cells, Cultured ; Coculture Techniques ; Male ; Mice ; Osteoclasts ; cytology ; Osteocytes ; cytology ; Osteoprotegerin ; metabolism ; RANK Ligand ; metabolism ; Stress, Mechanical

Objective:To study the expression of osteoprotegerin(OPG)and receptor activator nuclearfactor kappa B ligand(RANKL)at protein level in human periodontal ligament cells(HPDLCs),and theeffect of 1?,25(OH)_2 vitamin D_3[1,25(OH)_2 vitD_3] on the secretion of OPG protein in vitro.Meth-ods:HPDLCs were harvested in vitro by sequential digestion with trypsin and collagenase.The expressionof RANKL in HPDLCs at protein level was tested by immunocyto-chemistry.Enzyme-linked immuno-adsordent assay(ELISA)was used to detect the OPG protein which was secreted into the culture mediumby HPDLCs cultured with and without 10~(-8) mol/L 1?,25(OH)_2 vitD_3 on the 0,2nd,4th,and 6th days,respectively.Results:RANKL protein was detected on the membrane and plasma of HPDLCs,and OPGprotein was secreted in the culture medium.The secretion of OPG protein was down-regulated by 10~(-8)mol/L 1?,25(OH)_2 vitD_3.Conclusion:HPDLCs have the bone metabolism system of OPG/RANKL,which works during the process of 1?,25(OH)_2 vitD_3 inducing HPDLCs.The conclusion has laid thegroundwork for the study on bone remodelling mechanisms of HPDLCs.

OBJECTIVESTo construct the expression vector pGenesil-3-shRNA that can express the short hairpin RNAs (shRNA) silencing Kir6.2 gene and to study its influence on the proliferation and invasion of HepG2 cells.

METHODSTwo shRNA silencing Kir6.2 genes were transcript synthesized intracellularly by expressed templates of plasmid vector pSilence-3, and the target sequence of Kir6.2 gene was inserted into the upstream of the reporter gene in order to construct the recombinant plasmid vector pGenesil-3. Plasmids containing 2 different sequences of human Kir6.2 mRNA coding region were constructed and transfected into HepG2 cells by using lipofectamine 2000 methods. The experiment was divided into 4 groups: SK (normal), SK-HK (negative control), SK-K1 (transfected with the interfering sequence 1) and SK-K2 (transfected with the interfering sequence 2) groups. A selected single clone was cultured after screening by G418. The expression of Kir6.2 protein was detected by Western blot. MTT assay and Transwell system were used to observe the proliferation and invasion of HepG2 cells.

RESULTSThe recombinant expression plasmid pGenesil-3 was successfully constructed and underexpression of Kir6.2 gene in HepG2 cells was detected by Western blot. Underexpression of Kir6.2 gene significantly decreased the proliferation and invasion of the HepG2 cells.

CONCLUSIONshRNA can inhibit the expression of Kir6.2 gene in the HepG2 cells, and underexpression of Kir6.2 gene decreased the proliferation and invasion of the HepG2 cells.

Cell Proliferation ; Genetic Vectors ; Hep G2 Cells ; Humans ; Neoplasm Invasiveness ; Plasmids ; Potassium Channels, Inwardly Rectifying ; genetics ; RNA, Small Interfering ; genetics

OBJECTIVETo explore the role of the fibroblast transdifferentiation into myofibroblasts (MFBs) in the pathogenesis of systemic sclerosis (SSc) and investigate the influence of transforming growth factor β(1) (TGF-β(1)) and blocking of its signal transduction on fibroblast transdifferentiation.

METHODSFibroblasts derived from the skin lesions of SSc patients and normal skin tissue were cultured in vitro. The proportion of MFBs in the fibroblast culture was analyzed qualitatively using immunocytochemistry and quantitatively with ELISA for α-smooth muscle actin (α-SMA). The changes in fibroblast transdifferentiation were observed after addition of TGF-β(1) in the cell culture and after blocking the signal transduction of TGF-β(1).

