IGF-1 mRNA expression of adult rat thyroid cell cultured in vitro

Objective: To investigate the law of age-related changes of insulin-like growth factor-1 (IGF-1) expression of rat thyroid cells cultured in vitro. Methods: Rat thyroid of different age (10, 45, 65, 100, 150 weeks) was isolated and thyrocytes cultured. Total RNA was extracted in different rat age group when thyroid cells had been cultured for two weeks. mRNA IGF-1 expression was measured with reverse-transcription polymerase chain reaction( RT-PCR) in each group and compared. Results: Quantity of total RNA in thyroid cells decreased with ageing when the rat thyroid cells had been cultured for 2 weeks. There is significant difference among groups (P<0.05). Expression of IGF-1 mRNA could be detected in thyroid cells of different age cultured in vitro. Quantity of IGF-1 mRNA expression by RT-PCR analysis increased from 10 to 45 weeks old, and then decreased with ageing. Conclusion: Rat thyroid cells from different age cultured in vitro can express IGF-1 mRNA. Quantity of total RNA in thyroid cells cultured in vitro decreased with aging. IGF-1 mRNA expression was correlated to age (r=0.401, P<0.05).

In the Basis of Hashimoto’s Thyroiditis, to Form Papillary Thyroid Carcinoma, Metastasized and Then to De-Differentiate into Poorly Differentiated Squamous Cell Carcinoma

Thyroid squamous cell carcinoma is very rare. At present, it is limited to case reports. Since the thyroid follicular epithelium is the non-squamous epithelium, how primary squamous cell carcinoma (SCC) of the thyroid occurs is still a controversial issue. Hashimoto’s thyroiditis (HT) is considered to be an independent risk factor for thyroid cancer, under the basis of HT, how tumor cells evolve and develop to papillary thyroid carcinoma (PTC), and particularly to de-differentiate into SCC is elusive. We report a 72-year-old female patient who developed multiple subtypes of PTC on a basis of HT, and finally to de-differentiate into SCC within the local foci of lymph node metastasis. We found that there was a variety of sub-types of PTC in this patient in the background of HT. SCC was found within local lymph node metastasis. Pathomorphology, immunohistochemistry, and molecular pathology have confirmed that the SCC was derived from PTC, and then developed into poorly differentiated SCC and/or anaplastic carcinoma. We also conducted a comprehensive literature review.

Didactic surgical experience of thyroid metastasis from renal cell carcinoma: A case report

BACKGROUND The optimal therapeutic strategy in treating thyroid metastasis from renal cell carcinoma(RCC) has not been clearly established. Here we describe a case of didactic surgical experience of the disease which caused massive intraoperative bleeding.CASE SUMMARY A 59-year-old male patient presented with a thyroid left lobe soft mass detected by chest computed tomography scans prior to the surgical treatment of RCC of the left kidney. The thyroid mass was initially considered to be benign, then he underwent left radical nephrectomy. One year after the nephrectomy, stereotactic radiosurgery was performed for brain metastasis. During follow-up, the thyroid nodule gradually grew, and the patient manifested swallowing discomfort. Under a clinical diagnosis of thyroid follicular neoplasm, left hemithyroidectomy was performed. Although hemithyroidectomy is usually a safe and straightforward procedure, massive bleeding from markedly developed tumor vessels made the operation very difficult. The thyroid tumor was finally diagnosed as metastasis from clear cell RCC. CONCLUSION For proper timing of the surgery, a clinician should take into consideration the possibility of thyroid metastasis of RCC when a thyroid lesion is found in patients with RCC or in patients with a previous history of RCC. We recommend that thyroid metastasis of RCC should be resected as early as possible even if a patient has other metastatic sites.

Effect of parathyroid hormone on apoptosis of human medullary thyroid carcinoma cells

Objective The aim of the study was to investigate the effect of parathyroid hormone(PTH) on the apoptosis of human medullary thyroid carcinoma(MTC) cells.Methods In vitro cultured medullary thyroid carcinoma cell lines were treated with parathyroid hormone and parathyroid hormone receptor-monoclonal antibody,and the apoptosis of cells was detected by flow cytometry.Results The cell morphology changed significantly after treatment based on the observation using the inverted phase-contrast microscope.Various concentrations of parathyroid hormone and parathyroid hormone receptor-monoclonal antibody effectively induced apoptosis in a time-and concentrationdependent manner.When the concentration of parathyroid hormone was 2.0 μmol/L and that of parathyroid hormone receptor-monoclonal antibody was 1.0 μmol/L,the apoptotic rate was 13.24% and 20.78%,respectively,representing a statistically significant difference from that of the control cells(P < 0.05).Conclusion PTH plays a role in inducing apoptosis of human MTC cells.

