Interleukin-10 Gene Therapy Of Experimental Autoimmune Thyroiditis And Its Mechanism

PART â…  The pathologic effect of iodine on experimental autoimmuneanimal modelObjective: To make experimental autoimmune thyroiditis (EAT) animal model and investigate the pathologic effect of iodide on this model. Methods: Female Wistar rats were divided into four groups randomly: group A- normal control, group B- rats immunized with porcine thyroglobulin, group C- rats raised with 0.05 % iodine water, group D -rats treated with combination of 0.05 % iodine taking and immunization. Blood samples were taken at 0, 2, 4, 6, 8^th wk. TgAb and TmAb were tested by RIA. FT3, FT4 and TSH were measured by chemiluminescence method. The morphological features of thyroid tissues were observed through microscope. Results: The level of TgAb and TmAb of group B rose at 2^th wk and reached to the peak value at 4^th wk, then descended, though still higher than those of group A (P<0.001). The autoantibodies in group C increased gradually. The titer of TgAb and TmAb of group D rose most quickly and lasted for longest time in all groups. Absolute and relative weights of thyroid tissue of the rats from group D were heavier than those of group A and B (P<0.01). Lymphocyte infiltration in thyroid tissue presented in group B, anisometric folliculi were found in group C, severe inflammation and proliferation of fibrous tissue showed in group D. Sera FT₃ and FT₄ level from group B were higher than those from goup A and group C at 8th wk (P<0.001), whereas lower than those of group D(P<0.05).Conclusions: EAT model was established successfully. Excessive iodine intake deteriorates experimental autoimmune thyroiditis, impacts the thyroidfounction of EAT rats and induces goiter and local follicular atrophy .Key words: Iodine, autoimmune thyroiditis, thyroid function, morphologyPART â…¡The mechanism of the therapeutic effect of IL-10 gene transmissionon experimental autoimmune thyroiditis ratsObjectives:â‘ To investigate the impact of liposome in combination with poly-L-lysine on the efficiency of gene transmission.â‘¡To study the effect of L-10 gene therapy in thyroid on the course of ongoing EAT and the immune disorders. â‘¢ To detect expression of intercellular adhesion molecule-1 (ICAM-1) on thyroid follicular cells.â‘£To observe the effect of IL-10 gene transmission on thyroid apopotosis.Methods: â‘ SD rats were divided into four groups: group A(normal control), group B(pORFmIL10+lipofectamine),group C(PBS+lipofectamine), group D (pORF+lipofectamine). mIL-10 mRNA and protein were detected by RT-PCR and Western blot. â‘¡According as the different substance injected into thyroid glands, EAT rats were divided into four groups: group A(PBS+PLL), group B(pORF+PLL), group C(pORFmIL10 +PLL), groupD (pORFmIL10+MEM). The rats sacrificed at 8^th wk, thyroids were fixed in 10% neutral formalin solution and processed according to standard procedures.The specimens were stained with hematoxylin-eosin -safran. Histological thyroiditis was scored from one to four with reference to Charreive criteria. The level of serum TgAb,TmAb of rats were detected at 0, 2, 4, 6, 8^th wk. In vitro proliferative responses to ConA and different concentration PTg were measured by culturing 4×10⁵ splenocytes pulsed with 18.5KBq of [³H]thymidine for the final 12h and then harvested for liquid scintillation counting. Splenocytes werecultured with PTg 25mg/L, Thl cytokines IFN- y ,TNF- a and IL- 2 were detected by ELISA. (3)ICAM-1 protein expression on thyrocytes from the rats were examined by immunohistochemistry method. (4)RT-PCR and TUNEL were used to evaluated Fas/FasL mRNA expression and the in situ apoptotic ratio.Results: (1)mIL-10 mRNA and protein were expressed in throid tissue of rats, mIL-10 were detected on 3th day in liposome group. Poly-L-lysine plus lipofectamine enhanced the efficiency of transmission and prolong the expresssion of target gene till 14th day. (2)A dramatic disappearance of the lymphocytic infiltration was observed in thyroids from group C, infiltration index is l.l±0.18, lower than any other gr...