Biological Activity Of Fluorescent Probe YZP1 And Neuroprotective Effect Of AA13 And Related Mechanisms

The research is divided into two parts.The first part is to explore the biological activity of YZP1 in vitro and in vivo.The second part is to study the protective effect of AA13 and its related mechanism.The expression of COX-2 in inflammation and tumour environment is significantly increased,and local hypoxia often occurs in tumour microenvironment.Therefore,we designed a double-targeted probe YZP1 to explore new tracing methods for tumors and inflammation.Firstly,the biological activity of the probe was detected in vitro.The probe YZP1 was co-cultured with Hela cells.The survival rate of Hela cells was detected by MTT.The fluorescence was collected under confocal microscope.The results showed that YZP1 had no significant effect on cell growth and the fluorescence intensity of the probe was negatively correlated with the oxygen concentration,while positively correlated with the activity of COX-2.The lower the oxygen concentration,the higher the activity of COX-2,the stronger the fluorescence intensity of the probe.In organelle co-localization experiment,organelle-localization probes were co-localized with YZP1.It was found that the fluorescence coincidence of the probe with Golgi body and endoplasmic reticulum-localization probes was higher,indicating that the probe mainly played a role in these two organelles.The location of the probe is consistent with COX-2.In vivo imaging experiments,the mice with foot swelling as inflammation model,and the tail vein was injected with probe.The surrounding areas of breast tumors in nude mice were subcutaneously injected.And imaging was performed under the living imaging system respectively.The results showed that the probe could accurately locate the inflammation and tumor sites,and emit strong fluorescence signals.After the experiment,the mice were able to carry out normal life activities.The results of in vitro organ and tumor tissue imaging and H&E staining showed that the probe had a shorter metabolic time in vivo,and had no obvious damage to the main organs and no new inflammation.The probe YZP1 has good targeting ability in vivo and in vitro without toxic and side effects.The results of provide a reference for early diagnosis and precise resection of tumors in clinical practice.In the second part,the possible mechanism of neuroprotective effect of compound AA13 was studied.Our previous studies have shown that AA13 has potential anti-inflammatory and neuroprotective activities.Oxidative stress is one of the main causes of AD.Targeted autophagy therapy has recently become a promising strategy for many diseases including AD.In this experiment,SHSY5Y cells were injured by H2O2.The viability of SHSY5Y cells was detected by CCK method.The levels of Akt and p38MAPK proteins related to oxidative stress and autophagy proteins LC3 in AD-like environment were evaluated by Western Blotting.The results showed that the cell viability of SH-SY5Y cells exposed to H2O2 was decreased,and the cell viability was restored after pretreatment with AA13.Compared with those exposed to H2O2 only,the phosphorylation of p38 and Akt in H2O2-inducedSH-SY5Y cells decreased after pretreatment with AA13.AA13 also restored the ratio of autophagy-related protein LC3II to LC3I to normal level.This study suggests that AA13 can improve the survival rate of oxidative damaged cells induced by hydrogen peroxide and has potential neuroprotective activity.Its inhibition of the activation of p38MAPK pathway may contribute to the neuroprotective effect of AA13.Meanwhile,autophagy may be involved in the protective effect of AA13.Our results not only provide a new perspective for the neuroprotective effect of AA13,but also provide a new candidate compound for the prevention and treatment of neurodegenerative diseases.