Objective To screen the optimal medium formula for obtaining high proliferation rate and differentiation rate of protocorm-like body ( PLB) of Dendrobium officinale Kimura et Migo ( D.officinale) . Methods We observed the effect of different concentrations of phytohormones such as 6-benzylaminopurine (6-BA) , 1-naphthaleneacetic acid ( NAA) , 2,4-dichlorophenoxyacetic acid ( 2,4-D) and kinetin ( KT) , or their combinations together with organic additives such as potato extractive ( PE) , banana extractive ( BE) , apple extractive ( AE) and coconut milk ( CM) on the proliferation and differentiation of PLB of Dendrobium officinale Kimura et Migo. Results 6-BA, NAA and KT at certain concentrations were beneficial to the proliferation and growth of PLB, and when the concentration of KT was 1.0 mg/L, 50-d PLB proliferation times reached to 7.20. 2,4-D had obvious inhibitory effect on PLB proliferation and growth. The combination of 6-BA, NAA and KT showed stronger effect on PLB proliferation than the phytohormones used alone, and the combination of 0.5 mg/L 6-BA, 1.0 mg/L KT and 1.0 mg/L NAA had the best effect on PLB proliferation, 50-d PLB proliferation times reaching to 9.52. Certain concentrations of PE and AE could promote PLB differentiation, and 100 g/L PE brought well-grown seedling in test tube and the highest differentiation rate, being up to 92.6% . Conclusion The optimal hormone matching and additives concentration on proliferation and differentiation of PLB of Dendrobium officinale Kimura et Migo have been obtained, which can lay a foundation for its rapid propagation and artificial seed preparation.

Objective:To investigate the role and mechanism of splenic myeloid-derived suppressor cells (MDSCs) in sepsis-induced adrenal injury (SAI).Methods:Thirty male C57 mice aged 6-8 weeks were randomly divided into normal control group ( n = 5), sham operation group (Sham group, n = 5), sepsis model group [cecal ligation and perforation (CLP) group, n = 10] and sepsis+splenectomy group (CLPS group, n = 10). The sepsis model of mice was reproduced by CLP method. In Sham group, only the cecum was opened and separated, then closed, without CLP. In CLPS group, the spleen was removed before CLP. In normal control group, no challenge was given. After 24 hours, the rats were sacrificed by anesthesia, and peripheral blood, spleen, bone marrow, and bilateral adrenal glands were harvested. The pathological of adrenal gland was assessed by hematoxylin-eosin (HE) staining under optical microscope. The ratio of MDSCs in peripheral blood, spleen and bone marrow was determined by flow cytometry. The expressions of MDSCs surface antigen CD11b, Gr-1 and interleukins (IL-6, IL-1β) mRNA in adrenal tissue were measured by real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR). Western Blot was used to detect the expressions of mammalian rapamycin target protein (mTOR) pathway related proteins including total mTOR (T-mTOR), phosphorylation of mTOR (p-mTOR) and caspase-3. Results:The adrenal cortex and medulla of the normal control group and Sham group were intact and the structure was clear under optical microscope, while in the CLP group, the adrenal gland showed edema, cortical hemorrhage and cell edema. Compared with the CLP group, the adrenal tissue injury was significantly reduced in the CLPS group. Compared with the normal control group and Sham group, MDSCs ratio in the peripheral blood was significantly increased and significantly reduced in the spleen in the CLP group, but there was no significant difference in bone marrow, the expression levels of CD11b, Gr-1, IL-6, IL-1β mRNA and caspase-3 protein were increased significantly and p-mTOR protein expression was significantly decreased in adrenal tissue, there was no significant difference in the expression of T-mTOR protein. Compared with the CLP group, in the CLPS group, the MDSCs ratio in the peripheral blood was significantly decreased (0.143±0.011 vs. 0.324±0.023, P < 0.01), the expression levels of CD11b, Gr-1, IL-6 , IL-1β mRNA and caspase-3 protein in adrenal gland were significantly decreased [CD11b mRNA (2 -ΔΔCt): 2.90±0.56 vs. 5.74±0.13, Gr-1 mRNA (2 -ΔΔCt): 2.71±0.14 vs. 4.59±0.46, IL-6 mRNA (2 -ΔΔCt): 2.44±0.64 vs. 5.17±1.04, IL-1β mRNA (2 -ΔΔCt): 3.58±0.52 vs. 4.44±0.26, caspase-3 protein (caspase-3/GAPDH): 0.05±0.01 vs. 0.13±0.02, all P < 0.01], the p-mTOR protein expression was significantly increased (p-mTOR/GAPDH: 0.61±0.11 vs. 0.27±0.04, P < 0.01). Conclusions:The spleen is the major source of MDSCs in SAI. Splenectomy can attenuate SAI by reducing mobilization of MDSCs and activating the mTOR signaling pathway.