Skull base chordoma is a rare and aggressive bone tumor with a poor prognosis. The basement membrane (BM) plays an pivotal role in tumor progression. However, the involvement of BM-related genes in assessing the prognosis and influencing the biological behavior of skull base chordomas remains unclear. Patients with skull base chordoma undergoing endoscopic endonasal surgery were included in the study (77 patients for bulk transcriptome sequencing and 6 patients for single-cell RNA sequencing). A BM-related genes signature was established and validated using bulk transcriptome data. Additionally, we investigated the oncogenic potential of a key BM-related gene in chordoma cells in vitro. A prognostic signature consisting of five BM-related genes was identified through LASSO Cox regression analysis. The accuracy and reliability of this signature were validated by the validation cohort. Multivariate Cox analysis and a nomogram demonstrated that the risk score serves as an independent and reliable prognostic factor for skull base chordoma. Moreover, the BM-related gene signature was significantly associated with the immune microenvironment, immune checkpoint expression, and drug sensitivity. Single-cell RNA sequencing analysis revealed both the chordoma tumor cell and the fibroblast contributed to the overall BM signature. Finally, in vitro experiments demonstrated that the knockdown of ITGB3, the hub gene in the signature, inhibited the proliferation and migration of chordoma cells via the PI3K-Akt pathway. This study explored the critical role of BM-related genes in skull base chordoma, which affected postoperative recurrence and maligant behavior of chordoma via the PI3K-Akt signaling pathway.

The clinical diagnosis and treatment of pituitary neuroendocrine tumors (PitNETs) that invade the cavernous sinus are fraught with difficulties and challenges. Exploring the biological characteristics involved in the occurrence and development of PitNETs that invade the cavernous sinus will help to elucidate the mechanism of cavernous sinus invasion. There are differences between intrasellar tumors (IST) and cavernous sinus-invasion tumors (CST) in ultramicrostructure, tumor microenvironment (TME), gene expression, and signaling pathways. The microvascular endothelial cell is increased in CST. The VEGFR signaling pathway, VEGF signaling pathway, and chemokine signaling pathway are activated in CST. HSPB1 is upregulated in CST and promotes cell proliferation, cell viability, and migration. HSPB1 promotes the release of VEGF from GT1-1 cells and activates the VEGF signaling pathway in bEnd.3 cells. HSPB1 promotes the migration of bEnd.3 cells to GT1-1 cells and promotes the formation of blood vessels of bEnd.3 cells. bEnd.3 cells can release CCL3 and CCL4 and promote the vitality, proliferation, and migration of GT1-1 cells. HSPB1 promotes the formation of blood vessels of bEnd.3 cells and ultimately leads to tumor growth in vivo. HSPB1 acts as a key gene for invasion of the cavernous sinus in PitNETs, remodeling TME by promoting the formation of blood vessels of brain microvascular endothelial cells. The synergistic effect of tumor cells and microvascular endothelial cells promotes tumor progression. The mechanism by which HSPB1 promotes tumor invasion by inducing angiogenesis in PitNETs may be a new target for the treatment of PitNETs invading the cavernous sinus.

Background and Objectives: The diagnosis and treatment of pituitary adenomas with cavernous sinus invasion pose significant challenges for clinicians. The objective of this study is to investigate the expression profile and prognostic value of HSPB1 (heat shock protein beta-1) in pituitary adenomas with invasive and non-invasive features. Additionally, we aim to explore the potential relationship between HSPB1 expression and immunological functions in pituitary adenoma. Materials and Methods: A total of 159 pituitary adenoma specimens (73 invasive tumours and 86 non-invasive tumours) underwent whole-transcriptome sequencing. Differentially expressed genes and pathways in invasive and non-invasive tumours were analysed. HSPB1 was subjected to adequate bioinformatics analysis using various databases such as TIMER, Xiantao and TISIDB. We investigated the correlation between HSPB1 expression and immune infiltration in cancers and predicted the target drug of HSPB1 using the TISIDB database. Results: HSPB1 expression was upregulated in invasive pituitary adenomas and affected immune cell infiltration. HSPB1 was significantly highly expressed in most tumours compared to normal tissues. High expression of HSPB1 was significantly associated with poorer overall survival. HSPB1 was involved in the regulation of the immune system in most cancers. The drugs DB11638, DB06094 and DB12695 could act as inhibitors of HSPB1. Conclusions: HSPB1 may serve as an important marker for invasive pituitary adenomas and promote tumour progression by modulating the immune system. Inhibitors of HSPB1 expression are currently available, making it a potential target for therapy in invasive pituitary adenoma.

