碘过量和多聚肌苷酸-聚胞苷酸及甲状腺球蛋白诱发小鼠甲状腺炎对Toll样受体3表达的影响

Objective To observe the effect of iodine excess(HI),polyinosinic-polycytidylic acid[Poly(I:C),Poly]and thyroglobulin(TG)on the thyroid of mice by the expression of Toll-like receptor 3(TLR3)to reveal the functional role of TLR3 in autoimmune thyroiditis.Methods Forty-two non-obese diabetic mice,body weight (20±3)g,were divided into six groups:control group,HI group,Poly group,TG group,HI+TG group,HI+Poly group. Fed with deionized water and injected intraperitoneally with physiological saline 0.1 ml each day for a week, the mice in control group were injected with physiological saline every other day at the same dose for 1 week before they were sacrificed; HI group drank 0.05% NaI water and were injected intraperitoneally with physiological saline same as control group; Poly group drank deionized water and were injected intraperitoneally with poly 0.1 ml (1 g/L)each day of the week, then the mice were injected with Poly every other day at the same dose for 1 week before they were sacrificed; TG group drank deionized water and were injected intraperitoneally with physiological saline same as control group, immunized with 0.1 mg TG by subcutaneously injecting and the immunization was enhanced after they were fed half dose for 4 and 8 weeks separately. In HI + Poly group, the treatment was the same as HI group and Poly group; HI + TG group: the treatment was the same as HI group and TG group. Eight weeks later, mice were sacrificed and thyroids were taken to make frozen sections, Hematoxylin-Eosin (HE) staining was employed to observe the morphological change of the thyroids. The expression of TLR3 of thyroids was observed under fluorescence microscope after Immumofluorescence using TLR3 antibody and TR3-positive cells were analyzed in the thyroid density. Results HE staining showed thyroids of Poly group had no inflammation under microscope.There were different degrees of inflammatory cell infiltration in HI group and TG group. The inflammatory cell infiltration and the damage of follicular thyroid of HI + TG group and HI + Poly group were serious, and the degrees of inflammation were higher over "++". Thyroid follicular epithelial cell with TLR3 expression could be seen in Poly group and HI group, meanwhile, there were TLR3 strong positive inflammatory cells in HI group under fluorescent microscope. Using stereological analysis of TLR3-positive cell density in the thyroid, the difference between groups was statistically significant(F=7.870, P<0.01 ). TLR3-positive cell density in the thyroid of HI + Poly group was higher[ (9.287 ± 0.522)mm2] than control group[ (0.062 ± 0.025)mm2, P < 0.01] significantly, meanwhile, the density in HI + Poly group was higher than HI group [ (2.574 ± 0.257 )mm2] and Poly group[ (1.361 ± 0.148 )mm2, all P < 0.01]. The density in HI + TG group[ (4.843±0.405)mm2] was higher than HI group and TG group[(1.601 ±0.268)mm2, all P < 0.01 )]. Conclusions Excessive iodine and thyroglobulin can induce thyroiditis, and stimulate the expression of TLR3 in the thyroid follicular epithelial, Poly aggravated thyroiditis induced by iodine excess in NOD mice; TLR3 positive inflammatory cells also appeared in inflammatory region, suggesting that TLR3 is involved in the pathogenesis of autoimmune thyroiditis     