Inhibition effects of parathyroid hormone on human medullary thyroid carcinoma cells

Objective： The purpose of the study was to investigate the effects of parathyroid hormone and parathyroid hormone receptor monoclonal antibody on in vitro growth and proliferation of human medullary thyroid carcinoma cell lines. Methods： The medullary thyroid carcinoma cell line was cultured in vitro, with parathyroid hormone and parathyroid hormone receptor monoclonal antibody treatment intervention, the growth of the cells was observed under an inverted contrast micro scope, the MTT assay was used to detect the cell growth inhibition rate. Results： Under the inverted contrast microscope, the cells changed significantly, the parathyroid hormone and parathyroid hormone receptor monoclonal antibodies can effectively inhibit the proliferation of medullary thyroid cancer cells in a time and dose dependent. When parathyroid hormone concentra tion reached a concentration of 2.0 IJmol/L, the parathyroid hormone receptor monoclonal antibody reached a concentration of 1.0 μmol/L, the cell growth was most significantly inhibited （P 〈 0.05）. Conclusion： Parathyroid hormone and parathyroid hormone receptor monoclonal antibody were able to inhibit the proliferation of medullary thyroid carcinoma cells and signifi cantly reduce the proliferation index.

Characteristics of clear cell renal cell carcinoma metastases to the thyroid gland: A systematic review

BACKGROUND Thyroid gland is an uncommon site for metastases from clear cell renal cell carcinoma(CCRCC)and literature is scarce.Due to the variable and often long lag time before development of metastases in patients with CCRCC,thyroid nodules may be misdiagnosed initially as benign.This systematic review aims at a better understanding of the nature of these metastases.METHODS A bibliographic search was performed using PubMed(1990-2019),key words being“renal cell carcinoma,thyroid,kidney cancer,clear cell.”147 cases were analyzed.The patient’s characteristics assessed were:age,sex,stage,size of metastases,lag time,diagnostic modality,initial symptoms,treatment and outcome in last documented follow up.Binary logistic regression,Spearman’s rho and ANOVA were used to identify differences between the existing variables.RESULTS The mean age(±SD)was 64±(10)years in males and 64(±11)in females.The mean lag time to diagnosis of thyroid metastases was 8.7(±6.3)years.Gender distribution of the patients was 46.3%male,52.4%female.There was a weak correlation between lag time and size of metastases,not statistically significant.Size of metastases was significantly higher in symptomatic patients(6.06±3.51 cm)compared to those with painless mass(4.6±0.29 cm)and asymptomatic ones(3.93±1.99 cm)(P=0.03).Fine Needle Aspiration was diagnostic in 29.4%of cases,47.1%were non diagnostic.Most patients(80.3%)underwent thyroid surgery.At 1 year follow up,55.6%of patients operated were alive versus 35.3%who did not have surgery,though this was not statistically significant(P=0.1).CONCLUSION A larger size of thyroid metastasis was more likely to present with symptomatology.A high index of suspicion is warranted when evaluating thyroid nodules in CCRCC patients.There was no significant difference in outcome between patients who underwent surgery and those who did not.With the wider use of immune check-point inhibitors and tyrosine kinase inhibitors in metastatic CCRCC,surgery may eventually be reserved only for palliative purposes.

Thyroid follicular renal cell carcinoma excluding thyroid metastases:A case report

BACKGROUND Thyroid follicular renal cell carcinoma is a special type of renal cell carcinoma newly recognized in recent years.It has attracted attention because of its unique histology,immunophenotype,and clinical characteristics.It has a very low incidence,and the number of case reports available for review is limited.Moreover,a thyroid mass with type of tumour is rare.CASE SUMMARY We report a case of a renal mass with a bilateral thyroid mass that was accidentally discovered in a 60-year-old man during physical examination.B-mode ultrasound showed a hypoechoic mass in the middle and lower parenchyma of the right kidney,and computed tomography showed an iso-density shadow tumour in the right kidney.Contrast agents had a significant continuous enhancement effect on the tumour,and the enhancement was not uniform.After partial nephrectomy,pathological analysis was performed to rule out the possibility that the renal tumour was caused by thyroid tumour metastasis.Needle biopsy of the thyroid tumour confirmed that the renal cell carcinoma was not related to the thyroid tumour.The patient was alive at the last postoperative follow-up.CONCLUSION This is the third published case in which thyroid tumour biopsy was performed to confirm that thyroid follicular renal cell carcinoma is not thyroid related.