RESULTSThe fibroblasts isolated from SSc patients and control subjects showed a similar morphology. The mean number of cells positive for α-SMA in SSc group was significantly higher than that in the control group (P<0.01). As culture time extended, α-SMA levels of the two groups both increased gradually (P<0.01), but the increments were significantly greater in SSc group than in the control group at 24, 48, and 72 h (P<0.05 all). Addition of TGF-β(1) resulted in significantly increased α-SMA levels in both groups (P<0.05), and SSc group showed significantly higher α-SMA levels than the control group at 24, 48, and 72 h (P<0.01). In the presence of TGF-β(1), blocking of Smads, ERK/MAPK, and p38MAPK pathways, but not JNK/MAPK pathway, caused an obvious decrease in α-SMA levels in the fibroblasts in both groups.

CONCLUSIONThe fibroblasts in the skin lesion of SSc patients have strong potential of transdifferentiation into MFBs, and TGF-β(1) can promote this transdifferentiation process possibly involving Smads, and ERK/MAPK, and p38MAPK signalling pathways.

Actins ; metabolism ; Adult ; Cell Transdifferentiation ; physiology ; Cells, Cultured ; Female ; Fibroblasts ; pathology ; Humans ; Male ; Myofibroblasts ; pathology ; Scleroderma, Systemic ; pathology ; Signal Transduction ; Skin ; pathology ; Transforming Growth Factor beta1 ; metabolism

Objective:To explore the relationship among psychological distress,adult attachment,and social support in primary caregivers of cancer patients.Methods:A total of 208 primary caregivers of cancer patients in one third grade hospital in Anhui Province were recruited.The 10-item Kessler Psychological Distress Scale(K10),Experiences in Close Relationships Inventory(ECR) and Social Support Questionnaire(SSQ) were used to explore psychological distress,adult attachment,and social support status.Results:The average K10 score was (21.5 ± 7.5).Multiple linear regression analysis indicated that caregiver gender,pressure on patient care,and attachment anxiety had positive prediction on psychological distress (β =2.30,3.02,0.13),while residence had negative prediction on psychological distress (β =-3.22).Conclusion:It suggests that the psychological distress is related to attachment anxiety and social support among the primary caregivers.

To study relevant risk factors of Shenmai injection induced adverse reactions by using Logistic model and ROC curve, and made the prediction for the occurrence of relevant adverse reactions/events. Case data of patients treated with Shenmai injection were collected by using the prospective, multi-center, large-sample, nested-case control method, in order to analyze the risk factors of Shenmai injection-induced adverse reactions/events, establish the logistic model and draw the receiver operating characteristic (ROC) curve for risk factors. During the study, 7632 patients (including 3 477 males and 4 155 females) were included, and eight of them suffered adverse reactions/events. Based on a multi-factor Logistic model analysis, the age (> or = 50 years) (OR = 5.061, 95% CI: 2.197-7.924; P = 0.001), the total number of medication days (OR = -1.020, 95% CI: -l.652 - 0.388; P = 0.002) and the single dose (OR = 0.245, 95% CI: 0.127-0.364; P = 0.000) were significant independent risk factors for Shenmai injection-induced adverse reactions/events. According to the results, ROC curves were drawn with age (> or = 50 years), the total number of days of inedication and single dose; The area under ROC curves the joint predictor (0.9753, 95% CI: 0.9443-1.000, P < 0.005) was larger than that of the other three single indexes, with a higher risk prediction value. The independent risk factors for Shenmai injection-induced adverse reactions/events included the age (> or = 50 years), the total number of days of medication and single dose. In clinical practice, the age (> or = 50 years), the total number of days of medication and the medication dose can be substituted in the joint predictor calculation formula (P = 1 / [1 + e(-(-21.58 + 5.061 x Xage - 1.020 x Xd + 0.245 x X(mL)] to predict the potential adverse reactions of patients and adjust the dosage regimen.
Adolescent ; Adult ; Aged ; Aged, 80 and over ; Child ; Child, Preschool ; China ; epidemiology ; Drug-Related Side Effects and Adverse Reactions ; epidemiology ; etiology ; Drugs, Chinese Herbal ; administration & dosage ; adverse effects ; Female ; Humans ; Infant ; Logistic Models ; Male ; Middle Aged ; Prospective Studies ; ROC Curve ; Risk Factors ; Young Adult

