Objective To study the estradiolNGF regulatory cascade in retinal endothelial cells. Methods Retinachoroids vascular endothelial cell line RF/6A from rhesus was cultured in M199 medium contain 20% FBS.mRNA of ER,NGF and its receptors,TrkA and P75 NTR were detected with RTPCR.The cells were incubated with NGF,VEGF,antibodies against NGF or VEGF,and K252a(100nmol/L),the specific inhibitor of TrkA,separately or in different combination.MTT based cell counting assay was used to study the viability of the cells.The apoptosis was evaluated by FACS,mass migration by wound healing assay,and tubogenesis by AngioMatrix assay. Results We amplified the specific fragments of cDNA of ER,NGF and NGF receptor TrkA using RTPCR.10?nmol/L1??mol/L estradiol augments the proliferation and increases the viability of RF/6A in a dosage dependent manner.In the wound healing based migration assay,we found the similar alteration.This effects of estradiol was partially blocked by NGF neutralized antibody and K252a.Apoptosis rates were at the similar level among the groups.For the tubogenesis of RF/6A,we found no augmentation by NGF,and no blocked augmentation by estradiol.Conclusion NGF,first identified as the survival factor of nerve system,now also seemed to be an activator of retinal endothelial cell,is under the regulation of estrogen.The proliferation and migration,but not the tubogenesis of retinal endothelial cells are regulated by this regulatory cascade.

Objective To explore the roles of spinal microglial Src-family kinases (SFKs) in diabetic neuropathic pain. Methods Male Sprague-Dawley rats, weighing 200 ~ 220 g, were used in the experiments. The level of p-SFKs in spinal dorsal horn was detected by single immunofluorescence staining on day 7th , 14th and 28th after intraperitoneally injection of Streptozotocin (STZ) and its location was detected by double immunofluorescence staining. The changes of 50% paw-withdrawal thresholds of rat were detected by behavioral tests when PP2 , a specific inhibitor of SFKs , was intrathecally administered before intraperitoneally injection of STZ. Results Compared with vehicle group, the blood glucose level increased on day 1 (P < 0.001) and the hyperglycemia persisted at least for 28 days (P < 0.001) after intraperitoneally administered of STZ (50 mg/kg). Paw-withdrawal threshold (PWT) decreased gradually from day 1 (P < 0.05) and reached the minimum on day 28 (P < 0.001) after STZ administration. Meanwhile, the expression of p-SFKs in spinal dorsal horn markedly increased on day 7 (P < 0.01), day 14 (P < 0.01) and day 28 (P < 0.01). The p-SFKs was mainly co-localized only with microglia , but not with neurons or with astrocytes. Intrathecally administered of PP2 before STZ reversed STZ-induced mechanical hyperalgesia. Conclusion Microglial SFKs in spinal dorsal horn maybe play a pivotal role in diabetic neuropathic pain.

AIM:To investigate the effects of P2X4 receptor on peri-sciatic administration of recombinant rat TNF-α(rrTNF)-induced mechanical allodynia.METHODS:Male Sprague-Dawley rats (180~200 g) were used in the experiments.The levels of P2X4 receptor on day 3, day 7 and day 14 after peri-sciatic administration of rrTNF were exam-ined by Western blot, and the location of P2X4 receptor in the spinal dorsal horn was observed by double immunofluores-cence staining.The changes of 50%paw-withdrawal thresholds of the rat were detected by behavioral test, and the level of TNF-αin the spinal dorsal horn was also examined by Western blot when TNP-ATP was intrathecally injected before the ad-ministration of rrTNF.RESULTS:Compared with control group, the expression of P2X4 receptor in the spinal dorsal horn on the ipsilateral side significantly increased on day 3, day 7 and day 14 (P<0.01) after rrTNF (100 ng/L) administra-tion.P2X4 receptor was co-localized only with microglia, but not with neurons or astrocytes.Intrathecal injection of TNP-ATP before rrTNF administration prevented mechanical allodynia induced by rrTNF and inhibited the upregulation of TNF-αin the spinal dorsal horn.CONCLUSION:P2X4 receptors in microglia may be involved in rrTNF-induced mechanical allodynia by the upregulation of TNF-αin the spinal dorsal horn.

Based on the demand of enterprise talents and the characteristics of manufacturing process management in biotechnology, in order to make the students acquire the ability to solve complex engineering problems in the production process, we developed a "Comprehensive Biotechnology Experiment" course, where two-step enzymatic production of l-aspartate and l-alanine were the key processes. In this course, we drew lessons from the site management of the production enterprise, performed the experimental operation mode of four shifts and three operations. The content of this course includes principles, methods and experimental techniques of several core curricula and the site management mode of enterprises. As to the evaluation, the summary of the experimental staff's handover records and the content of teamwork were examined and scored. Through teaching practice and continuous improvement, we developed a complete experimental teaching process and assessment mechanism. Overall, the Comprehensive Biotechnology Experiment course achieved good teaching effect, which may serve as a reference to promote the development of experimental teaching of biotechnology.
Humans ; Biotechnology ; Curriculum ; Students