Inhibitory effect and mechanism of metformin on human ovarian cancer cells SKOV-3 and A2780.

OBJECTIVE: Ovarian cancer is the most common malignant tumor in female reproductive system. Metformin is an orally taken hypoglycemic agent, which is extensively applied in the clinic. Clinical trials find that there may be a certain degree of action of the metformin in inhibiting malignant tumors. This paper aims to investigate the inhibitory effect and mechanism of metformin on human ovarian cancer cells. MATERIALS AND METHODS: Through in vitro cell experiment, the influences of metformin on the proliferation, colony formation and apoptosis of ovarian carcinoma cells were studied. Ovarian cancer cells SKOV-3 and A2780 in logarithmic growth phase were selected and cell proliferation was measured by MTT method. The metformin was processed for 48 h to calculate the survival rate of cells. Also, metformin was processed for 24 h and two weeks or stained with crystal violet, after which Quantity One (Bio-Rad, Hercules, CA, USA) method was used to quantitatively analyze the cell clone formation, meanwhile, the FCM (flow cytometry) was used for the detection and analysis. RESULTS: Intervened by metformin with different concentrations for 48 h, the cell viabilities of SKOV-3 and A2780 cells were respectively reduced by 19.49 ± 2.92%, 45.41 ± 7.95%, 53.84 ± 5.53%, 64.04 ± 4.36% and 11.45 ± 3.12%, 35.42 ± 7.55%, 43.77 ± 5.77%, 53.05 ± 5.55% as compared with that in the control group with statistical significances. After processed by metformin with different concentrations for two weeks, the cells clone numbers of SKOV-3 and A2780 were significantly reduced. Treatment of metformin on SKOV-3 and A2780 cells of human ovarian cancer showed significant apoptosis. CONCLUSIONS: The metformin has the inhibitory effect on the cells of human ovarian cancer, which may be through inducing ovarian cancer cell apoptosis.

Importance of CD44 on umbilical cord mesenchymal stem cells for expansion of hematopoietic cells.

Human umbilical cord mesenchymal stem cells (hUCMSCs) have important functions on the expansion of hematopoietic stem cells (HSCs) through providing the essential microenvironment for hematopoiesis. In order to test whether CD44 on hUCMSCs could have a key function for the ability of hUCMSCs to expand human HSCs, the soluble anti—CD44 antibody was added to the co—cultures of hUCMSCs and cord blood (CB) CD34+ cells, which blocked the ability of hUCMSCs to expand CB CD34+ cells significantly. Long—term culture initiating cell (LTC—IC) assay revealed that the ability of multipotent differentiation of CB CD34+ cells co—cultured with CD44 knockdown hUCMSCs could only retain lasting at most for 5 weeks in vitro. In vivo assay, based on non—obese diabetic/severe combined immunodeficient disease (NOD/SCID) mice, revealed that the hematopoietic reconstitution potential of CB CD34+ cells co—cultured with CD44 knockdown hUCMSCs is significantly reduced. The hematopoietic supporting ability of hUCMSCs in vivo and in vitro is reduced upon the knockdown of CD44. CD44 has important functions on the ability of hUCMSCs to expand human HSCs in the cell— extrinsic control.

Effect of insulin on oxygen free radicals and oxidative phosphorylation in liver mitochondria of diabetic rats.

AIM: To observe the effects of insulin on liver mitochondrial respiratory function, activity of H+-ATPase, and superoxide anion free radicals production in diabetic rats. METHODS: Rats were injected iv with alloxan 40 mg/kg to induce diabetes. The liver mitochondrial respiratory function was assayed by measurement of oxygen consumption using a Clark oxygen electrode. Superoxide anion production was assayed using chemiluminescence method. Activities of H+-ATPase were measured by luciferin-luciferase system and inorganic phosphorus's method. RESULTS: Insulin 1 U/kg sc daily for 9 weeks improved oxidative phosphorylation, respiratory rate state 3 (P < 0.05), respiratory control ration (P < 0.01), and ADP:O ratio (P < 0.01), but there were no obvious effect on respiratory rate state 4 (P > 0.05). In the insulin group, synthesis activity of H+-ATPase was obviously increased (P < 0.05) and hydrolytic activity of H+-ATPase was remarkably decreased (P < 0.01), compared with the diabetes group. Insulin 1 U/kg for 9 weeks apparently decreased the production of O2.- (P < 0.01) in liver mitochondria of diabetic rats. CONCLUSION: Insulin can prevent the injury from superoxide anion in liver mitochondria, and improve the function of the liver mitochondria oxidative phosphorylation.

Rapid assay of A1c-type glycosylated hemoglobin in blood of diabetic rats using fast protein liquid chromatography.

