Aged ; Diabetes Mellitus, Type 2 ; Exercise ; Humans ; Physical Fitness

In-vitro assay methods were established to evaluate transactivation and binding activity of compounds on peroxisome proliferator-activated receptor y (PPARγ). Firstly, plasmids were constructed for transactivation assay of PPARγ response element (PPRE) triggered reporter gene expression, and for cell-based binding activity assay of the chimeric receptor, which was fused with PPARγ ligand binding domain (LBD) and yeast transcriptional activator Gal4. Secondly, by using PPARy competitive binding assay based on time resolved-fluorescence resonance energy transfer (TR-FRET), affinities of compounds and drugs to PPARγ were evaluated. In application of these above methods, the PPARγ activating potency and characteristics of different compounds were evaluated, and a novel benzeneselfonamide derivative, ZLJ01, was found to have comparable binding activity and affinity with the well-known PPARy agonist, but lack of PPRE mediated transactivation activity. In preliminary study on in-vitro hypoglycemic activity, ZLJ1 was found to promote insulin-stimulated glucose uptake by liver cells. Therefore, we believe that combining transactivation and binding activity as well as affinity evaluation, the system could be used to screen non-agonist PPARγ ligand as anovel PPARγ modulator
Genes, Reporter ; Hepatocytes ; Hypoglycemic Agents ; chemistry ; Ligands ; PPAR gamma ; agonists ; chemistry ; Plasmids ; Response Elements ; Sulfonamides ; chemistry ; Transcriptional Activation

Kidney injury and decreased chemosensitivity of tumor cells are obstacles with cisplatin (CDDP) chemotherapy. Down-regulation of the organic cation transporter 2 (OCT2) and multidrug resistance-associated protein 2 (MRP2) is a key means to alleviate CDDP-induced kidney injury and increase chemosensitivity. Astragaloside IV (AS IV) is obtained from the well-known traditional Chinese herb Astragalus membranaceus. This study explored the role of AS IV in preventing kidney injury and enhancing the antitumor effect of CDDP by suppressing OCT2 expression in kidney and MRP2 in tumors. This project was reviewed and approved by the Animal Ethics Committee of the First Hospital of Jilin University. The effects of AS IV on CDDP inhibition of tumor growth and promotion of apoptosis were assessed in Lewis lung tumor (LLC)-bearing mice by H&E and TUNEL staining. Kidney injury was assessed by serum biochemical parameters and H&E staining. We used Western blotting and immunohistochemistry assays to detect OCT2 and MRP2 expression in kidney and tumor. The concentration of CDDP in kidney and tumor was measured by HPLC-MS/MS. AS IV enhanced CDDP chemosensitivity by increasing tumor cell apoptosis and slowing tumor growth, and decreased kidney injury as evidenced by lower blood creatinine (Cr) and blood urea nitrogen (BUN). Co-administration of AS IV suppressed MRP2 overexpression induced by CDDP in tumor tissues and may be an important mechanism for enhancing CDDP chemosensitivity. Moreover, AS IV reduced CDDP-induced kidney injury in mice along with suppression of OCT2 expression in kidney. The concentration of CDDP was increased in tumor but decreased in kidney. In total, AS IV not only enhanced the antitumor effect of CDDP by suppressing MRP2 expression in tumor cells, but also decreased kidney injury induced by CDDP. The results provide new insight into the combined use of a chemotherapy drug and natural ingredients to treat cancer.

OBJECTIVEWe aim to investigate the sonodynamic effect induced by hydroxyl acetylated curcumin (HAC) on THP-1 macrophages.

METHODSTHP-1 derived macrophages (1 x 10(5) per milliliter) were cultured with HAC at a concentration of 5 µg/mL for 4 h and then exposed to pulse ultrasound treatment (0.5 W/cm2) for 5 min. Six hours later, cell viability analysis was performed with CCK-8 assay, apoptosis and necrosis analysis were detected with Annexin V/PI staining by flow cytometery.

RESULTSThe cell viability of THP-1 macrophage decreased significantly in the group treated with the combination of HAC and ultrasound (P < 0.01), and HAC-SDT induced both apoptosis and necrosis in THP-1 macrophages, the apoptotic rate was higher than the necrotic rate with appropriate conditions, the maximum apoptosis/necrosis ratio was detected in sonodynamic therapy (SDT) group (P < 0.01).