目的 分析Delta样蛋白1同源物(DLK1)在生长激素(GH)腺瘤中的表达情况及其临床意义.方法 本实验时间为2020年10月至2022年6月.肿瘤标本来自2016—2020年于首都医科大学附属北京天坛医院神经外科行手术切除的34例GH腺瘤患者.采用免疫组织化学染色检测肿瘤标本中DLK1、GH表达情况.取对数生长期的大鼠GH3细胞,将其随机分为A组、B组、C组、D组〔B组、C组、D组分别加入1、5、20μg/ml抗DLK1抗体,A组加入等体积的二甲基亚砜(DMSO)〕,分别于培养0、24、48、72 h后采用细胞增殖实验检测各组细胞活力.取对数生长期的大鼠GH3细胞,将其随机分为E组、F组(F组加入5μg/ml抗DLK1抗体,E组加入等体积的DMSO),分别于培养0、24、48、72 h后采用ELISA检测各组GH3细胞培养上清液中GH水平.取对数生长期的大鼠GH3细胞,将其随机分为G组、H组、I组、J组(H组、I组、J组分别加入1、5、20μg/ml抗DLK1抗体,G组加入等体积的DMSO),采用Western blot法检测各组GH3细胞中磷酸化p70核糖体蛋白S6激酶(p-p70S6K)、磷酸化起始因子4E结合蛋白1(p-4EBP1)、磷酸化雷帕霉素靶蛋白(p-mTOR)水平.结果 DLK1主要位于稀疏颗粒型肿瘤标本的细胞核及致密颗粒型肿瘤标本的细胞质,GH主要位于疏松颗粒型、致密颗粒型肿瘤标本的细胞质.Pearson相关分析结果显示,肿瘤标本的DLK1评分与GH评分呈正相关(r=0.550,P<0.001).根据DLK1评分中位数将肿瘤标本分为高DLK1评分组(≥110分,n=17)和低DLK1评分组(<110分,n=17).高DLK1评分组患者血清GH及肿瘤标本GH评分、临床表型为致密颗粒型者占比高于低DLK1评分组(P<0.05).B组培养48、72 h后细胞活力高于A组,C组、D组培养24、48、72 h后细胞活力高于A组(P<0.05);C组培养48 h后细胞活力高于B组,D组培养24、48、72 h后细胞活力高于B组(P<0.05);D组培养48、72 h后细胞活力高于C组(P<0.05).A组、B组、C组、D组培养24、48、72 h后细胞活力分别高于本组培养0 h后,培养48、72 h后细胞活力分别高于本组培养24 h后,培养72 h后细胞活力分别高于本组培养48 h后(P<0.05).F组培养48、72 h后GH3细胞培养上清液中GH水平低于E组(P<0.05).E组培养48、72 h后GH3细胞培养上清液中GH水平高于本组培养0 h后,F组培养48、72 h后GH3细胞培养上清液中GH水平低于本组培养0 h后(P<0.05);E组培养72 h后GH3细胞培养上清液中GH水平高于本组培养24 h后,F组培养72 h后GH3细胞培养上清液中GH水平低于本组培养24 h后(P<0.05).H组、I组、J组GH3细胞中p-p70S6K水平高于G组,J组GH3细胞中p-p70S6K水平低于H组、I组(P<0.05);I组、J组GH3细胞中p-4EBP1水平低于G组、H组(P<0.05);H组、I组、J组GH3细胞中p-mTOR水平高于G组,I组、J组GH3细胞中p-mTOR水平高于H组,J组GH3细胞中p-mTOR水平高于I组(P<0.05).结论 DLK1主要在致密颗粒型GH腺瘤中表达;DLK1可抑制GH3细胞增殖,增加血清GH水平,其机制可能与DLK1可抑制GH3细胞中p70核糖体蛋白S6激酶、雷帕霉素靶蛋白的磷酸化及促进起始因子4E结合蛋白1的磷酸化有关.