碘过量和多聚肌苷酸-聚胞苷酸及甲状腺球蛋白诱发小鼠甲状腺炎对Toll样受体3表达的影响

Objective To observe the effect of iodine excess(HI),polyinosinic-polycytidylic acid[Poly(I:C),Poly]and thyroglobulin(TG)on the thyroid of mice by the expression of Toll-like receptor 3(TLR3)to reveal the functional role of TLR3 in autoimmune thyroiditis.Methods Forty-two non-obese diabetic mice,body weight (20±3)g,were divided into six groups:control group,HI group,Poly group,TG group,HI+TG group,HI+Poly group. Fed with deionized water and injected intraperitoneally with physiological saline 0.1 ml each day for a week, the mice in control group were injected with physiological saline every other day at the same dose for 1 week before they were sacrificed; HI group drank 0.05% NaI water and were injected intraperitoneally with physiological saline same as control group; Poly group drank deionized water and were injected intraperitoneally with poly 0.1 ml (1 g/L)each day of the week, then the mice were injected with Poly every other day at the same dose for 1 week before they were sacrificed; TG group drank deionized water and were injected intraperitoneally with physiological saline same as control group, immunized with 0.1 mg TG by subcutaneously injecting and the immunization was enhanced after they were fed half dose for 4 and 8 weeks separately. In HI + Poly group, the treatment was the same as HI group and Poly group; HI + TG group: the treatment was the same as HI group and TG group. Eight weeks later, mice were sacrificed and thyroids were taken to make frozen sections, Hematoxylin-Eosin (HE) staining was employed to observe the morphological change of the thyroids. The expression of TLR3 of thyroids was observed under fluorescence microscope after Immumofluorescence using TLR3 antibody and TR3-positive cells were analyzed in the thyroid density. Results HE staining showed thyroids of Poly group had no inflammation under microscope.There were different degrees of inflammatory cell infiltration in HI group and TG group. The inflammatory cell infiltration and the damage of follicular thyroid of HI + TG group and HI + Poly group were serious, and the degrees of inflammation were higher over "++". Thyroid follicular epithelial cell with TLR3 expression could be seen in Poly group and HI group, meanwhile, there were TLR3 strong positive inflammatory cells in HI group under fluorescent microscope. Using stereological analysis of TLR3-positive cell density in the thyroid, the difference between groups was statistically significant(F=7.870, P<0.01 ). TLR3-positive cell density in the thyroid of HI + Poly group was higher[ (9.287 ± 0.522)mm2] than control group[ (0.062 ± 0.025)mm2, P < 0.01] significantly, meanwhile, the density in HI + Poly group was higher than HI group [ (2.574 ± 0.257 )mm2] and Poly group[ (1.361 ± 0.148 )mm2, all P < 0.01]. The density in HI + TG group[ (4.843±0.405)mm2] was higher than HI group and TG group[(1.601 ±0.268)mm2, all P < 0.01 )]. Conclusions Excessive iodine and thyroglobulin can induce thyroiditis, and stimulate the expression of TLR3 in the thyroid follicular epithelial, Poly aggravated thyroiditis induced by iodine excess in NOD mice; TLR3 positive inflammatory cells also appeared in inflammatory region, suggesting that TLR3 is involved in the pathogenesis of autoimmune thyroiditis     

