Clinical Significance of Eukaryotic Initiation Factor 4E (eIF4E) Level among Cases Suffering Basal Cell Carcinoma of Skin.

BACKGROUND Eukaryotic initiation factor 4E (eIF4E) has been reported to act as a prognostic biomarker in various cancers, but its actual effect on basal cell cancer (BCC) of the skin is rarely reported. Our research measured eIF4E levels and discussed its consequence in BCC of the skin. MATERIAL AND METHODS Semi-quantitative real-time polymerase chain reaction (RT-PCR) and western blotting analysis were used to detect relative expression level of eIF4E in specimens at both mRNA and protein levels. The relationship of eIF4E level with clinical profiles was analyzed via chi-square test. Additionally, prognostic value of eIF4E was analyzed via Kaplan-Meier and cox regression analysis. RESULTS We found that eIF4E was over-expressed in tumor tissues, in comparison to bordering cancer-free tissue samples. Besides, elevated eIF4E level exhibited a strong relation to metastasis, TNM stage, and differentiation. Kaplan-Meier analysis revealed cases harboring high eIF4E levels faced shortened overall survival compared to cases of low levels (log rank test, P=0.018). Moreover, eIF4E could act as an independent biomarker for the prognosis of BCC of the skin, according to Cox regression analysis. CONCLUSIONS The level of eIF4E was upregulated and significantly correlated with the development of BCC of the skin. Thus, it might be a promising prognostic biomarker and therapy target for BCC of the skin.

miR-339-5p inhibits metastasis of non-small cell lung cancer by regulating the epithelial-to-mesenchymal transition.

Metastasis is a common event in cancer pathology, and represents the primary cause of cancer-associated mortality. Metastasis, which is the process in which cancer cells at the primary tumor site spread to a different location in the body and form a new tumor, is regulated by multiple factors and includes a number of steps and stages. In our previous study, it was demonstrated miR-339-5p inhibits cell migration and invasion in vitro and is associated with the tumor-node-metastasis stage and the lymph node metastasis status of non-small cell lung cancer. In the present study, expression of miR-339-5p was first determined in the tissues and peripheral blood of patients with non-small cell lung cancer (NSCLC) and in NSCLC cell lines. It was then demonstrated that miR-339-5p inhibits A549 and H1299 cell invasion. The underlying molecular events of miR-339-5p action in NSCLC were also explored. By luciferase assay and western blot analysis, B-cell CLL/lymphoma 6 (BCL6) was verified as the direct target gene of miR-339-5p. miR-339-5p may inhibit lung cancer cell invasion and migration by regulating the epithelial-to-mesenchymal transition via BCL6 in vitro. It was also demonstrated that the relative expression of miR-339-5p in the peripheral blood is associated with cancer metastasis in patients with non-small cell lung cancer.

Thyrotrophic status in patients with pituitary stalk interruption syndrome.

Pituitary stalk interruption syndrome (PSIS) is associated with simultaneous or subsequent pituitary hormone deficiencies (PHDs). Although the clinical features of multiple PHDs are well known, the status of the thyrotrophic axis in PSIS has not been thoroughly investigated.The clinical data of 89 PSIS patients and 34 Sheehan syndrome (SS) patients were retrospectively analyzed.The prevalence of central hypothyroidism in the PSIS patients and the SS patients was 79.8% and 70.6%, respectively. The thyroid-stimulating hormone (TSH) levels in the PSIS patients were significantly higher in comparison with the SS patients (5.13 ±â€Š3.40 vs 1.67 ±â€Š1.20 mU/L, P < .05). TSH elevation (8.79 ±â€Š3.17 mU/L) was noticed in 29 of 71 (40.85%) hypothyroid PSIS patients but not in the 24 hypothyroid SS patients. The TSH levels in the hypothyroid PSIS patients were significantly higher in comparison with the euthyroid PSIS patients (5.42 ±â€Š3.67 vs 3.66 ±â€Š1.50 mU/L). Thyroid hormone replacement significantly reduced the TSH levels in the PSIS patients with elevated TSH levels from 7.24 ±â€Š0.98 to 1.67 ±â€Š1.51 mU/L (P < .05). The logistic regression analysis suggested that TSH level was not significantly associated with pituitary stalk status and height of the anterior pituitary gland.PSIS is a newly recognized cause of central hypothyroidism. The proportion and amplitude of TSH elevations are higher in PSIS than in other causes of central hypothyroidism.

Multi-genic pattern found in rare type of hypopituitarism: a whole-exome sequencing study of Han Chinese with pituitary stalk interruption syndrome.

Pituitary stalk interruption syndrome (PSIS) is a rare type of hypopituitarism manifesting various degrees of pituitary hormone deficiency. Although mutations have been identified in some familial cases, the underpinning mechanisms of sporadic patients with PSIS who are in a vast majority remain elusive, necessitating a comprehensive study using systemic approaches. We postulate that other genetic mechanisms may be responsible for the sporadic PSIS. To test this hypothesis, we conducted a study in 24 patients with PSIS of Han Chinese with no family history using whole-exome sequencing (WES) and bioinformatic analysis. We identified a group of heterozygous mutations in 92% (22 of 24) of the patients, and these genes are mostly associated with Notch, Shh, Wnt signalling pathways. Importantly, 83% (20 of 24) of the patients had more than one mutation in those pathways suggesting synergy of compound mutations underpin the pathogenesis of sporadic PSIS.