Frequency of IL-10-producing regulatory B cells associated with disease activity in thyroid-associated orbitopathy

AIM： To investigate the association between IL-10-producing regulatory B（B10） cells and the clinical features of thyroid-associated orbitopathy（TAO）. METHODS： A total of 30 patients with TAO were recruited at Zhongshan Ophthalmic Center from May 2015 to December 2015. Peripheral blood mononuclear cells（PBMCs） were separated from blood samples of 30 TAO patients and 16 healthy controls and stimulated with CD40 ligand and CpG for 48h. The frequency of IL-10＋ B cells was examined by flow cytometry and the correlation between the frequency of IL-10＋ B cells and clinical features of TAO was analyzed by SPSS. RESULTS： The frequency of IL-10＋ B cells among CD19＋ B cells in TAO patients was significantly lower than in healthy controls（TAO： 4.66%±1.88% vs healthy control： 6.82%±2.40%, P〈0.01）. The frequency of IL-10＋ B cells showed a positive correlation with disease activity of TAO measured by Clinical Activity Score（CAS）（r=0.50, P〈0.01）, and became higher in TAO patients with family history of Graves＇ disease（GD）（P=0.04）. CONCLUSION： The decrease of the frequency of IL-10＋ B cells in TAO patients indicates the deficiency of B10 cells in TAO, and the positive association with disease activity suggests its important role in TAO inflammation regulation.

Lenvatinib-induced multiorgan adverse events in Hurthle cell thyroid cancer: A case report

BACKGROUND The management of metastatic progressive radioiodine-resistant differentiated thyroid cancer remains challenging for clinicians.The availability of tyrosine kinase inhibitors(TKIs),sorafenib and lenvatinib,within the last decade has expanded treatment options;however,these lead to significant adverse effects,which may curtail their use.CASE SUMMARY We report the case of a 47-year-old female with Hurthle cell thyroid cancer who underwent total thyroidectomy followed by radioiodine ablation.During followup,she developed noniodine-avid renal and pulmonary metastases.With respect to her pre-existing diabetes,hypertension,and polycystic kidney disease,the tumor board decided against performing renal metastasectomy because of the risk of future renal decline requiring dialysis.Metastases were treated using sorafenib,which provided stability followed by progression within a year.We switched to lenvatinib,which led to disease regression.However,the patient experienced severe adverse effects,including cardiomyopathy,bicytopenia,renal impairment,and the rarely reported nephrotic syndrome.Renal metastasis is a rare manifestation of Hurthle cell thyroid cancer with only two reported cases in literature.We report the experience of our first case of renal metastasis and its treatment with TKIs.This case serves as a reminder of the adverse drug reactions associated with TKI use.CONCLUSION We advocate close monitoring of patients’hematological and renal profiles as well as their cardiac status using an echocardiogram.

Concurrent pulmonary Mycobacterium avium complex (MAC) infection and active Hürthle cell thyroid carcinoma: is there a connection?

We present two cases of pulmonary MAC infection in women with Hürthle cell thyroid carcinoma. Both cases were asymptomatic octogenarian women with active Hürthle cell thyroid carcinoma and prolonged periods of hypothyroidism prior to diagnosis of pulmonary MAC. Mycobacterium avium complex has never been reported in association with any type of thyroid cancer, specifically Hürthle cell carcinoma. A review of the literature and possible associations between the two are discussed in this article.

