Ethnopharmacological relevance: Fuzi-Lizhong pill (FLZ) is a traditional Chinese medicine for treating patients with Spleen Yang deficient syndrome. Ghrelin, a peptide with 28 amino acid residues, plays multiple roles in thermogenesis. This study aims to explore FLZ regulating ghrelin to compensate hypothermia in rats with hypothyroid and indigestion.Materials and methods: In litter-matched rats, hypothermia was developed with both thyroidectomy at d(1) and interscapular brown adipose (IBA) removal at d(42), indigestion was induced with both high fat diet and fasting-feeding cycle from d(56); the littermates with hypothermia and indigestion were administrated with FLZ from d(70). Adaptive thermogenesis, thyroid hormones, metabolites, ghrelin dynamics were measured at d(98).Results: The results showed that plasma ghrelin levels were inversely correlated with the gastric ghrelin levels and adaptive thermogenesis in rats undergone both thyroidectomy and IBA removal. Fatty diet and FLZ enhanced the increase of plasma ghrelin of hypothyroid rats. These were supported by the changes of plasma thyroid related hormones, plasma metabolites, gastric ghrelin mRNA and protein, and the effects of fatty diet or FLZ.Conclusions: Our results suggest that more ghrelin release compensate chronic hypothermia in rats with both hypothyroidism and indigestion. It could explain the mechanisms of FLZ in relieving chronic hypothermia. (C) 2013 Elsevier Ireland Ltd. All rights reserved.

Potent effects of Flt3 ligand (FL) on the development of the immune system have generated much interest in application of FL in cancer immunotherapy.Objective: To evaluate the effects of Pichia pastoris secreted rhFL on the growth of mouse EL-4 lymphoma and C26 colon adenocarcinoma injected in syngeneic mice for the first time.Methods: Mice were placed into one of two treatment groups. 2 x 10(5) EL-4 or C26 cells were injected subcutaneously (SC.) into mice on day 0. Group 1 received subcutaneous PBS injections from Day -7 to Day 14 and group 2 received subcutaneous rhFL injections at 30 mu g/day from Day -7 to Day 14. Serial tumor areas were measured. On Day 22, mice from each group were sacrificed, and weight of tumors and spleens were evaluated. Data analysis used Student t tests.Results: Pichia pastoris secreted rhFL resulted in tumor growth delay for both EL-4 lymphoma and C26 colon adenocarcinoma compared with control (P < 0.01). Tumors from rhFL-treated mice were smaller (P < 0.01) than controls while spleens larger (P < 0.01) than controls. Histological examination of tumor sections revealed an obvious increase in regions composed largely of infiltrating cells in the rhFL-treated tumors. Infiltrating cells could be detected in clusters among tumors from mice treated with rhFL whereas these cells were only occasionally detected in sections of control tumors.Conclusion: treatment of rhFL expressed from Pichia pastoris resulted in an antitumor response against EL-4 and C26 tumors injected in syngeneic mice.

OBJECTIVE:To study the expression of hHSF in E. coli and its effect on the mobilization of hematopoietic stem/progenitor cells.METHODS:The hHSF gene was obtained by overlapping PCR and cloned into the vector pET30a to yield pET30a-hHSF, which was transformed into E. coli BL21(DE3) and expressed with IPTG induction. Subsequently, rhHSF was purified by gel filtration and cation exchange chromatography and subjected to refolding. Molecular weight of hHSF was measured by MALDI-TOF Mass Spectroscopy. The N terminal amino acid sequence rhHSF was determined by protein sequencing. rhHSF was profiled in rhesus monkey for mobilization of peripheral blood stem cells. Eight rhesus monkeys were equally divided into two groups. The first group was administered single subcutaneous injection of 500 microg/kg hHSF, while the other one was administered 10 microg.kg(-1).d(-1) G-CSF for 4 days followed by a single subcutaneous injection of 500 microg/kg rhHSF.RESULTS:The sequence coding hHSF was confirmed by sequencing and the induced-expression level was about 30% of total cell proteins. The purity of target protein was over 95%. The sequence of N terminal 10 amino acids and the amino acid composition were consistent with the theoretical parameters; molecular weight of rhHSF was 7540. The peripheral CD34(+) cells, CFU-GM yields, and neutrophils peaked at 3 h (16.3-folds increase compared with baseline), 1 h (1.9-folds increase) and 45 min (4.4-folds increase) respectively after the single injection of rhHSF. The addition of rhHSF after the last dose of G-CSF boosted these levels to 25.8-folds, 8.7-folds and 8.3-folds respectively.CONCLUSION:hHSF is highly expressed in E. coli and rapidly mobilizes the hematopoietic stem/progenitor cells and neutrophils in rhesus monkeys. hHSF shows distinct synergistic effect with G-CSF.