The Metabolism Of Sodium Selenite And Selenium-enriched Yeast In Pig Tissues And The Effect Of Se Metabolites On The Proliferation Of Human Small Cell Lung Cancer Cell Line

To study the metabolism of dietary selenium-enriched yeast (SeY) and sodium selenite (SS) in pig tissues and the effect of selenium metabolites on the proliferation of human small lung cancer cell NCI-H446, four experiments were couducted.Exp 1. Effect of dietary SeY and SS on the antioxidant system and Se retention in pig liver and muscle.To study effect of dietary SeY and SS on the antioxidant system and Se retention in pig tissues,60 piglets were allocated into two experiments (eperimentl and experiment 2), with 3 treatments per experiment and 10 replicates per treatment. The 6 treatments were fed basal diet supplemented with 0,0.3,3.0 mg kg-1 SeY (period 1) and 0,0.3 and 3.0 mg kg-1 SS (period 2) for 8 wk, respectively. Results showed there was no influence of dietary SeY or SS level on pig performance. Compared with 0 mg kg-1 dietary 0.3 mg kg-1 SeY and SS remarkably (P< 0.05) increased GPX activity in liver, muscle and plasma, but there was no further increase of GPX activity in these tissues from 0.3 to 3.0 mg kg-1 dietary SS and SeY. There was no remarkable effect of dietary SeY and SS level on tissue SOD and GST activity. With the increase of dietary SeY, Se concentration in pig liver, muscle and plasma was all significantly (P< 0.01) increased. In the range of 0-3.0 mg kg-1 SS, Se concentration in liver and muscle were remarkably increased (P< 0.01), while plasma Se reached plateau at the level of 0.3 mg kg-1. These suggested there was no effect of dietary SeY and SS on pig performance and tissue antioxidant system within 8 wk, but tissue Se level could be increased by the two Se in this period.Exp 2. Influence of dietary SS and SeY on the distribution of selenocompounds in pig liver and muscle.There are two forms of Se in mammal tissues, protein-bound selenium (PBS) and nonprotein-bound selenium (NBS). The results of Exp 1 indicated dietary SeY and SS increased Se concentrations in both liver and muscle. To study the effect of dietary SeY and SS on the distribution of NBS and PBS, one procedure for separating NBS and PBS in pig tissues was firstly developed, and then selenocompounds in pig liver and muscle obtained from Exp 1 was separated with this procedure. Results showed the NBS and PBS content in both liver and muscle increased (P< 0.05) with dietary SeY and SS. The relative percentage of PBS amount to total Se in liver decreased (P < 0.05) with the increase of dietary SeY, and that of NBS amount to liver Se increased (P< 0.05), accordingly. In contrasted to liver, the relative percentage of NBS and PBS in pig muscle kept consistent at 6% and 94%, respectively. Similar with SeY, the relative of NBS and PBS in the liver of pig fed SS increased (P< 0.05) and decreased (P< 0.05), respectively, with increase of dietary SS level. The distributing pattern of NBS and PBS in the muscle of pigs fed SS was similar with that in liver. These suggested that the distribution of NBS and PBS were similarly affected in liver and differently affected in muscle by dietary SeY and SS.Exp 3. Effect of LE containing NBS from the liver of pig fed excess SeY and SS on the proliferation of human lung cancer cell line.The results of Exp 2 indicated that NBS concentrate and relative percentage in pig liver was increased by ditary SeY and SS, and the distribution of NBS and PBS was similar in liver of pigs fed SS and SeY. To study whether there was potent anticancer potential of NBS, livers from pigs fed basal diet and the diet plus 3.0 mg kg-1 SS and 3.0 mg kg-1 SeY for 16 wk were homogenated, then the liver homogenate was precipitated with chloroform and ethanol. The superant was freez-dried, condensed, then three LE containg NBS was obtained. The LE from pigs fed basal diet, plus SeY and SS Se was expressed as LEB, LEY and LES, which contained 0.98μM,2.62μM and 7.84μM Se. After the addition of LEB, LEY and LES into cell culture medium, results showed that cell viability were decreased by LEY and LES when compared with LEB. The mechanism of LES and LEY inhibiting cancer cell was via the induction of DNA fragmentation. Sodium selenite is the standard material for evaluating the relative anticancer potential. After the balance of Se content between LEB and LEY by addition of SS into LEB, the inhibition effect of LEY was still higher than that of LEB. This is also true for LES. These suggested that there were powerful anticancer NBS in the liver of pigs fed excess SeY and SS.Exp 4. Effect of pig serum on the proliferation of human lung cancer cell The results of Exp 3 demonstrated there was potential cancer-preventive NBS in the liver of pigs fed 3.0 mg kg-1 SS. To study whether there was cancer cell suppressing selenocompounds in the serum of pigs fed 3.0 mg kg-1 SS and SeY, blood were collected via vena cava anterior and serum was separated and filter with 0.22 uM membrane. Se concentration in serum of pigs fed basal diet and supplemented with 3.0 mg kg-1 SeY and SS was 165,1316 and 834 ng ml-1, that's Se(deficient), SeY(excess) and SS (excess). To proquest the opitimal addition level of pig serum to cell medium in place of FBS,7 graded levels (8-20%) of pig serum of Se(deficient), SeY(excess) and SS(excess) were added. The optimal addition level of pig serum was 17.79%,15.39% and 14.31%, respectively based on relation between the cell MTT OD value and serum level, and the average 14.31% was chosen as the optimal level. The optimal culture time was 144 h based on cell morphology. After the addition of 15.83% pig serum to cell culture for 144h results showed the MTT OD value of cell cultured with Se (deficient) was significantly higher than that of cell treated with SS(excess), while the MTT OD value of SeY(excess) treatment cell was between Se (deficient) and SS (excess). Compared with SeY (excess) and SS (excess), Se(deficient) treat cell had lower (P< 0.05) LDH activity in medium, while no difference between SeY(excess) and SS(excess) treated cells. After stained with Annexin V and PI and analyzed with cytometry, cell apoptotic rate was significantly lower (P< 0.05) in Se (deficient) than in Se (excess) treated cell, while the treatment of SeY (excess) was still between Se (deficient) and SS (excess) and no difference was observed.In summary, we found the Se content in the liver and muscle of pigs fed SeY or SS ranging from deficient to excess significantly increased with dietary Se level, but the NBS content and relative perentage increased with dietary SeY and SS level after NBS was separated from PBS. Furthermore, the distribution pattern of NBS/PBS in liver was similar. So, we concluded there would be anticancer NBS in pig liver based on the above finding and the potent anticancer mechanism of Se. After the addition of extracted NBS into lung cancer cell media, we found there were powerful anticancer NBS in the pig liver. Since blood was the carrier from liver to the peripheral tissue, we also suspect there was anticancer selenocompound in the serum of pig fed excess SeY and SS. With serum pharmacology, we found there was anticancer selenocompound in the serum of pig fed 3.0 mg/kg Se as SS.