Fine-needle aspiration cytology of an intrathyroidal nodule diagnosed as squamous cell carcinoma: A case report

BACKGROUND Both squamous cell carcinoma(SCC)and papillary thyroid carcinoma(PTC)are common malignant tumors in the neck.However,seldom has SCC of the thyroid been diagnosed.Further,cytological features of SCC and PTC have rarely been reported.The significance of fine-needle aspiration cytology(FNAC)in the diagnosis of neck masses has been established.Herein,we present an exceedingly rare case of an intrathyroidal SCC diagnosed using FNAC,along with its cytological features.CASE SUMMARY A 66-year-old man presented with a left-sided neck mass.Ultrasound examination showed an ill-defined nodule.The appearance was hypoechoic with a few hyperechoic spots.FNAC of the left thyroid nodule was performed.A cellular smear was obtained,and it showed a large number of neoplastic cells with rich cytoplasm and poor cell adhesion.Tumor cell nuclei showed coarse nuclear chromatin and a few enlarged prominent nucleoli.An increased nuclear/cytoplasm ratio was observed.Thus,malignancy was diagnosed without a confirmed tumor type.Percutaneous tumor biopsy was performed to make a definite diagnosis.The tumor cells showed typical squamous cell characteristics.CONCLUSION Head and neck SCC and PTC have different cytologies.Measures are needed to ensure accurate diagnosis using FNAC.

Thyroid Metastases from Clear Cell Renal Carcinoma: Presentation of an Unusual Case and Literature Review

Clear cell type renal cell carcinoma is a malignant tumor that can metastasize to many locations, but unusually spreads to thyroid. Therefore it is important to know the oncological background of the patient and to perform a complete immunohistochemical analysis of the thyroid lesion to obtain a correct diagnosis. Thyroidectomy can be considered in patients with no other metastasis or in those who present compressive symptoms as a palliative measure. We expose the case of a patient with clear cell type renal cell carcinoma, with a previously known bone metastasis of renal origin and symptomatic goiter containing a nodule that was found to be a metastasic lesion.

Clear Cells Carcinoma of Thyroid: Difficult Diagnosis and Rare Case

Clear cell carcinoma of the thyroid is rare. We report a case observed in a 17-year-old woman with a nodule in the left lobe of the thyroid. Cervical ultrasound showed a single nodule of the left lobe of the thyroid, measuring 28 × 14 × 21 mm with microcalcifications, classified TIRADS 4. Pathological and immunohistochemical examination of the excisional specimen concluded with clear cell carcinoma. The epidemiological and histological aspects were discussed.

Scabrous patrinia root inhibits circulating tumor cells in differentiated thyroid carcinoma–a clinical observation

Objective To observe the clinical inhibition of circulating tumor cells(CTCs) in differentiated thyroid carcinoma(DTC) by the extract of scabrous patrinia root(Huikangling).Methods Eighty-seven DTC patients tested positive for CTCs were randomly divided into two groups; all patients were treated with oral levothyroxine sodium in accordance with the DTC endocrine inhibition treatment criteria. Patients(n = 45) in the treatment group were provided the standard endocrine therapy along with oral Huikangling(0.4 g/tablet, 0.4 g × 3/time, 3 times/day, 12 weeks). Patients(n = 42) in the control group were only provided the standard therapy. After 4 and 12 weeks, CTCs in the blood were detected by flow cytometry.Results After 4 weeks of oral Huikangling treatment, CTCs were detected in 18(40%) and 29(69%) patients in the treatment and control groups, respectively; the difference was statistically significant(χ2 = 8.49, P < 0.05). After 12 weeks, CTCs were detected in 7(15.6%) and 17(44.7%) patients in the treatment and control groups, respectively; the difference was statistically significant(χ2 = 5.68, P < 0.05). Follow-up evaluation revealed two patients with lung metastasis and one patient with bone metastasis in the control group; one patient showed lateral neck lymph node metastasis without local recurrence in the treatment group.Conclusion Huikangling treatment reduces the number of CTC-positive DTC cases; however, further studies are needed to elucidate the underlying mechanisms.