碘过量和多聚肌苷酸-聚胞苷酸及甲状腺球蛋白诱发小鼠甲状腺炎对Toll样受体3表达的影响

目的 观察碘过量(high iodine,HI)和多聚肌苷酸-聚胞苷酸[Polyinosinic-Polycytidylic acid,Poly (I:C),Poly]及甲状腺球蛋白(Thyroglobulin,TG)诱发小鼠甲状腺炎对Toll样受体3(Toll-like receptor 3,TLR3)表达的影响,探讨TLR3在自身免疫性甲状腺炎发病中的作用.方法 NOD(Non-obese diabetic)小鼠42只,体质量(20±3)g.按体质量将小鼠随机分为6组:对照组、HI组、Poly组、TG组、HI+TG组、HI+Poly组,每组7只.对照组:饮用去离子水,腹腔注射生理盐水0.1 ml,每天1次,连续1周,在处死小鼠前1周隔日1次,同样剂量生理盐水再注射3次;HI组:饮用0.05%的碘化钠去离子水,腹腔注射生理盐水(同对照组);Poly 组:饮用去离子水,腹腔注射0.1 ml Poly(1 g/L,按5 mg/kg体质量),每天1次,连续1周,处死前1周隔日1次,同剂量Poly再注射3次;TG组:饮去离子水,腹腔注射生理盐水(同对照组),皮下免疫猪TG 0.1 mg,在喂养第4、8周时分别再加强免疫1次,剂量减半;HI+Poly组:给药方法同HI组和Poly组;HI+TG组:给药方法同HI组和TG组.喂养8周后处死小鼠,取出甲状腺组织,冰冻切片、常规HE染色,光镜下观察小鼠甲状腺组织形态学变化:根据甲状腺组织炎细胞浸润数量及浸润范围、滤泡破坏范围等进行炎症程度分级;应用TLR3抗体对甲状腺切片进行免疫荧光染色,荧光显微镜下观察TLR3的表达,体视学分析甲状腺TLR3阳性细胞数密度变化.结果光镜下,Poly组甲状腺未见炎细胞浸润,HI组和TG组小鼠甲状腺都有不同程度的炎细胞浸润,HI+TG组和HI+Poly组甲状腺炎症细胞浸润和甲状腺滤泡破坏严重,炎症分级均在"++"以上.免疫荧光显示.HI组和Poly组的甲状腺滤泡上皮细胞可见到TLR3表达,在HI组和HI+Poly组炎症区域出现TLR3表达强阳性的炎症细胞.体视学分析甲状腺TLR3阳性细胞数密度,对照组、HI组、Poly组、TG组、HI+TG组、HI+Poly组组间比较差异有统计学意义(F=7.870,P<0.01);与对照组[(0.062±0.025)mm2]比较,HI+Poly组[(9.287±0.522)mm2]增加最为显著(P<0.01),而且HI+Poly组高于HI组[(2.570±0.257)mm2]和Poly组[(1.361±0.148)mm2,P均<0.01],HI+TG组[(4.843±0.405)mm2]高于HI组和TG组[(1.601±0.268)mm2,P均<0.01].结论 HI和TG免疫可诱发NOD鼠发生甲状腺炎,并刺激甲状腺滤泡上皮表达TLR3,Poly加重了HI诱发的NOD鼠甲状腺炎的病理变化过程;浸润的炎症细胞中亦有TLR3强阳性的细胞,提示TLR3途径参与了自身免疫性甲状腺炎的发病过程.     

Antibody response of mice to chemically induced tumors.

BALB/c mice immunized with syngeneic methylcholanthrene-induced fibrosarcomas did not have antibodies against the unique tumor-specific transplantation antigens, even though they were capable of rejecting a lethal dose of tumor cells. Endogenous murine leukemia virus antigens expressed by certain of the tumors did, however, elicit high titers of antibodies, accounting for serological crossreactions that occurred between those tumors.     

Effect of cholecystokinin-B gastrin receptor blockade on chemically induced colon carcinogenesis in mice: follow-up at 52 weeks

BACKGROUND/AIMS: The potential role of gastrin and the cholecystokinin-B (CCK-B)/gastrin receptor in the genesis of colon cancer is debated. Aberrant crypt foci (ACF) are considered to be preneoplastic lesions of colon cancer. We aimed to assess whether the CCK-B/gastrin receptor antagonist, CR2945, may prevent the development of ACF and adenocarcinoma in the experimental model of dimethylhydrazine (DMH)-induced colorectal cancer. MATERIALS AND METHODS: 226 CD1 mice were randomized into 3 groups (sham, control and treated) and received intraperitoneal injections of NaCl 0.9%, DMH, and DMH + CR2945, respectively, for 5 weeks. 168 mice were sacrificed at 15, 38, 45 and 52 weeks after the first injection day. The colon and rectum were investigated for frequency, multiplicity and distribution of ACF as well as for adenocarcinoma at histology. The expression of gastrin was assessed in tumor samples at histology by immunohistochemistry. RESULTS: ACF frequency and multiplicity significantly increased with time in both controls and treated mice with no difference between groups except that at week 45. 38.8% of controls and 14.3% of treated mice developed cancer (p = 0.004). No cancer was positive for gastrin at immunohistochemistry. The mean number of cancers per mouse and the proportion of mice with cancer increased with time with statistically significant difference between controls and treated mice at week 38 only but not afterwards. A significant correlation between cancer and ACF frequency (r = 0.35) and multiplicity (r = 0.25) was observed. CONCLUSIONS: Our findings support the preneoplastic significance of ACF and indicate that CR2945 treatment does not interfere with the DMH-induced carcinogenic process.     