Introduction: Aberrant miR-320a has been reported to be involved in the tumorigenesis of several cancers. In our previous study, we identified the low expression of circulating miR-320a in patients with somatotroph pituitary neuroendocrine tumor (PitNET); however, the role of miR-320a in somatotroph PitNET proliferation is still unclear. Methods: Cell viability and colony formation assays were used to detect the effect of miR-320a and BCAT1 on GH3 cells. TargetScan was used to identify the target genes of miR-320a. Dual-luciferase reporter gene assay was used to explore the relation between miR-320a and BCAT1. Transcriptome and proteome analyses were performed between somatotroph PitNETs and healthy controls. The expression level of miR-320a in somatotroph PitNETs were detected by RT-qPCR and Western blot. Results: miR-320a mimics inhibit cell proliferation, while miR-320a inhibitors promote cell proliferation in GH3 cells. An overlap analysis using a Venn diagram revealed that BCAT1 is the only target gene of miR-320a overexpressed in somatotroph PitNETs compared to healthy controls, as revealed by both microarray and proteomics results. A dual-luciferase reporter gene assay showed that miR-320a may bind to the BCAT1-3′UTR. The transfection of miR-320a mimics downregulated the expression and miR-320a inhibitors and upregulated the expression of BCAT1 in GH3 cells. The interference of BCAT1 expression in GH3 cells downregulated cell proliferation and growth. Pan-cancer analyses demonstrated that high BCAT1 expression often indicates a poor prognosis. Conclusion: Our findings illustrate that miR-320a may function as a tumor suppressor and BCAT1 may promote tumor progression. miR-320a may inhibit the growth of somatotroph PitNETs by targeting BCAT1.

Background and Objective: Non-functioning pituitary neuroendocrine tumors (NF-PitNETs) represent a heterogeneous tumor type that lacks effective medical treatment. MDM2, the main negative regulator of p53, binds to and forms a stable complex with p53 to regulate its activity. In this study, we measured the expression levels and role of MDM2 in non-functioning PitNET patients' combined clinical features and investigated the effect of etoposide on the cell bioactivity of the GT1-1 cell line in vivo and in vitro. Methods: RT-PCR and immunochemistry measured the expression levels and role of MDM2 in 103 NF-PitNET patients' combined clinical features. Cell proliferation, migration, colony and apoptosis experiments measured the effect of etoposide on the GT1-1 cell line in vivo and in vitro. Results: There was more invasive behavior (p = 0.013) in patients with high MDM2, who were also younger (p = 0.007), were more frequently female (p = 0.049) and had larger tumor sizes (p = 0.018) compared with patients with low MDM2. Patients with high p53 were younger (p = 0.017) and had larger tumor sizes (p = 0.034) compared with patients with low p53. Univariate (p = 0.018) and multivariate (p = 0.023) Cox regression analysis showed that MDM2 was the independent factor for invasive behavior in NF-PitNET patients. Log-rank analysis showed that the average progression-free survival (PFS) time in the low MDM2 patients was longer than that in the high MDM2 patients (p = 0.044). Functional studies indicated that etoposide inhibited cell proliferation and cell migration and induced apoptosis in p53 independence in GT1-1 cells. Furthermore, etoposide significantly inhibited the growth of GT1-1-xenograft in BALB/c nude mice. The tumor growth inhibition rate of etoposide was 67.4 ± 4.6% after 14 d of treatment, which suggested the anti-tumor activity of etoposide. Conclusions: MDM2 played the role of tumorigenesis of NF-PitNET in a p53 independence manner, and an MDM2 inhibitor could be a potential choice for the treatment of NF-PitNET patients.