5例甲状腺嗜酸细胞癌患者的临床病理分析

目的:探讨甲状腺嗜酸细胞癌(thyroid oncocytic cell carcinoma,OCA)的临床病理学特征、诊断、鉴别诊断及预后。方法:收集5例OCA患者的临床病理资料,分析总结其病理形态学特征及其预后关系,并复习相关文献。结果:5例患者中3例为女性,2例为男性,年龄42~70岁,平均年龄53.4岁。肿瘤最大径2~7.5 cm,平均4.9 cm。镜下肿瘤均可见纤维性包膜,3例显示血管侵犯(其中2例同时存在包膜侵犯),其余2例仅见包膜侵犯,肿瘤细胞胞浆丰富、嗜酸性、颗粒状,可见核仁,未见甲状腺乳头状癌(papillary thyroid carcinoma,PTC)的核特征,核分裂象罕见,未见肿瘤性坏死。5例患者随访时间9~41个月,均无复发及转移。结论:甲状腺OCA是一种侵袭性肿瘤,发病率低,转移风险较高,预后较差,具有其独特的病理形态学特点及分子改变,需与其他伴有嗜酸性变的甲状腺肿瘤相鉴别。

补中益气汤对自身免疫性甲状腺炎小鼠脾脏树突状细胞免疫耐受状态的量效研究

目的研究补中益气汤对自身免疫性甲状腺炎(Autoimmune thyroiditis,AIT)小鼠脾脏树突状细胞(Dendritic cells,DCs)免疫耐受状态的影响及其量效关系。方法选取90只6^8周龄SPF级雌性NOD.H-2^(h4)小鼠,随机分为正常(NC)组、对照(AIT)组、补中益气汤高剂量(BZH)组、补中益气汤中剂量(BZM)组、补中益气汤低剂量(BZL)组、亚硒酸钠(SS)组,共6组,每组15只。BZH组每日予以16.38 g·kg^(-1)补中益气汤灌胃,BZM组每日予以8.19 g·kg^(-1)补中益气汤灌胃,BZL组每日予以4.10 g·kg^(-1)补中益气汤灌胃,SS组每日予以0.26 mg·kg^(-1)亚硒酸钠片灌胃,NC组及AIT组每日予以等体积蒸馏水灌胃,连续灌胃8周后,HE染色检测各组小鼠甲状腺淋巴细胞浸润情况;ELISA检测各组小鼠血清中甲状腺球蛋白抗体(Thyroglobulin antibody,TgAb)、TSH及脾脏IL-10水平;流式细胞术检测各组小鼠脾脏树突状细胞表面MHC-II及CD80、CD86、CD40分子表达,以及脾脏Th17/Treg细胞比例。结果(1)HE染色可见AIT组小鼠甲状腺淋巴细胞浸润显著,各治疗组甲状腺淋巴细胞浸润情况均有所恢复,其中BZM组小鼠每日灌胃8.19 g·kg^(-1)补中益气汤效果最为显著。(2)Elisa检测各组小鼠血清TgAb、TSH水平发现,AIT组小鼠血清TgAb较NC组显著升高,其余各治疗组均能够降低小鼠血清TgAb水平,其中BZH组效果最为明显,但BZH组与BZM剂量组之间差异没有统计学意义;与NC组相比,其余组小鼠血清TSH均有所升高,但各组间差异没有统计学意义。(3)AIT组小鼠脾脏IL-10分泌较NC组降低,其余各治疗组均能够升高脾脏IL-10的分泌水平,与TgAb结果相似,BZH组效果最为明显,但BZH组与BZM组之间差异没有统计学意义。(4)AIT组小鼠较NC组比较脾脏DCs表面因子CD40、CD80、CD86及MHC-Ⅱ表达升高,脾脏T细胞Th17/Treg水平升高,各治疗组脾脏DCs表面因子水平降低,Th17/Treg水平降低。结论补中益气汤能够降低AIT小鼠甲状腺自身抗体水平,改善甲状腺组织病理损伤,降低DCs表面CD80、CD86、CD40及MHC-II的表达,促进IL-10的分泌,维持Th17/Treg轴平衡,延缓AIT发生。推荐小鼠最佳灌胃剂量为9.18 g·kg^(-1)。