Lung tumorigenesis by 1,2-diformylhydrazine in mice

Summary 1,2-Diformylhydrazine was administered in drinking water as a 2% solution to randomly bred Swiss albino mice for life from 6 weeks of age. As a result of treatment, the lung tumor incidence rose from 15 to 96% in females and from 22 to 82% in males. The treatment had no statistically significant effect on the development of other types of tumors. Histopathologically, the neoplasms exhibited the characteristic appearance of adenomas and adenocarcinomas of the lungs.The work is part of a series of studies aimed at revealing the relative carcinogenic potency of mono and dialkyl-hydrazines and also their possible environmental significance as cancer causative agents.Supported by USPHS Contract N01 CP33278 from the National Cancer Institute, NIH, USA     

Hyaluronic acid produced by human synovial fibroblasts. Effect of polyinosinic-polycytidylic acid (POLY I:C) and interferon

Poly I:C (polyinosinic-polycytidylic acid) stimulated hyaluronic acid production by rheumatoid and non-rheumatoid human synovial fibroblasts. Stimulation was dose dependent and was inhibited by acetylsalicylic acid and indomethacin. Poly I and Poly C, when separately added, had no stimulatory effect on hyaluronic acid production, and Poly A:U had only a slight effect on this parameter. Cells grown with Poly I:C were virus resistant and interferon was detected in their medium. Human interferon had also a dose-dependent stimulatroy effect on hyaluronic acid production by synovial cells. A possible interferon-mediated relationship between virus infection and pathologic accumulation of joint fluid is suggested.     

Lung tumorigenesis by 1,2-diformylhydrazine in mice.

1,2-Diformylhydrazine was administered in drinking water as a 2% solution to randomly bred Swiss albino mice for life from 6 weeks of age. As a result of treatment, the lung tumor incidence rose from 15 to 96% in females and from 22 to 82% in males. The treatment had no statistically significant effect on the development of other types of tumors. Histopathologically, the neoplasms exhibited the characteristic appearance of adenomas and adenocarcinomas of the lungs. The work is part of a series of studies aimed at revealing the relative carcinogenic potency of mono- and dialkyl-hydrazines and also their possible environmental significance as cancer causative agents.     

Effect of chemically induced diabetes mellitus on glutamine transport in rat intestine

The effect of chemically induced diabetes mellitus on the intestinal transport of glutamine was examined using a brush-border membrane vesicle technique. Diabetes was induced by a single intraperitoneal injection of streptozotocin (100 mg/kg body weight). Control and diabetic rats were studied 5 days following the induction of diabetes. Na(+)-dependent and Na(+)-independent glutamine (0.5 mM) transport was found to be significantly higher in the diabetic rats than in the control rats. This increase was found to be caused by a significant increase in the Vmax of the Na(+)-dependent and the Na(+)-independent glutamine transport processes in the diabetic rats (Vmax of 3742 +/- 487 and 2055 +/- 279 pmol/mg protein per 7 s, respectively) compared with that of the control rats (2183 +/- 75 and 1271 +/- 83 pmol/mg protein per 7 s, respectively). The apparent Km values of glutamine transport systems, on the other hand, were similar in the two rat groups. Insulin treatment of the diabetic rats significantly reduced the Vmax of glutamine transport by both the Na(+)-dependent and the Na(+)-independent processes to a level similar to that of the control rats (Vmax in the insulin-treated diabetic rats of 2036 +/- 123 and 1247 +/- 105 pmol/mg protein per 7 s, respectively). This study demonstrates that chemically induced diabetes mellitus is associated with an increase in intestinal glutamine transport. This increase is the result of the diabetic condition itself and appears to be mediated through an increase in the number of the transport carriers of glutamine.     