Ubiquitination is reported to be a critical biological event on ACTH secretion in corticotroph adenomas. However, the effect of ubiquitylation on ACTH secretion in silent corticotroph adenomas (SCAs) remains unclear. The aim of our study was to explore the mechanism of decreased secretion of ACTH in SCAs with ubiquitinomics. The differently expressed ubiquitinated proteins between SCAs and functioning corticotroph adenomas (FCAs) were identified by 4D label-free mass spectrometer, followed by bioinformatics analysis. The function of the candidate ubiquitinated protein ATP7A (K333) was validated in AtT20 cells. A total of 111 ubiquitinated sites corresponding to 94 ubiquitinated proteins were typically different between SCAs and FCAs. Among all the ubiquitinated sites, 102 showed decreased ubiquitination in SCAs, which mapped to 85 ubiquitinated proteins. Pathway enrichment analysis revealed that ubiquitinated proteins were mainly enriched in vesicle pathway and protein secretion pathway. ATP7A (K333) was one of the proteins enriched in vesicle pathway and protein secretion pathway with decreased ubiquitination level in SCAs. In vitro assay indicated that both ATP7A siRNA and omeprazole (ATP7A protein inhibitor) increased the secretion of ACTH in AtT20 cell supernatant compared to control groups (p<0.05). These results indicated that ATP7A might be related to the abnormal expression of ACTH in SCAs and potential for the treatment of SCAs.

Objective:To determine risk factors and management for the development of a postoperative cerebrospinal fluid (CSF) leak after an endoscopic endonasal surgery (EES) for pituitary adenomas.Methods:The clinical data of 400 patients who underwent EES for resection of pituitary adenomas from December 2018 to November 2019 in the Department of Neurosurgery of Beijing Tiantan Hospital were retrospectively reviewed. Age, gender, body mass index (BMI), tumor size, Knosp grade, suprasellar extension grade, sellar floor erosion grade, repeated transsphenoidal surgery, intraoperative CSF leak, use of pedicled nasoseptal flap and lumbar drain were collected and analyzed.Results:Postoperative CSF leak occurred in 14 of 400 patients (3.5%). Age, gender, BMI, tumor size, Knosp grade and repeated transsphenoidal surgery were not risk factors for CSF leak. Suprasellar extension grade (≥B 6.0% vs. <B 1.4%; p = 0.024), sellar floor erosion grade (≥III 5.7% vs. <III 0.6%; p = 0.020) and intraoperative CSF leak (Yes 7.5% vs. No 2.0%; p = 0.009) were factors associated with an increased postoperative CSF leak rate.Conclusions:Higher suprasellar extension grade, higher sellar floor erosion grade and intraoperative CSF leak were risk factors for postoperative CSF leak after endoscopic treatment of pituitary adenoma. Strict skull base reconstruction including use of a pedicled nasoseptal flap and perioperative lumbar drainage may avoid postoperative CSF leak.

目的 探讨生长激素腺瘤血清Micr0-RNA(miRNA)的表达情况,及miR-423-5p对生长激素腺瘤增殖的影响.方法 各检测6例生长激素腺瘤病人和正常人血清外泌体miRNA的表达情况,对比两者之间的差异.结果 外泌体miRNA表达谱显示:生长激素腺瘤病人和正常人血清之间有169个差异表达miRNA(P<0.05,差异表达倍数>2倍).在169个差异表达的miRNA中,miR-423-5p在生长激素腺瘤病人中的表达量明显低于正常人(P<0.05).这一结果在qPCR阵列芯片实验结果中也得到证实.体外实验证实:miR-423-5p能够抑制GH3细胞增殖,诱导细胞凋亡,减少生长激素释放和细胞迁移.结论 miR-423-5p在生长激素腺瘤中低表达,具有促进肿瘤细胞增殖的作用.

