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Study on in Vitro Infection of Tachyzoites of Toxoplasma Gondii RH Strain in Rat Brain Astrocytes

Author: LiDongNa
Tutor: ZhuGeHongXiang
School: Suzhou University
Course: Pathogen Biology
Keywords: Astrocytes Toxoplasma Tachyzoites Living cells workstation Autophagy
CLC: R382.5
Type: Master's thesis
Year: 2010
Downloads: 86
Quote: 0
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Abstract


Purpose of Toxoplasma infection can cause a variety of tissues and organs lesions, one of the most serious harm to Toxoplasma human brain across the blood-brain barrier invasion of Toxoplasma encephalitis. The established Toxoplasma gondii RH strain tachyzoites SD rats infected brain astrocytes in vitro model to study autophagy in astrocytes process of Toxoplasma gondii invasion explore arched mechanism The worm Tachyzoites possible non-specific immune cells infected brain astrocytes provide the foundation. Method 1. Primary cultured mixed glial cells of the cerebral cortex of newborn SD rats oscillation method to obtain purified astrocytes, immunohistochemical identification of cell purity. Recovery cryopreservation of Toxoplasma gondii abdominal infection Kunming mice, until the time of the mice showed symptoms of infection, to extract the peritoneal fluid, observation of Toxoplasma gondii virulence for three generations until inoculation collection Toxoplasma gondii tachyzoites . 3. Pending astrocytes reached the third generation, were seeded in petri dishes placed in advance slides, wait until the cells covered 80% of the petri dish, Toxoplasma gondii RH strain tachyzoites astrocytes interstitial cells co-cultured 0.5 to 72h, Giemsa staining, observed tachyzoite proliferation. 4. Toxoplasma gondii tachyzoites co-cultured with astrocytes on living cells workstation, observe the the RH tachyzoites role of astrocytes way. 5.MDC staining observed by fluorescence microscopy 0 ~ 48h, co-cultured astrocytes autophagic vesicles changes. Results (1) astrocytes shaken by thermostat separated, spread to the third generation, the cells mature, stable state. And GFAP antibody on astrocytes identification, purity 95% or more. (2) tachyzoites in Kunming mice passaged preserving, every 2 to 3 days passaged once. Transmitted to the third generation of Toxoplasma gondii virulence intact. (3) of Toxoplasma gondii cultured astrocytes 2h, the worm began to enter the cell; 8h enter the cell parasites increased significantly; 24h vast majority of the parasite has entered the cell; 48h, most of the star can see the parasite cysts false, glial cells were daisy-like arrangement cysts originally formed parasites, and then to binary fission proliferation gradually formed mature cysts, waiting for a breakthrough in the cell membrane when; 72h large some cells have been parasites breaking up a large number of parasites are released from the cells and start a new infection - fake cysts - proliferation - the process of rupture. After this cycle, the proliferation of parasites can be 10 to 20 times. (4) the living cells workstation, observed first Toxoplasma tachyzoites contact its side and astrocyte cell membrane, and then drill into the top of Toxoplasma gondii tachyzoites active invasion mouse astrocytes. (5) MDC staining of Toxoplasma gondii astrocytes co-cultured 1h, the intracellular punctate fluorescent particles; 2h, 4h, 8h, intracellular fluorescent particles gradually increased most significantly to 8h; at 12h , intracellular fluorescent particles decreased significantly, accompanied by cell injury, but still intact morphology; cells when the parasites infected cells 24h and 48h, fluorescent particles almost disappeared, while the cells are destroyed. Conclusion Toxoplasma gondii RH tachyzoites in vitro infection model of rat astrocytes; cycle of Toxoplasma gondii in vitro infection of mouse astrocytes 72h, its invasion of astrocytes in vitro active offensive; initial recognition autophagy process of Toxoplasma gondii in vitro infection of astrocytes was inhibited.

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CLC: > Medicine, health > Basic Medical > Medical Parasitology > Medical protozoa school > Toxoplasma gondii
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