Host genes affecting survival period of chemically induced bladder cancer in mice

Treatment of C57BL/6?J (B6) and NON male mice with N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) resulted in a high incidence of bladder cancer. The mean survival period, however, differed significantly by strain: 481?±?219 days in B6 (n?=?31) and 203?±?119 days in NON (n?=?30) (P?<?0.0001). Major causes of death were renal failure due to obstruction of the urinary tract, or local invasion of tumors. The fact that the BBN-treated NON?×?B6 reciprocal F1 mice had survival periods as short as those of the parental NON mice suggests a genetically dominant susceptibility in NON or recessive resistance in B6. A linkage analysis of 248 back-cross mice to B6 suggested at least two quantitative trait loci determining the length of the survival period: one was mapped close to D2Mit260 (logarithm of odds, LOD, score 2.21), a microsatellite marker locus 83?cM from the centromere on chromosome 2, and another was close to D6Mit159, 7?cM from the centromere on chromosome 6 (LOD score 2.51).     

二甲双胍、塞来昔布对化学诱导小鼠结直肠异常隐窝病灶预防作用的比较

[目的]探讨二甲双胍对氧化偶氮甲烷(AOM)诱导的小鼠结直肠异常隐窝病灶(ACF)的预防作用,并与塞来昔布进行比较。[方法]45只6周龄BALB/c雌性小鼠随机分为对照组、二甲双胍组、塞来昔布组,每组15只;均腹腔注射AOM(10mg/kg),每周1次,连续2周以建立ACF模型;同时对照组、二甲双胍组、塞来昔布组小鼠分别予0.9%氯化钠溶液0.5ml·只~(-1)·d~(-1)、二甲双胍250mg·kg~(-1)·d~(-1)、塞来昔布20mg·kg~(-1)·d~(-1)灌胃处理,每周监测小鼠体质量变化。6周后处死小鼠并分离结直肠组织,美蓝染色后计数每只小鼠结直肠ACF个数和每个ACF中畸变隐窝(AC)个数。[结果]各组小鼠体质量随周龄增加均呈现增长趋势,干预后体质量比较均差异无统计学意义(P0.05)。对照组小鼠结直肠ACF及AC数量分别为(8.80±0.84)个、(26.75±2.63)个,二甲双胍组分别为(3.67±0.58)个、(8.33±0.58)个,塞来昔布组分别为(5.60±0.89)个、(13.40±1.67)个;与对照组相比,二甲双胍组、塞来昔布组小鼠ACF及AC均显著减少(P0.05);二甲双胍组小鼠ACF及AC均比塞来昔布组显著减少(P0.05)。[结论]二甲双胍、塞来昔布均能显著抑制ACF形成,对结直肠癌起到预防的作用,且二甲双胍的作用明显优于塞来昔布。     

Selection of liver-colonizing tumor cells from a murine fibrosarcoma induced by methylcholanthrene

