Effect Of Thyroid Hormone Deficiency In The Duration Of Pregnancy And Lactation On Offsprings Rats Auditory Function And Inner Ear Otoferlin Protein Expression

1. ObjectiveTo establish the animal models of TH defiency of rats in the duration of pregnancy and lactation with 0.02% MMI water, and investigate the influence of TH defiency on the off springs hearing ability such as ABR, DPOAE, etc. as well as the fine structure of inners and the expression of otoferlin protein in the inner ear.2. Methods2.1 24 rats of about 6～8 weeks old (12 male and 12 female,Ⅱdegree) were provided by the animal center of our university. Rats ABR were tested to make sure that there were with normal hearing. The animals were fed in quiet environment for one week before the experiment and then the male and female were 1:1 matched. Vagina discharge of females were detected in the next morning, and defined this day as the 0 day of embryo (E0) if any the in the charge were found. 2.2 Animals were divided into 4 experiment groups and a control one. The experiment groups were given 0.02% MMI water. For the group one, the MMI water was provided from the 0 day of embryo (E0) to the 15th day of embryo (E15), the group two was from the 0 day of embryo (E0) to the 0 day after birth (P0), the group three was from the 15th day of embryo (E15) to the 5th after birth (P5), and the group four was from the 15th day of embryo (E15) to the 12th after birth (P12).2.3 The blood Serum of both offspring rats and their mothers were collected 1, 3, 5, 10 days after adding drugs and 1, 3, 5 days after stopping drug water to examine the FT3,FT4 level. Frozen sections of adult offering rats thyroid gland were produced to observe the morphous with HE staining.2.4 ABR and DPOAE of offspring rats at p30, p60 and p90 were examined and the threshold of ABR, latency ofⅠ,Ⅲ,Ⅴwave in a SPL of 80 dB, DPOAE amplitude and SNR were recorded.2.5 Scanning electron microscope was used to detect the morphologic change of IHCs and OHCs in offsprings rats at 30,60,90days after birth of both control and experiment groups. Immunofluorescence staining was used to observe the hair cell's development and otoferlin protein expression in the inner ear.3. Results3.1 The free thyroxine level of both mother rats and offerings rats decreased after given the MMI water. but gradually came to normal 3 days after stopping drug. Frozen section of adult offering rats thyroid gland in the experiment groups shew no difference from that of the control group after HE staining. 3.2 ABR threshold of the experiment group one had no difference from that of the control group , while that of experiment grouptwo, three, and four were significantly higher than that of the control group. The rising degree of ABR threshold was related to the duration of the MMI water and was irreversible. DPOAE and ABRⅠ,Ⅲ,Ⅴwave latency had no statistically significant difference among all the groups (P>0.05).3.3 Scanning electron microscope and stretched preparation shew that the hairs cells in all the experiment groups lined up in order, without loss, fusion or lodging. Immunohistochemistry staining of brain stem sections were observed with a microscope and shew no difference in the neuron and gliocyte quantity.3.4 otoferlin protein expression in the inner ears shew no difference among the experiment group 1, 2, 3 and the control group, as well as in the p30,p60 group 4 animals. In p90 experiment group 4 offspring rats, otoferlin protein expression in the basal turn of the inner ears shew no difference from that of the control group at P30 and P60, while in the apical turn, otoferlin protein expression was obviously dimlier than that of the control group.4. Conclusions4.1 We produced a stable animal models of TH defiency in the duration of pregnancy and lactation with 0.02% MMI water. This animal models could provide a foundation for our further study.4.2 Thyroid hormone deficiency of mothers during the development period of offsprings hearing could lead to the lesion of hearing ability of the offsprings, and the lesion was related to the thyroid hormone dificiency period and was gradual and irreversibily. But the ABR latency, DPOAE and the conduction pathway shew no damage. 4.3 In p90 experiment group 4 offspring rats, otoferlin protein expression in the apical turn of the inner ears was obviously dimlier than that of the control group,which may cause the damage to the low frequency transduction. It suggests that thyroid hormone deficiency during the development period of hearing may cause the damage of the hearing ability, Which may be related to the otoferlin protein expression in the inner ear and AN development.