CONCLUSIONhAC-SDT was effective to induce THP-1 macrophages apoptosis.

Apoptosis ; Cell Line ; Cell Survival ; Curcumin ; pharmacology ; Humans ; Macrophages ; cytology ; drug effects ; Necrosis ; Ultrasonics

To evaluate the effects of rhG-CSF on mobilization of mesenchymal stem cells (MSCs) of mouse bone marrow at different time point, thirty mice were randomly divided into rhG-CSF treatment group and control group. The mice were subcutaneously injected with rhG-CSF in a dose of 80 microg/kg or saline for 5 days. The bone marrow and peripheral blood were obtained at time points of 6, 12, 168 hours after final injection of rhG-CSF or saline. Bone marrow mononuclear cells (BMMNCs) were seeded at density of 1 x 10(6) MNCs onto 12-well plate for culture expansion in DMEM supplemented with 10% FBS, and the number of colony forming unit - fibroblast (CFU-F) was counted after 14 days. The cells were collected by trypsinization and the surface antigens CD34, CD133, CD90 and CD105 were analyzed by flow cytometry. The multi-differentiation of MSCs were done in the culture condition of induced-adipocyte and osteocyte. Peripheral blood MNCs examination was same as the bone marrow. The results indicated that the number of CFU-F of bone marrow in rhG-CSF group was more than that in control group (p < 0.01), the number of CFU-F in rhG-CSF group at 6 hours was more than that at 12 hours and 168 hours, respectively (p < 0.01). There was no obvious difference between CFU-F at 12 hours and at 168 hours (p > 0.05). MSCs were positive for CD90, CD105 and negative for CD34 and CD133. MSCs were found to differentiate into adipocyte and osteocyte in vitro. The CFU-F of PBMNCs obtained and cultured in vitro in the same culture conditions could be observed after the rhG-CSF injection at 6 hours, but cloning efficiency was (0.50 +/- 0.11) x 10(-6) MNCs and showed statistical difference as compared with control. It is concluded that rhG-CSF to mobilize hemopoietic stem cells can be used to induce mouse MSCs in vivo expansion, which showed the peak value within 6 hours after final injection of rhG-CSF. rhG-CSF have the mini-mobilization effect on murine MSCs derived from bone marrow.
Animals ; Cells, Cultured ; Female ; Granulocyte Colony-Stimulating Factor ; pharmacology ; Hematopoietic Stem Cell Mobilization ; Mesenchymal Stromal Cells ; cytology ; Mice ; Random Allocation ; Recombinant Proteins

lerosis. MMP-3 may be an independent risk factor of atherosclerosis exclude TC and LDL

BACKGROUNDTransplantation of mensenchymal stem cells (MSCs) has been proposed as a promising way for tissue engineering. However, the application of MSCs for transplantation will undergo apoptosis due to the extremely harsh microenvironment such as excessive inflammation. Apigenin (API) has been reported to protect cells against inflammatory damage and cell death by exhibiting anti-inflammatory and anti-oxidative capacity. Here we investigated the modulatory effects of API in lipopolysaccharide (LPS)-mediated inflammation and apoptosis of MSCs, and further defined the underlying mechanism.

METHODSEffects of different concentrations of API (0, 5, 10, 20, 40 and 80 µmol/L) for 24 hours, and LPS (0, 0.5 and 5.0 µg/ml) for 6 hours and 24 hours on MSCs viability were assayed by MTT. Based on this, MSCs were pretreated with different concentrations of API (0 - 40 µmol/L) at the indicated times (6, 12 and 24 hours) followed by exposure to 5 µg/ml LPS for 24 hours. MTT, phase-contrast microscopy, annexinV/propidium iodide (PI) double stain flow cytometry (FCM) and Hoechst staining were applied to explore the effects of API on MSCs induced by 5 µg/ml LPS for 24 hours. In addition, reverse-transcription polymerase chain reaction (RT-PCR) was applied to detect the mRNA expression of pro-inflammatory factors including cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), nuclear factor-kappa B (NF-κB), pro-apoptotic gene caspase-3, Bad, and anti-apoptotic gene Bcl-2. Moreover, AutoDock software was used to imitate the docking score of API and vitamin D receptor (VDR). In parallel, Western blotting and RT-PCR were used to investigate protein and mRNA expression of VDR.

