1.Effect of activated microglia induced by LCN2 on depression pathogenesis in rats
Zilu JIANG ; Fen WANG ; Tao XIE ; Weiyan LI
Journal of Medical Postgraduates 2015;28(10):1038-1042
Objective Lipocalin-2(LCN2) can promote the M1 approach of microglia.The study was to explore the effect of activated microglia induced by LCN 2 on depression pathogenesis in rats and its mechanism . Methods According to the chronic stress depression model created by Willner , 40 adult male SD rats were divided into 4 groups(n=10): normal control group;UCMS group;UCMS+LCN2 siRNA group;LCN2 siRNA control group .A series of stress stimulation was given on UCMS group and UCMS +LCN2 siRNA group for 21 days to create depression model .At 7 days after the stress stimulation , the rats in UCMS+LCN2 siRNA group and LCN2 siRNA control group were anaesthetized by 0.4mg intraperitoneal injection of 10% chloral hydrate , followed by intrathecal injection of LCN2 siRNA(0.015 μL/g, 3 times a week) to the rats till the end of the stress (21 days).At the same time, the same volume of isotonic saline was given to normal control group and UCMS group .The weight of rats was measured every week and the sucrose preference and the forced swimming test were applied to measure behavior of the rats after the experiment .The hippocampus of the rats were extracted and immunofluorescence and western blot were applied to detect the expressions of microglia specific markers:LCN2 and the Iba. Results At 3 week, the weight of rats in UCMS+LCN2siRNA group was higher than that of UCMS group ([262.82 ±0.01]g vs [179.98 ±0.08]g, P<0.05).The weight of rats in normal control group and LCN2 SiRNA control group increased significantly higher than the other two groups (P<0.05).The sucrose preference values of normal control group (0.82 ±0.01),UCMS+LCN2 siRNA group(0.81 ±0.01) and LCN2 siRNA control group(0.82 ±0.01) were higher than that of UCMS group (0.25 ±0.04) (P<0.05).The fixed time of the forced swimming test of UCMS+LCN2siRNA group decreased significantly compared with UCMS group ([4.64 ±0.8]s vs [23.11 ±2.63]s, P<0.05).The LCN2 expression of UCMS group was significantly greater than the other groups (P<0.05).The Iba1 expression in the hippocampus of the UCMS group increased significantly compared with other groups . Conclusion LCN2 is associated with the pathogenesis of de-pression induced by chronic stress reaction and is mechanism may be related to the activation of microglia in the central nervous system of rats.
2.Effect of spinal cord lipocalin-2 on development of morphine tolerance in normal rats
Fen WANG ; Qingzhen LIU ; Jian LIU ; Zilu JIANG ; Tao XIE ; Weiyan LI
Chinese Pharmacological Bulletin 2015;(4):565-569
Aim To explore the effect of knockdown spinal cord LCN2 by RNAi on the development of mor-phine tolerance in normal rats.Methods After suc-cessful intrathecal implantation, fourty-eight male Sprague-Dawley rats weighing 1 80 -220 grams were randomly divided into 4 groups (n =1 2):group I:control group,group II:morphine tolerance group, group Ⅲ:mismatch siRNA group,group IV:LCN2 siRNA group.The sixth day after intrathecal implanta-tion,rats were tested to ensure the position of cathe-ters,and it was recorded as d 0.On d 2 -8,rats were subcutaneously (s.c)injected of normal saline (NS) (group I)or morphine (group Ⅱ,Ⅲ,Ⅳ)1 0 μg· g -1 twice a day at 8:00 and 1 6:00.Before everyday s. c injection,rats were intrathecally injected of 1 0 μL DEPC solution (group Ⅰ,Ⅱ),1 0 μL DEPC solution containing 4 μg mismatch siRNA (group III)and 4 μg LCN2 siRNA solution (group IV).Paw withdrawal la-tencies to thermal stimuli (PWTL)were tested before morphine injection and 45 minutes after morphine in-jection on d 1 and d 9.The percentage of maximal pos-sible effect (% MPE)was calculated later.Animals were sacrificed on d 9 after the behavioral test and the lumbar enlargement segments of the spinal cord were removed for detecting the expression of phosphorylated-p38 mitogen-activated protein kinase (p-p38 MAPK) and LCN2 by Western blot and microglia marker Iba1 by immunofluorecence.Results On d 1 ,there was no significant difference in %MPE among four groups. On d 9,compared to group Ⅰ,%MPE was signifi-cantly reduced (P <0.05)while p-p38MAPK,LCN2 and Iba1 were markedly up-regulated in group Ⅱ andⅢ (P <0.05 ).On d 9,compared to group Ⅱ,%MPE was significantly increased while p-p38MAPK, LCN2 and Iba1 were markedly reduced in group IV (P<0.05).Conclusion Using LCN2 siRNA to knock-down spinal LCN2 relieves the development of mor-phine tolerance in normal rats possibly through inhibi-ting the activation of microglia and p38 MAPK in the spinal cord.
3.Construction of the siRNA expression vector and the silencing influence on focal adhesion kinase gene of Tca8113 cell line
Hualian LIU ; Hongbing JIANG ; Shuzhong XING ; Laikui LIU ; Zilu WANG ; Yangyu ZHENG
Journal of Practical Stomatology 2001;0(01):-
Objective:To construct the siRNA expression vector of focal adhesion kinase(FAK) gene and inhibit the expression of FAK gene in tongue cancer cell line Tca8113 by RNA interfering technique. Methods:According to the encoding sequence of FAK mRNA, two pieces of oligonucleotide sequences were designed and synthesized. The annealed oligonucleotide fragments were subcloned into pGCSilencerTM-U6/Neo siRNA expression vector. After being identified by restriction enzyme method, the recombinant pSilencer-FAK plasmids were transfected into Tca8113 cells. The transfected cells were selected by G418 method. Immuocytochemistry and Western blotting were used to evaluate FAK gene silencing efficiency. Results:The oligonucleotide fragments were correctly inserted into pGCSilencerTM-U6/Neo vector. FAK expression of the transfected cells was significantly down-regulated by pSilencer-FAK. Conclusion:The siRNA expression vector of FAK is successfully constructed and FAK expression of Tca8113 cells can be inhibited by RNA interfering technique.
4.The Role of Signal Transducer and Activator of Transcription 3 in Bone Homeostasis and Load-Driven Bone Remodeling
Yiwen CUI ; Zilu WANG ; Weiqi LI ; Yingqi CHENG ; Zhiyun YE ; Xinyi GONG ; Siru ZHOU ; Yiling YANG ; Lingyong JIANG
Journal of Medical Biomechanics 2021;36(5):E818-E823
Bone homeostasis is a relative balance between bone formation and resorption. Signal transducer and activator of transcription 3 (STAT3), which is closely related to bone homeostasis, takes part in multiple intracellular and extracellular signal pathways. STAT3 participates in the process of osteoblast differentiation regulated by several factors. It can also maintain bone homeostasis by regulating the recruitment, differentiation and activation of osteoclasts. In addition, STAT3 is involved in the interaction between osteoblasts and osteoclasts. Patients with STAT3 mutations can have several inherited bone metabolism diseases. Furthermore, STAT3 plays a critical role in load-driven bone remodeling. Mechanical stimulation promotes osteoblast differentiation and bone formation through activating or enhancing STAT3 expression during bone remodeling process. This review summarizes the participation of STAT3 in maintaining bone homeostasis together with its possible mechanisms and discusses the connection between STAT3 and mechanical stimulation in bone remodeling, so as to provide a potential pharmacological target for the treatment of bone diseases.