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Peripheral Nerve Injury of Hartley Guinea Pigs Induced by Gastrointestinal Infections of Campylobact

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Tutor: LiZhenZhong
School: Hebei Medical University
Course: Neurology
Keywords: Campylobacter jejuni,Pertussis toxin,Peripheral nerve injury,Guillain-Barre synd
CLC: R745
Type: Master's thesis
Year:  2014
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Abstract:
Objective: In order to provide more experimental supports for the buildingof animal models of peripheral neuropathy-Guillain-Barre syndrome, weinfected Hartley guinea pigs with Campylobacter jejuni and injected them withPertussis toxin, and try to investigate the influence of pertussis toxin togetherwith Campylobacter jejuni on the peripheral nerve injury compared withCampylobacter jejuni alone.Method:1Campylobacter jejuni culture, identification and bacterial suspensionpreparation: Campylobacter jejuni was isolated from the Guillain-Barresyndrome patients, identified by the China Center for Disease Control andPrevention. Campylobacter jejuni lulei was cultured on Colombia blood agarplate under microaerobic condition of42℃,5%O2,10%CO2,85%N2.Campylobacter jejuni lulei was identified by Gram staining, hippurate hydrolysisexperiments and catalase test. We purificated the bacteria and prepared bacterialsuspension with a concentration of3×108cfu/ml.2Experimental animal groups:30Hartley guinea pigs were randomly dividedinto experimental group and control group. Experimental group was furtherdivided into two subgroups: Group A: Campylobacter jejuni strains lulei pluspertussis toxin; Group B: Campylobacter jejuni strains lulei plus saline. Controlgroup was also further divided into three subgroups: Group C: Brucella brothplus pertussis toxin; Group D: Brucella broth plus saline; Group E: completelyblank control.3Experimental animal treatment: Hartley guinea pigs were orally fed with Campylobacter jejuni. Foods were deprivated for12hours and water wasdeprivated for4hours before Campylobacter jejuni feeding. Hartley guinea pigsin group A, group B were orally fed2ml bacterial suspension for4weeks with aconcentration of3×108cfu/ml per animal,3days a week. Hartley guinea pigs ingroup C, group D were given2ml Brucella broth for4weeks,3days a week.Hartley guinea pigs in group A and group C were injected with0.5ml of salinecontaining of1ug pertussis toxin during the first and the third time ofCampylobacter jejuni feeding of the first week and the second week. Hartleyguinea pigs in group B and group D were injected with0.5ml saline only.4Experimental animals’ clinical symptoms observation, pathological change andstatistics: Guinea pigs’ clinical symptoms were observed every day. Guinea pigs’body weight were weighed regularly. Sciatic nerves were extracted one weekafter the last time of Campylobacter jejuni feeding. Sciatic nerves were observedby transmission electron microscopy after osmium tetroxide staining. And χ2testwere conducted according to the lesions rate of osmium tetroxide staining.Results:1Experimental animal observation: According to the clinical symptom scorestandard, all the scores of group A, group B, group C, group D and group E were0. However, one guinea pig in group A had dirty fur on its abdomen from thethird week. The body weight of guinea pigs increased slowly in group A andgroup B and the hair of guinea pigs began to loss from the third week in group Aand group B.2Peripheral nerve pathology:2.1Osmium tetroxide staining: The thickness of partial sciatic nerve fibers ofguinea pigs in group A is inhomogeneous and the myelins are segmentallyatrophyedd and ruptured, just like the Great Wall. Some of the nerve fibers arealso fractured. The thickness of partial sciatic nerve fibers in group B is alsoinhomogeneous and part of the myelins are segmental collapsed, just like the sawtooth. Sciatic nerve fibers of guinea pigs in the control group show noabnormalities.2.2Transmission electron microscope observation: In group A, the thickness ofsciatic nerve myelin is inhomogeneous. The lamellar structure of myelin isdisappeared and part of the myelins are fractured. For the repression of myelin,some axons are disappeared. In group B, part of the lamellar structure of myelinis disappeared and anastomosised. Microtubules and filaments are packed tightly.Mitochondrial cristae are blurred. In the control group, the myelin thickness ishomogeneous and the lamellar structure is normal, microtubules and filamentsare also arranged properly.3Nerve fiber lesions rates and statistical analysis:3.1Nerve fiber lesions rates: All of the Hartley guinea pigs appeared peripheralnerve injury in experimental groups. The average nerve fiber lesions rates ofHartley guinea pigs in experimental groups and control groups are different.Nerve fiber lesions rate of group A is47.3%. Nerve fiber lesions rate of group Bis18.8%. Nerve fibers lesions rate of group C is3.8%. Nerve fiber lesions rate ofgroup D is2.5%. Nerve fiber lesions rate of group E is2.8%.3.2Statistical analysis: All the data were processed by SPSS13.0. Chi-square testwith bilateral significant testing were used to analysis the average rates of nervefiber lesions and α=0.05was set as the test level. We consider that the nervefiber lesions rates of Hartley guinea pigs in different groups are not all equall(P<0.05). Further comparison among different groups used the partitions of χ2method and α=0.05was set as the test level. We need to make the comparisonbetween group A and group E, group B and group E, group C and group E, groupD and group E, group A and group B. Test level was adjusted α’=0.01. Thelesion rates of Hartley guinea pigs nerve fiber between group A and group E arestatistical significantly different (P <0.01) and nerve fiber lesion rates in group Aare higher than group E. The lesion rates of Hartley guinea pigs nerve fibersbetween group B and group E are statistical significantly different (P <0.01) and the lesion rates in group B are higher than group E. The lesion rates of Hartleyguinea pigs nerve fibers between group C and group E are not statisticalsignificantly different (P>0.01). The lesion rates of Hartley guinea pigs nervefibers between group D and group E are not statistical significantly different (P>0.01). The lesion rates of Hartley guinea pigs nerve fibers between group A andgroup B are statistical significantly different (P <0.01) and the lesion rates ofHartley guinea pigs in group A were higher than group B.Conclusion:1Gastrointestinal infections with campylobacter jejuni can lead to peripheralnerve injury in the three-week-old Hartley guinea pigs. All of the guinea pigsappeared peripheral nerve injury, and the average rate of peripheral nerve lesionreached18.8%.2Gastrointestinal infections with campylobacter jejuni and combined withintraperitoneal injection of pertussis toxincan lead to peripheral nerve injury inthe three-week-old Hartley guinea pigs. All of the guinea pigs appearedperipheral nerve injury, and the average rate of peripheral nerve lesion increasesto47.3%.Corollary:Gastrointestinal infections of campylobacter jejuni can lead to peripheral nerveinjury on Hartley guinea pigs. Pertussis toxin can exacerbate the peripheral nerveinjury through changing the immune status or vascular permeability near theblood-nerve barrier of guinea pigs.
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