AIM: To develop a fast protein liquid chromatography (FPLC) method for the assay of Alc-type glycosylated hemoglobin in diabetic rats. METHODS: Venous blood was collected from rats. After the erythrocytes were washed and sedimented, the hemolysate was prepared and diluted with isotonic saline as a sample for the assay. Samples were then separated on Mono_S_HR_5/5 cation exchange column by a lithium chloride gradient elution system. The Alc-type glycosylated hemoglobin was monitored by measuring the absorbance at 415 nm. RESULTS: The Alc-type glycosylated hemoglobin was well separated from total hemoglobin. The average HbAlc amount in diabetic rats was determined to be (3.6% +/- 0.6%, n = 7), while the corresponding figure in normal rats was (1.4% +/- 0.4%, n = 7, P < 0.01). CONCLUSION: This FPLC method is easy, rapid and reproducible, and can be used for an assay of diabetic rat HbAlc to evaluate and screen new drugs for diabetes mellitus therapy.

[Studies on the saponin constituents of jiu jielong (Ardisea pusilla)].

Two saponins were isolated from Ardisea pusilla A. DC. which have improved immunological function and antitumor activity. Their structures were determined on the basis of chemical and spectral analysis (IR, MS, 1HNMR, 13CNMR and two new NMR technique). The two compounds were 3-O-[alpha-L-rhamnopyranosyl-(1-->2)-beta-D-glucopyranosyl-(1-->3)] [beta-D-glucopyranosyl-(1-->2)]-alpha-L-arabinopyranosyl cyclamiretin A (I) and 3-O-[beta-D-xylopyranosyl-(1-->2)-beta-D-glucopyranosyl-(1-->4)] [beta-D-glucopyranosyl-(1-->2)-beta-D-glucopyranosyl-(1-->2)]-alpha-L- rhamnopyranosyl cyclamiretin A (II). They were new saponin and named as ardipusilloside I and ardipusilloside II.

[Selective long-range DEPT NMR technique and its application to structural study of glycoside].

This paper reports a new NMR technique: selective long-range DEPT. The result obtained showed that the technique can be used to assign NMR signal of 13C, 15N and 31P nuclei, and connect the spin systems separated by quaternary carbon or heteroatom. This paper deals mainly with applying the technique for determining the glycosidation position in aglycone moiety and sequence of sugar moiety in a natural glycoside.

[Chemical structure of glutinic acid, a new diterpenoid from Caryopteris glutinosa].

A new diterpenoid compound named glutinic acid had been isolated from Caryopteris glutinosa Rehd by liquid chromatography on Al2O3 and silica gel column. The structure of glutinic acid was elucidated on the basis of IR, UV, MS, H1-NMR, 13C-NMR, DEPT and 13C-1H-COSY spectral analyses.

[Studies on the chemical constituents from Aralia elata].

Eight compounds have been isolated from the root bark of Aralia elata. Their structures have been identified by means of physico-chemical and spectral analysis. They are (6'-O-palmitoyl)-beta-sitosterol-3-O-beta-D-glucoside (A5), silphiosideA (A9), chikusetusaponin Ib (A11), araloside A (A12), araloside C (A15), acanthoside D (B1). Compound A10 is a new natural product, named as araloside A methyl-ester. 3-O-beta-D-glucopyranosyl (1----3)[beta-D-glucopyranosy (1----4)]-beta-D-glucopyranosyl-oleanolic acid-28-O-beta-D-glucopyranoside (A16) is a new compound, named as araloside G. Compounds A5, A9, A10, A11, A16, and B1 were isolated for the first time from the plant. 13C-NMR chemical shifts of compounds A9 and A15 were assigned for the first time.

[1H, 13C NMR and stereochemistry studies of 3S-substituted alkoxylquinuclidine by two-dimensional and NOE difference NMR techniques].

By using optically pure 3S-quinuclidinol, two diastereoisomers (3S-1) and (3S-2) of 3-(substituted) alkoxy-quinuclidine were synthesized. 1H and 13C NMR spectra of (3S-1) and (3S-2) have been completely analyzed utilizing double-quantum filtered COSY, 13C-1H COSY and NOE difference experiment. The NOE difference experiment was used to determine the absolute configuration at C-11 of the diastereomers. According to the results of NOE difference and variable concentration NMR experiments, the configuration about C-11 of (3S-1) is designated as R (1A), whereas the configuration about C-11 of (3S-2) is designated as S (1B). The result was confirmed by X-ray diffraction analysis.

[The structure determination of a new alkaloid phlegmariuine-N by long-range two-dimensional and NOE difference NMR spectroscopy].

A new alkaloid, phlegmariuine-N, was isolated from Phlegmariurus fordii (Baker) Ching. In the present work, a technique of long-range 1H-1H and 13C-1H COSY in combination with NOE difference spectroscopy has been successfully used to connect the spin systems separated by quatermary carbons and heteroatom. Its structure has been elucidated as (I).