OBJECTIVEIn order to observe the therapeutic efficacy of food control to recurrent oral ulcer (ROU), alimentary control of certain foods was employed to relieve outbreak of ROU.

METHODSThe kits for food intolerant IgG of certain food were used to test the intolerant food of fifty patients with ROU. Observations and assessments for alimentary control were made after three months' treatment on these patients.

RESULTSThe top three of intolerant foods were crab, egg and milk and the remission rate of ROU reached 74% after treatment.

CONCLUSIONThe result of food intolerant IgG testing has certain function to alimentary control therapy for remitting the outbreak of ROU.

Adult ; Diet ; Female ; Food ; Humans ; Male ; Middle Aged ; Oral Ulcer

OBJECTIVETo compare between MLO-Y4 osteocyte and osteoblast to support osteoclast formation in co-culture system.

METHODSMLO-Y4 cells and murine osteoblast cells were co-cultured with bone marrow cells with or without vitamin D₃ presence.Bone marrow cells were as control group. Tartrat resistant acid phosphatase (TRAP)+ giant cells with three or more nuclei were counted and compared under a microscope at day 9.

RESULTSIn the absence of vitamin D₃, (1963.3 ± 93.1)/plate osteoclasts were observed when MLO-Y4 cells co-cultured with bone marrow cells in 24-well plate.While only (12.7 ± 5.5)/plate osteoclasts were found in the osteoblast group, and (6.0 ± 1.0)/plate in control group. The statistical difference occurs for any two groups (P < 0.05). Vitamin D₃ could significantly increase osteoclast formation in the three groups.

CONCLUSIONSOsteocytes could induce osteoclastogenesis without the presence of vitamin D₃ and vitamin D₃ could enhance the induction effects of MLO-Y4 and osteoblast cells.

Animals ; Cell Line ; Cholecalciferol ; chemistry ; Coculture Techniques ; Culture Media ; chemistry ; Mice ; Osteoblasts ; cytology ; Osteoclasts ; cytology ; Osteocytes ; cytology

AIMTo investigate protective effects of ginkgolide B (GB) in different administration modes on glutamate-induced neuronal damage.

METHODSEssential GB were obtained by supercritical CO2 fluid extraction. Glutamate excitotoxicity were examined in primary cultures from neonatal Wistar rat, by using of Trypan blue dye staining, testing the lactate dehydrogenase leakage from cultured neurons and terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) method. The protective effects of GB in different administration modes (pre-treatment and post-treatment) were adopted and compared with the NMDA receptor uncompetitive antagonist-MK-801 in acute-treatment.

RESULTSTreatment with GB in two administration modes both could increase ratio of surviving neuron, decrease LDH efflux and reduce ratio of neuron apoptosis in different degree, depended on dose in certain range. The protective effect of pre-treatment was superior to post-treatment, but inferior to MK-801.

CONCLUSIONGB can protect neurons against glutamate damage, and preventive using has more efficiency. The potential mechanism of its neural protection may be not only related to PAF receptor. If the predominant protection effect of GB in pretreatment is considered, precautionary intervention to high-risk population could have more value.

Animals ; Cells, Cultured ; Dizocilpine Maleate ; pharmacology ; Ginkgolides ; administration & dosage ; pharmacology ; Glutamic Acid ; adverse effects ; Hippocampus ; drug effects ; metabolism ; Lactones ; administration & dosage ; pharmacology ; Neurons ; drug effects ; metabolism ; Rats ; Rats, Wistar