Expression Profiles Of Cathepsins From Infected Mice With Angiostrongylus Cantonensis And The Cloning Of The Gene Coding AcHTCP

Angiostrongyliasis cantonensis, which is caused by Angiostrongyluscantonensis (Ac), can lead to diseases in central nervous system. Its intermediatehost is mollusc, including ampullaria gigas, and rat as its definitive host. Human,like mice, is the non-permissive host. Man can be infected with Ac by eatingundercooked snails.Angiostrongyliasis cantonensis is very popular in Southeast Asia and Pacificislands. Since the first case in Taiwan1945, this disease is appearing in mainland.Ministry of health listed Angiostronglyliasis cantonensis as new born communicabledisease. At present, the diagnosis of the disease was based primarily on contacthistory, clinical symptoms and ELISA testing result. The golden standard to diagnosethe disease is to find worm in cerebrospinal fluid (CSF). Although this method isdependable, the detection rate is very low, and this method is invasive. So this testmethod is difficult to spread. The most popular detection method in lab is ELISA. Itcan only use crude antigens from worm, so the sensibility and specificity is low. Inconclusion, it’s very important to find biomarkers, which are very important forimproving the test method.Past research has shown that the cathepsin play an important role in Parasite-host interaction. Mice have a lot of cathepsin. We find18kinds of cathepsins, namedas cathepsin A,B,C,D,E,F,G,H,J,K,L,M,O,Q,R,S,W,Z. By analyzing literatures abouthost’s cathepsin, we guessed that mice’s cathepsins may take part in Meningoencep-halitis process.We got the sequence of Angiostrongylus cantonensis hemoglobinase-typecysteine proteinase from GenBank database. Past research told us that wormcathepsins associated with invasion of the host, self protection, and immune evasion.My research can be divided into two parts: First, Expression Profiles ofCathepsins from infected mice with Angiostrongylus cantonensis. Second, we cloneworm cathepsin. Part one: Expression Profiles of Cathepsins from infected mice withAngiostrongylus cantonensis.RESEARCH OBJECTIVEWe used BALB/c as infection model, and we observed expression change of thecathepsins of mice. By doing this, we could provide experimental basis for findingbiomarker to diagnose the disease.CONTENT AND METHOD1. We found mice cathepsins from GenBank database.2. We designed Real-time PCR primers on the website: https://www.genscript.com/ssl-bin/app/primer.3. We constructed Ac infection model by using BALB/c mice. And we got thebrain at different time points(7d、14d、21d). We tested the gene expression situation.RESULT1. We found18kinds of mice cathespins, named cathepsin A, B,C,D,E,F,G,H,J,K,L,M,O,Q,R,S,W,Z.2. We succeed in constructing BALB/c mice infection model. By testingexpression change during infection, we could classified them as three kinds: gradualincrease、sweep increase and irregular curve.CONCLUSIONThis work indicates that mice cathepsins changed while it was infected with Ac,and most kinds of cathepsins had expression increasing. Analysis results pointed outthat cathepsin C、S and M may be used as biomarkers for diagnosing Ac infection. Part two: Cloning of Angiostrongylus cantonensishemoglobinase-type cysteine proteinaseRESEARCH OBJECTIVECloning of Angiostrongylus cantonensis hemoglobinase-type cysteineproteinase, which is useful for studying the immune function.CONTENT AND METHOD1. We found the sequence in the GenBank database and did bioinformaticsanalysis.2. We constructed pET32a(+)/AcHTCP recombinant plasmid And didprokaryotic expression.CONCLUSIONWe construct pET32a(+)/AcHTCP prokaryotic expression system.