Pituitary adenoma and meningioma are two of the most common benign tumors in the central nervous system. Pituitary adenoma associated with meningioma (PAM) is a rare disease, the tumorigenesis of which remains unclear. Therefore, the aim of the present study was to investigate the tumorigenesis of PAM. A total of 8,197 patients with pituitary adenoma were analyzed. Furthermore, the clinical data of 57 patients with PAM were compared with patients with multiple endocrine neoplasia 1 (MEN-1) syndrome. Whole exome sequencing (WES) was performed on 23 samples from patients with PAM and the germline mutation was verified by Sanger sequencing. The age of tumor penetrance (age of patients at diagnosis) for PAM was significantly higher than that for patients with MEN-1. Compared with MEN-1 patients, there was a significant association between PAM and female sex (P=0.004). Clonal analysis and phylogenetic tree construction suggested that the pituitary adenoma and meningioma in PAM don't originate from a common progenitor. WES revealed that 5/23 PAM samples had the recurrent germline mutation MEN1 c.1523G>A; p.G508D, which may be a genetic risk factor for PAM. Compared with patients with sporadic pituitary adenoma, the difference was statistically significant (P=0.0004). Compared with wild-type MEN1, there was a significant association between the MEN1 mutation and recurrence of pituitary adenoma, young age and larger diameter of the meningioma. The present study indicated that germline mutations in MEN1 may be associated with the tumorigenesis of PAM.

The genetic basis and corresponding clinical relevance of prolactinomas remain poorly understood. Here, we perform whole genome sequencing (WGS) on 21 patients with prolactinomas to detect somatic mutations and then validate the mutations with digital polymerase chain reaction (PCR) analysis of tissue samples from 227 prolactinomas. We identify the same hotspot somatic mutation in splicing factor 3 subunit B1 (SF3B1(R625H)) in 19.8% of prolactinomas. These patients with mutant prolactinomas display higher prolactin (PRL) levels (p=0.02) and shorter progression-free survival (PFS) (p=0.02) compared to patients without the mutation. Moreover, we identify that the SF3B1(R625H) mutation causes aberrant splicing of estrogen related receptor gamma (ESRRG), which results in stronger binding of pituitary-specific positive transcription factor 1 (Pit-1), leading to excessive PRL secretion. Thus our study validates an important mutation and elucidates a potential mechanism underlying the pathogenesis of prolactinomas that may lead to the development of targeted therapeutics.

目的 探寻垂体嗜酸性细胞瘤线粒体功能异常相关蛋白.方法 本研究样本取材于行手术治疗的14例垂体嗜酸性细胞瘤,正常垂体组织取材于8例交通意外后尸体捐献者.采用超高效液相色谱和蛋白质质谱分析垂体嗜酸性细胞瘤和正常垂体之间的差异蛋白质.进一步通过生物信息学分析方法,识别与垂体嗜酸性细胞瘤线粒体功能异常相关的蛋白及蛋白信号通路.采用实时定量PCR方法验证线粒体功能异常相关的蛋白VDAC1、VDAC2、VDAC3、SLC25A4、SLC25A5和pG-1β在垂体嗜酸性细胞瘤和正常垂体的表达情况.结果 蛋白质质谱分析结果显示:598个蛋白质在垂体嗜酸性细胞瘤和正常垂体的表达差异有统计学意义(均P＜ 0.05);其中331个蛋白质在垂体嗜酸性细胞瘤中上调,267个蛋白质在垂体嗜酸性细胞瘤中下调.IPA生物信息学分析结果显示:最显著富集的经典通路是线粒体功能异常通路.选择该信号通路中的蛋白质VDAC1、VDAC2、VDAC3、SLC25A4、SLC25A5和PGC-1β进行进一步验证.QPCR结果显示:VDAC2、VDAC3和PGC-1β在垂体嗜酸性细胞瘤表达显著高于正常垂体(P分别为0.0011,0.045,0.0003).结论 在垂体嗜酸性细胞瘤中,与线粒体功能异常相关蛋白表达升高,且线粒体合成蛋白PGC-1β表达升高.