虎杖苷通过Hippo/YAP通路影响甲状腺癌8505C细胞的恶性生物学行为和顺铂敏感性

目的:探究虎杖苷通过Hippo/Yes相关蛋白(YAP)通路对人甲状腺癌8505C细胞的恶性生物学行为和顺铂(DDP)敏感性的影响。方法:体外培养8505C细胞,构建其DDP耐药细胞8505C/DDP,用CCK-8法检测0、25、50、75、100 nmol/L虎杖苷处理8505C和8505C/DDP细胞的增殖能力,以筛选虎杖苷的最佳作用浓度。将8505C细胞分为对照组、虎杖苷组、空载组、虎杖苷+YAP1过表达组;将8505C/DDP细胞分为对照组、DDP组、DDP+虎杖苷组、DDP+空载组、DDP+虎杖苷+YAP1过表达组。WB法检测各组8505C细胞中Hippo/YAP通路[YAP1、转录辅激活因子(TAZ)]和EMT(E-cadherin、N-cadherin)相关蛋白,8505C/DDP细胞中YAP1、TAZ、耐药相关蛋白[P-糖蛋白(P-gp)、多药耐药相关蛋白1(MRP1)]、凋亡相关蛋白(C-caspase-3、BAX、Bcl-2)的表达。Transwell小室和细胞划痕实验分别检测各组8505C、8505C/DDP细胞的侵袭、迁移能力。结果:虎杖苷可显著抑制8505C细胞的增殖活性(P<0.05)明显抑制8505C细胞中YAP1、TAZ蛋白、N-cadherin的表达(均P<0.05),提升E-caderin蛋白的表达(P<0.05),显著抑制8505C细胞的迁移和侵袭能力(均P<0.05),而8505C/DDP细胞对低浓度的虎杖苷具有耐药性(P<0.05);过表达YAP1则可逆转虎杖苷对8505C细胞的影响。50 nmol/L虎杖苷明显抑制DDP处理的8505C/DDP细胞中YAP1、TAZ、P-gp、MRP1、Bcl-2的蛋白的表达(均P<0.05),提升cleaved caspase-3、BAX蛋白的表达(均P<0.05)并诱导其细胞凋亡(P<0.05),过表达YAP1则可逆转虎杖苷对8505C/DDP细胞的影响。结论:虎杖苷抑制Hippo/YAP信号通路,从而抑制8505C细胞的恶性生物学行为和增强其对的DDP敏感性。

川楝素调节Notch信号通路对甲状腺癌细胞增殖、迁移和侵袭的影响

目的 研究川楝素通过调节Notch信号通路对甲状腺癌细胞增殖、迁移和侵袭的影响。方法 体外培养人甲状腺癌细胞8505C,CCK-8法检测不同剂量川楝素对8505C细胞存活率的影响;将8505C细胞分为对照组、川楝素-L、M、H组、川楝素+DAPT(Notch信号通路抑制剂)组;EDU法检测8505C细胞增殖;Transwell检测8505C细胞迁移与侵袭能力;RT-PCR检测不同8505C细胞组Notch1、HES1、HEY1、Ki67、MMP-2的mRNA水平;western blot检测不同8505C细胞组Notch1、HES1、HEY1、Ki67、MMP-2的蛋白水平。结果 与对照组相比,川楝素-L、M、H组8505C细胞EDU阳性细胞率、迁移与侵袭细胞数、Ki67、MMP-2的mRNA以及蛋白水平均显著性降低,Notch1、HES1、HEY1的mRNA以及蛋白水平显著性升高,且呈剂量依赖性(P<0.05);与川楝素-H组相比,川楝素+DAPT组EDU阳性细胞率、迁移与侵袭细胞数、Ki67、MMP-2的mRNA以及蛋白水平均显著性升高,Notch1、HES1、HEY1的mRNA以及蛋白水平显著性降低(P<0.05)。结论 川楝素可能通过激活Notch信号通路,抑制甲状腺癌细胞8505C增殖、迁移和侵袭。