RESULTSMSCs stimulated with LPS 5 µg/ml for 24 hours was used as a model of apoptosis induced by over inflammatory stimulus. API (0 - 40 µmol/L) had non-toxic effect on MSCs; however, it could decrease mRNA expression of COX-2, iNOS and NF-κB at different time points in MSCs induced by LPS, except for API at the concentration of 5 µmol/L.

RESULTSfrom phase-contrast microscopy, MTT, Hoechst staining and AnnexinV/PI double stain FCM demonstrated that with the increasing concentrations of API and extension of administrating time, significant morphological changes of MSCs occurred, viability of cells was strongly inhibited, and meanwhile, apoptosis of LPS-administrated MSCs was exacerbated, compared with LPS individual group. In addition, API promoted caspase-3, Bad mRNA expression and inhibited Bcl-2 mRNA expression in a time-dependent and concentration- dependent manner. Further study found that pro-apoptosis effect of API was related to suppress VDR expression.

CONCLUSIONSAPI could inhibit the expression of inducible inflammatory factors, therefore exert the strong anti-inflammatory function. However, API could not protect MSC apoptosis induced by LPS but amplified the apoptosis. The apoptosis is related to Bad/Bcl-2 increasing and caspase-3 activation, which is mediated through suppressing VDR expression.

Animals ; Apigenin ; pharmacology ; Apoptosis ; drug effects ; Blotting, Western ; Cell Survival ; drug effects ; Cells, Cultured ; Flow Cytometry ; Lipopolysaccharides ; pharmacology ; Male ; Mesenchymal Stromal Cells ; cytology ; drug effects ; metabolism ; Rats ; Rats, Sprague-Dawley ; Receptors, Calcitriol ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction

The dialysis method was employed to load adriamycin into the micelles formed by temperature and pH sensitive polyhistidine-co-DL-lactide-co-glycolide-polyethylene glycol poly DL-lactide-co-glycolide-co-histidine (OLH-b-PLGA-b-PEG-b-PLGA-b-OLH). The critical micelle concentration (CMC) of the copolymer was measured with pyrene fluorescent probe method under different temperatures. The entrapment rate and drug-loading rate were determined with dialysis method. The diameter, morphology and surface potential of the copolymer micelles were investigated by corresponding instruments, respectively. The release behavior of adriamycin from copolymer micelles and the pH sensitivity were studied. The CMC of the copolymers ranged from 0.022 4 to 0.001 7 microg x mL(-1). The entrapment rate and drug-loading rate were 92.8% and 15.7%, respectively. The micelles have a mean diameter of (61.7 +/- 13.4) nm, and zeta potential was -9.88 mV. The in vitro adriamycin release rate increased with the pH dropping from 7.4 to 5.0. The results indicated that the CMC of the copolymers decreased as the raising of temperature, drug release behavior from the micelles possessed clearly pH sensitivity, and the copolymers may have a potential in targeted delivery system for anticancer drugs.
Doxorubicin ; administration & dosage ; chemical synthesis ; chemistry ; Drug Carriers ; Hydrogen-Ion Concentration ; Micelles ; Polyethylene Glycols ; chemistry ; Polyglactin 910 ; chemistry ; Technology, Pharmaceutical ; methods ; Temperature

Fingerprint technology is the key technology in modern Chinese medicine research, while spectrum-effect relationship research is the advanced stage of fingerprint research. Spectrum-effect relationship research can reveal the relationship between fingerprint and pharmacological effect through multiple statistical analyses, which can be used in Chinese medicine research. Spectrum-effect relationship has been used in many areas of Chinese medicine research, such as effective basis of single and compound Chinese medicine research, component compatibility research, processing mechanism research, pharmacological effect forecast research, technology optimization research, and so on. This paper systematically reviewed the application of spectrum-effect relationship in Chinese medicine research, and indicated some problems in spectrum-effect relationship research. At last, the authors give an outlook of the future of spectrum-effect relationship research.
Animals ; Biomedical Research ; China ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; Humans ; Spectrum Analysis

Adolescent ; Adult ; Aged ; Female ; Femoral Fractures ; diagnostic imaging ; surgery ; Fracture Fixation, Internal ; methods ; Humans ; Male ; Middle Aged ; Radiography