Objective To identify the role of abnormal metabolism signaling pathways in growth hormone-secreting pituitary adenomas (GH-PAs) based on phosphoproteome profiles.Methods In this study,twelve GH-PAs specimens were collected from patients who were surgically treated at Department of Neurosurgery,Beijing Tiantan Hospital,Capital Medical University from August 2010 to August 2017.Normal pituitary tissues were collected from 3 body donors who died from traffic accidents.The differential phosphoprotein expression patterns involved in GH-PAs were investigated by nano LC-MS/MS in a group of samples.The phosphoprotein expression data were analyzed by bioinformatics.The expression levels of the candidate phosphoproteins were validated by western blot analysis in another group of samples.Results A total of 1 213 phosphorylated protein sites corresponding to 667 proteins were significantly different between GH-PAs and healthy pituitary glands (P ＜ 0.05,FC ≥ 1.5 or ＜ 0.67).Among those phosphorylated sites,342 exhibited lower levels of phosphorylation in GH-PAs and 871 exhibited lower levels of phosphorylation in GH-PAs.The 1 213 phosphorylated protein sites were mapped to 667 phosphoproteins.The majority of differentially expressed phosphorylated proteins were significantly enriched in glycolysis and the AMPK signaling pathway in GH-PAs by IPA (P ＜ 0.05).In glycolysis pathway,9 of 10 phosphorylated proteins exhibited lower phosphorylation levels in GH-PAs than healthy pituitary glands.The AMPK signaling pathway was demonstrated to be inhibited in GH-PAs by Pathway Activity Analysis.Notably,western blot proved that the level of AMPK(1.75 ± 0.36 vs.1.59 ± 0.75,P =0.663) had no difference and the phosphorylated levels of AMPK (ser496) was significantly lower in GH-PAs than in healthy pituitary glands(1.47 ± 0.58 vs.0.32 ±0.16,P =0.016).Conclusion Phosphorylation proteomics revealed the abnormal metabolism in GH-PAs.Decreased phosphorylation of the AMPK pathway may be critical for glucose metabolism in GH-PAs.

BackgroundPituitary adenomas are common brain tumors. Although transsphenoidal surgery are able to achieve extensive tumor removal, the rate of recurrence ranges from 5 to 20% depending on the different subtype. Further understanding of these tumors is needed to develop novel strategies to improve the prognosis of patients. But their metabolic characteristics are largely unknown.MethodsWe used metabolomic, transcriptomic, and proteomic approaches to systematically investigate eight subtypes of pituitary adenomas and normal pituitary glands. By blocking IDH2, we investigate IDH2 play an inhibitory role in GH tumor cell growth and tumor secretion.ResultsWe found that all of the pituitary adenomas displayed downregulated glucose metabolism and glycolysis compared to normal tissues. Together with the differences in amino acids and fatty acids, we categorized these tumors into three clusters. We then re-established the reprogrammed metabolic flux in pituitary adenomas based on multiomic analyses. Take growth hormone-secreting pituitary adenomas as an example, we revealed that IDH2 is a key player in the reprogrammed metabolism of such tumors. By blocking IDH2, we confirmed that IDH2 is a potential target for the inhibition of tumor cell growth and tumor secretion.ConclusionsOur study first uncovered the metabolic landscape of pituitary adenomas and demonstrated a possible way to inhibit tumor growth by regulating aberrant metabolism.

Background To investigate the relationship between cyclin B1 (CCNB1) gene expression and cavernous sinus invasion in pituitary adenomas. Methods Twenty-four pituitary adenoma tissue samples were examined by RT-qPCR and Western blot to assess the mRNA expression levels and protein levels of CCNB1, E-cadherin and N-cadherin. Correlation analyses between the expression levels of E-cadherin, N-cadherin and CCNB1 were performed. After lentivirus-mediated knockdown of CCNB1 in rat pituitary adenoma cell lines (GH3 and GT1-1), cell function changes were studied. The relationship between CCNB1 and epithelial-mesenchymal transition (EMT) was further verified by animal experiments. Results CCNB1 and N-cadherin gene expression were significantly higher in the invasive pituitary adenomas than in the non-invasive pituitary adenomas. Conversely, E-cadherin expression in the invasive pituitary adenomas was significantly lower. CCNB1 gene expression was downregulated in the GH3 and GT1-1 pituitary adenoma cell lines; N-cadherin expression was also decreased, but E-cadherin expression was increased. These results were confirmed in vivo. After downregulation of CCNB1, cell invasion and migration was significantly reduced in Transwell experiments. Conclusion High CCNB1 expression in pituitary adenoma affects cavernous sinus invasion through EMT.