miR-100-5p对甲状腺癌细胞增殖与凋亡调控作用的实验研究

目的通过实验探讨微小核糖核酸(microRNA,miR)-100-5p在甲状腺癌细胞中的表达情况及其对细胞增殖与凋亡的调控作用。方法使用荧光定量PCR检测miR-100-5p在甲状腺癌细胞系(TPC-1,KTC-1)与甲状腺正常细胞系(Nthy-ori3-1)中的相对表达情况。TPC-1细胞分别转染miR-100-5p模拟物(miR-100-5p mimic)、抑制物(miR-100-5p inhibitor)及相应阴性对照(miR-mimic NC,miR-inhibitor NC)后,用CCK-8检测TPC-1细胞增殖情况,流式细胞仪检测TPC-1细胞凋亡情况。通过miRTarBase和TargetScan7.2数据库对miR-100-5p的靶基因进行预测和功能富集分析,用蛋白印迹实验与双荧光素酶报告基因实验验证miR-100-5p对成纤维细胞生长因子受体3(fibroblast growth factor receptor 3,FGFR3)的靶向调控作用。结果与Nthy-ori3-1细胞相比,miR-100-5p在TPC-1细胞中表达水平(1.87±0.03 vs 1.00±0.03)与KTC-1细胞中表达水平(6.33±0.47 vs 1.00±0.03)均上调,差异具有统计学意义(t=-34.220,-19.588,均P<0.05)。转染miR-100-5p mimic组在24,48,72h细胞450nm吸光度(A_(450nm))均高于miR-mimic NC组,差异具有统计学意义(t=-7.516,-17.828,-8.445,均P<0.05);转染miR-100-5p inhibitor组在24,48,72h A_(450nm)均低于miR-inhibitor NC组,差异具有统计学意义(t=6.720,6.782,6.073,均P<0.05)。与miR-mimic NC组相比,转染miR-100-5p mimic后凋亡率(7.43%±0.49%vs 10.55%±0.80%)下降(t=5.767,P=0.004),与miR-inhibitor NC组相比,转染miR-100-5p inhibitor后凋亡率(3.19%±0.22%vs 2.64%±0.15%)上升(t=-3.606,P=0.023),差异均有统计学意义。蛋白印迹实验显示,与miR-mimic NC组相比,FGFR3在miR-100-5p mimic组蛋白表达水平(0.78±0.12 vs 1.00±0.00)下调(t=3.071,P=0.037),与miR-inhibitor NC组相比,FGFR3在miR-100-5p inhibitor组蛋白表达水平(1.17±0.07 vs 1.00±0.00)上升(t=-4.509,P=0.046),差异均有统计学意义。与miR-mimic NC相比,miR-100-5p mimic没有降低FGFR33’UTR野生型组荧光素酶活性(1.01±0.17 vs 1.00±0.00)与突变型组荧光素酶活性(0.99±0.11 vs 1.00±0.00),差异无统计学意义(t=-0.057,0.181,P=0.96,0.873)。结论miR-100-5p在甲状腺癌细胞中表达上调,可促进甲状腺癌细胞增殖、抑制细胞凋亡,其可能成为甲状腺癌诊疗中新的生物标志物与调控靶点。

Hsa_circ_0016600通过调控miR-149/PTK7轴促进甲状腺癌细胞迁移的机制

目的:探究环状RNA hsa_circ_0016600对甲状腺癌细胞迁移的影响及其作用机制。方法:收集温州医科大学附属第二医院颈部外科收治的15例甲状腺癌患者的癌组织及正常组织标本,采用RT-qPCR法检测hsa_circ_0016600、miR-149和PTK7在甲状腺癌组织及甲状腺细胞系中的表达情况。体外培养甲状腺癌细胞系,分别敲低hsa_circ_0016600和过表达miR-149,采用CCK-8、Transwell和划痕实验检测甲状腺细胞增殖、迁移和侵袭能力。建立小鼠异种移植皮下瘤模型,观察敲低hsa_circ_0016600的甲状腺癌细胞在体内的成瘤作用,并利用荧光原位杂交实验检测hsa_circ_0016600在裸鼠移植瘤模型中的表达水平。采用RNA荧光原位杂交实验检测hsa_circ_0016600和miR-149在甲状腺癌细胞中的定位,双荧光素酶报告基因实验检测hsa_circ_0016600与miR-149及miR-149与PTK7的靶向调控关系。结果:在人甲状腺癌组织和细胞系中,hsa_circ_0016600(P<0.05)和PTK7表达上调(P<0.01),miR-149表达下调(P<0.05)。敲低hsa_circ_0016600表达后,甲状腺癌细胞的增殖(P<0.01)、侵袭(P<0.01)和迁移能力(P<0.01)受到抑制。hsa_circ_0016600可通过靶向miR-149促进甲状腺癌细胞的增殖(P<0.01)、迁移(P<0.01)和血管形成,并上调PTK7的表达(P<0.01)。此外,PTK7是miR-149的一个靶基因。结论:Hsa_circ_0016600在甲状腺癌组织和细胞中高表达,并通过调控miR-149/PTK7轴促进甲状腺癌细胞的迁移。