Summary An experimental tumor model was developed to study the organ preference of malignant tumors. The primary tumor ER 15-P was induced by in-oculation of 1 mg methylcholanthrene in 0.1 ml sesame oil into the left femoral muscle of a female C57/B1₆J mouse. The tumor was palpable 100 days after induction. Spontaneous lung metastases were found at autopsy on day 128. Serial IM transplantation of tumor cells from the primary ER 15-P resulted in pulmonary metastases in all male and female mice. After IV injection of tumor cells from ER 15-P to male mice, colonies were found in lungs, thoracic cavity, liver, kidneys and occasionally also adrenals; female mice sometimes had ovarian metastases in addition, but no hepatic metastases. Liver-colonizing tumor cells were selected in male mice as follows: (a) IV injection of tumor cells from primary ER 15-P; (b) removal of tumor cells from liver tumor nodules, reinjection into mesenteric vein; (c) preparation of resulting tumors in the liver, reinjection of these cells through the portal system in one group of mice, and IV administration into tail vein in another group: (d) IM inoculation of tumor cells of the mesenteric passage in the left gastrocnemius muscle of mice prior to IV injection via tail vein in another group. Steps c and d were repeated three times. The procedure resulted in a highly significant decrease of tumor cell colonization to lungs and other organs, and a preferential increase of liver colonization. The liver preference of cell lines thus selected was obvious. Possible mechanisms for the organ preference of malignant tumors are discussed.     

硫酸软骨素对大鼠酒精性肝纤维化模型的影响

目的探讨硫酸软骨素对酒精性肝纤维化的治疗作用。方法用连续灌服酒精的方法建立大鼠酒精性肝纤维化模型,在造模成功后给予硫酸软骨素治疗6周(25mg/Kg,腹腔注射,每日一次)。测定血清和肝脏匀浆丙二醛(MDA)水平,采用VanGieson染色法显示肝脏组织中胶原的表达,免疫组化检测肝组织转化生长因子β1(TGFβ1)、金属蛋白酶组织抑制因子1(TIMP1)的表达。结果硫酸软骨素能减轻酒精性肝纤维化的组织病理改变,降低酒精性肝纤维化大鼠肝组织胶原和TIMP1的表达。酒精性肝纤维化中,肝组织MDA和TGFβ1的表达增加,使用硫酸软骨素可显著抑制MDA和TGFβ1的表达(P<0.01)。结论硫酸软骨素对酒精性肝纤维化有治疗作用,其作用机制可能与影响MDA和TGFβ1的表达水平相关。     

聚肌胞苷酸诱导的早期原发性胆汁性肝硬化小鼠模型的建立

目的：探讨应用聚肌胞苷酸建立早期原发性胆汁性肝硬化动物模型。方法30只雌性C57BL/6小鼠被随机分为模型组和对照组。模型组小鼠腹腔注射聚肌胞苷酸（5 mg.kg-1）,对照组小鼠注射等体积生理盐水,均2次/w,于8 w、16 w和24 w分别检测血清抗线粒体抗体M2亚型、外周血CD4＋CD25＋Foxp3＋计数,以及肝组织CK19表达情况。结果在24 w末,模型组AMA-M2阳性率为100%（5/5）,显著高于对照组（0/5）;在8 w、16 w和24 w,模型组小鼠外周血CD4＋CD25＋Foxp3＋细胞计数分别为（3.48±0.95）%、（3.30±1.55）%和（2.67±0.97）%,均显著低于相应时间点对照组水平[（7.25±1.63）%、（6.33±1.06）%和（5.58±1.52）%,P〈0.05];两组肝组织CK19表达无显著性差异;病理学检查显示模型组小鼠肝组织汇管区和胆管周围炎性细胞浸润呈进行性加重,细小胆管增生,出现胆汁性碎屑样坏死和桥接纤维化,但未出现肝硬化。结论聚肌胞苷酸注射可成功建立早期原发性胆汁性肝硬化动物模型,CD4＋CD25＋Foxp3＋T细胞可能在发病中发挥重要作用。     

3H-arginine-rich cationic proteins in cytosol of fibrosarcoma, induced by methylcholanthrene in rats

With the use of 3H-arginine it is shown that among soluble cytosol of fibrosarcoma, induced by 20-methylcholanthrene in rats there appear cationic proteins rich in arginine with content of amino acid of about 10%. The amount of these proteins in cytosol fraction is higher than in cytosol of another types of experimental tumors. The role of arginine-rich cationic proteins in cytosol fraction is briefly discussed.