BACKGROUND:Pituitary adenoma and meningioma are the most common benign tumors in the central nervous system. Pituitary adenoma associated with meningioma (PAM) is a rare disease and the clinical features and mechanisms of PAM are unclear.METHODS:We summarized the clinical data of 57 PAM patients and compared with sporadic pituitary adenoma (SPA) and sporadic meningioma (SM). 5 pituitary adenomas of PAM and 5 SPAs were performed ceRNA microarray. qRT-PCR, Western Blot, siMEN1 and rapamycin inhibition experiment were validated for ceRNA microarray.RESULTS:Clinical variable analyses revealed that significant correlations between PAM and female sex as well as older age when compared with SPA and significant correlations between PAM and transitional meningioma as well as older age when compared with SM. Additionally, the characteristics of PAM were significantly different for MEN1 patients. Functional experiments showed lower expression of MEN1 can upregulate mTOR signaling, in accordance with the result of ceRNA microarray. Rapamycin treatment promotes apoptosis in primary pituitary adenoma and meningioma cells of PAM.CONCLUSIONS:MEN1 plays an important role in PAM by upregulating mTOR signaling pathway. Rapamycin represents a potential therapeutic strategy for PAM in the future.

Circulating miRNAs are novel disease biomarkers that are valuable for diagnosis and prognosis. But the circulating miRNAs profile in somatotroph adenomas is still unknown. Therefore, serum exosomal miRNAs expression profiling in somatotroph adenomas was performed on 6 somatotroph adenomas and 6 normal controls. From the exosomal miRNAs expression profiling, we found 169 miRNAs differently expressed between somatotroph adenomas and healthy pituitary samples (p< 0.05, FC > 2). Among the 169 miRNAs, miR-423-5p was expressed lower in somatotroph adenomas than in healthy pituitary samples, which was proved by miRSCan Panel Chip™ qPCR. PTTG1 and SYT1 were the target mRNAs of miR-423-5p, and transcriptomics and proteomics profile both indicated the high expression of PTTG1 and SYT1 in somatotroph adenomas. H-scores were 223.1 ± 34.7 for PTTG1 and 163.4 ± 42.3 for SYT1 in 62 somatotroph adenomas specimens and 84.2 ± 21.3 for PTTG1 and 47.4 ± 17.2 for SYT1 in 6 healthy pituitary specimens by IHC. miR-423-5p inhibited the expression of SYT1 and PTTG1 at the mRNA and protein levels. Dual luciferase reporter gene assay shown was significantly reduced in the presence of miR-423-5p in GH3 cells transfected with wild-type PTTG1 3'UTR luciferase reporter plasmid but not reduced when transfected with the mutation PTTG1 3'UTR luciferase reporter plasmid (p<0.01). In vitro experiments showed that miR-423-5p induced cell apoptosis, inhibited cell proliferation, and reduced growth hormone release and migration of GH3 cells. The activity of miR-423-5p in GH3 cell was nearly blocked by its inhibitor. These results verified the central role of low miR-423-5p in promoting tumorigenesis in somatotroph adenomas. PTTG1 may act as biomarkers for clinical treatment of somatotroph adenomas.

The 14-3-3 protein family has attracted much attention in research into the pathogenesis of human tumors because of its involvement in tumorigenesis. In previous studies, we found that 14-3-3η was highly expressed in pituitary oncocytoma. However, the mechanism by which 14-3-3η regulates tumorigenesis in pituitary oncocytoma is unclear. 14-3-3η-binding proteins were investigated in pituitary oncocytoma by immunoprecipitation and proteomic analysis. A total of 443 proteins were identified as 14-3-3η binding proteins. The interactions of 14-3-3η and its binding partners were identified by a network analysis using the STRING database. The network included 433 nodes and 564 edges. PRAS40 (AKT1S1) was a binding protein of 14-3-3η and showed experimental interactions with 14-3-3η in the STRING database. The combined score was 0.407, which suggested a functional link. The 443 binding proteins of 14-3-3η showed enriched molecular signatures in GSEA and GO analysis. PRAS40 (AKT1S1) was enriched in the mTOR signaling pathway. Western blot analysis showed that the relative expression of p-PRAS40 (T246)/PRAS40 was significantly higher in pituitary oncocytoma than in normal pituitary tissues (p < 0.05). R18, a 14-3-3 protein inhibitor, inhibited MMQ cell proliferation after treatment with 8 μM R18 for 48 h compared to the control group (p < 0.01). These results suggest that 14-3-3η may be involved in promoting tumorigenesis in pituitary oncocytoma by interacting with PRAS40 (T246) via the mTOR signaling pathway.

The pituitary gland is a small but important organ located in the base of the brain. Although mostly noncancerous, pituitary adenomas (PAs) can cause serious health problems such as headaches, visual field defects, double vision, and hypopituitarism by invasion of regional structures. Nonfunctioning PAs (NFPAs) approximately account for one-third of PAs manifested by no circulating hormone hypersecretion. Lipid reprogramming has been recognized as a hallmark of tumor cells and proven to play a crucial role in tumorigenesis. However, the lipid molecular pathogenesis of NFPAs has remained obscure to date. To uncover lipid alterations that may contribute to the development of NFPAs and define their molecular characteristics, we investigated tissue lipids of patients with NFPAs including eight null cell adenomas (NCAs) and eight oncocytomas (OCMs) and of five normal pituitary glands as the control (Ctrl) using nontargeted lipidomics based on ultrahigh-performance liquid chromatography-Orbitrap Q-Exactive HF mass spectrometry. The lipidomic results were further validated in another set of subjects consisting of 8 NCAs, 10 OCMs, and 6 Ctrls to define crucial lipids discriminating NFPAs from the normal pituitary tumors. Lipidomic analyses revealed that OCM showed more pronounced changes in lipid compositions than NCA and Ctrl. As expected, mitochondria abundant cardiolipins were remarkably increased in OCM, which was accordant with the biochemical evidence of mitochondria hyperplasia in OCM. Significantly increased levels of phospholipids (PLs), especially arachidonic acid (AA)-enriched PLs, were unique characteristics of lipid profiling in OCM vs Ctrl. Our results indicate that AA-PLs may have diagnostic potential for OCM.

目的 观察ERK1/2通路抑制剂U0126对大鼠生长激素垂体腺瘤GH3细胞增殖的影响,以探索ERK1/2通路作为生长激素腺瘤潜在治疗靶点的可行性.方法 将大鼠GH3细胞分为对照组和U0126组,U0126组给予不同剂量(10、20、40nM)的U0126,对照组给予等体积二甲基亚砜(DMSO).细胞增殖实验(MTS法)检测细胞活力的变化;凋亡试剂盒检测细胞凋亡水平,Western-blot检测p-ERK1/2蛋白的水平.结果 与对照组相比,不同剂量U0126处理24 h的GH3细胞细胞活力分别下降14％、15％和19％;48h下降19％、28％和36％;72 h下降24％、37％和52％.处理72 h后,U0126组膜联蛋白V(Annexin V)阳性细胞占总细胞数的比率分别为5.3％、11.4％和18.5％;碘化丙啶(PI)阳性细胞的比率分别为3.7％、7.3％和10.5％.处理24 h后U0126不同剂量组ERK的磷酸化蛋白表达水平呈下降趋势,40 nM组为对照组的41.7％,差异具有统计学意义(P＜0.05).结论 ERK1/2通路抑制剂可以诱导大鼠GH3细胞凋亡,抑制细胞增殖.