Clinical Features of Pituitary Stalk Interruption Syndrome in 114 Cases.

Objective To analyze the clinical characteristics of pituitary stalk interruption syndrome(PSIS). Methods The clinical data including clinical manifestations,laboratory tests,and imaging findings of 114 PSIS patients in our hospital were retrospectively analyzed. Results Of these 114 PSIS patients,102 cases (89.4%) were male. The average age was 21.1?6.1 years. A history of breech delivery was documented in 91 cases (91.9%). Short stature was found in 89 cases (71.8%) and bone age delayed (6.1?5.1) years. Secondary sex characteristics were poor or undeveloped in most patients. The prevalence of deficiencies in growth hormone,gonadotropins,corticotropin,and thyrotropin were 100.0%,94.0%,84.2%,and 74.6%,respectively. Hyperprolactinemia was found in 28.1% of patients. Three or more pituitary hormone abnormalities were found in 105 cases(92.1%). Compared with the 5 cases with history of cephalic delivery,no difference were found in the aspects of height(t=0.297,P=0.634),penile length(t=1.205,P=0.882),testicular volume (U=99.000,P=0.348),growth hormone peak (U=89.000,P=0.186),adrenocorticotropic hormone peak(U=131.000,P=0.967),luteinizing hormone peak(U=98.500,P=0.582),thyroid-stimulating hormone (U=82.000,P=0.162),and the height of anterior pituitary (t=1.676,P=0.107) in the 53 cases with history of breech delivery. Conclusions The clinical manifestations,symptoms,hormone deficiencies were severe in our series. The condition severities were not remarkably different in patients with different delivery ways.

Correlation between Pituitary Stalk Interruption Syndrome and Prokineticin Receptor 2 and Prokineticin 2 Mutations.

OBJECTIVE: To analyze the correlation between pituitary stalk interruption syndrome (PSIS) and prokineticin receptor 2 (PROKR2) and prokineticin 2 (RROK2) mutations. METHODS: PROKR2 and RROK2 genotypes were identified by multiplex polymerase chain reaction analysis with exon-flanking primers and by automated sequencing techniques with peripheral blood DNA samples from 59 patients with PSIS. RESULTS: Of these 59 PSIS patients, 6 showed intragenic deletions at the PROKR2 locus. Of them, 5 patients exhibited intragenic subsititution of exon 2 (c.991G>A), and the remaining one patient exhibited intragenic subsititution of exon 2 (c.1057C>T). No PROK2 mutation was found in these PSIS patients. CONCLUSION: PROKR2 may be the susceptibility gene of PSIS.

Preoperative Diagnosis of Juxtaglomerular Cell Tumors in Eight Patients.

The aim of this study was to improve the diagnostic efficiency for juxtaglomerular cell tumors (JCTs) and to determine whether clinical and magnetic resonance imaging features can help to differentiate JCTs from clear cell renal cell carcinoma (ccRCC). The clinical features of eight patients with JCTs and 27 patients with ccRCCs were analyzed. A flow diagram for young people with hypertension was applied to facilitate the diagnosis. Clinical presentations were analyzed, including age, hypertension, and hypokalemia. The results of our study produced a flow diagram that narrowed the scope of diagnosis. The statistical results demonstrated that patients with a renal mass aged 14 to 30 years, had grade 3 hypertension, or had moderate hypokalemia had a greater possibility of having a JCT than a ccRCC (P<.0000, P<.01, P<.0005, respectively). In addition, the flow diagram and magnetic resonance imaging features were useful to distinguish JCTs from other renal tumors.

[Construction of human Bcl-6 3'UTR reporter vector and expression vector and their functional assessment].

OBJECTIVE: To observe the direct regulation of miR-127 on Bcl-6 and the effect of Bcl-6 in rescuing miR-127-induced cell cycle and cell growth inhibition. METHODS: The 3'UTR and coding region of human bcl-6 gene were amplified by PCR and cloned into pcDNA3.0-Luc and pcDNA3.0-Flag vectors, respectively. Mutations were introduced into the seed sequences of the predicted miR-127 target sites within the Bcl-6 3'UTR using recombinant PCR. Luciferase assay was used to verify the direct targeted regulation of miR-127 on Bcl-6. In HepG2 cell models with overexpression or knockdown of miR-12, the changes of cell cycle and cell growth were investigated after transfection with the constructed vectors. RESULTS: The recombinant plasmids were successfully obtained as confirmed by double digestion and sequence identification. Luciferase assay showed that in 293T and HepG2 cells, miR-127 inhibited the activation of wild-type Bcl-6 3'UTR reporter vector but not mutated Bcl-6 3'UTR vector. Overexpression of miR-127 induced cell cycle arrest at G(2)/M phase and suppressed the growth of HepG2 cells, and these effects were reversed by Bcl-6 overexpression. CONCLUSION: We successfully cloned wild-type and mutated 3'UTR reporter vectors and expression vector of bcl-6 gene and confirmed their biological functions.

MiR-125a suppresses tumor growth, invasion and metastasis in cervical cancer by targeting STAT3.

MiR-125a has been characterized as a tumor suppressor in several cancers. However, the role of miR-125a in cervical cancer is unknown. In this study, we found the expression of miR-125a was downregulated in cervical cancer patients, and negatively correlated with the tumor size, FIGO stage, and preoperative metastasis. Kaplan-Meier analysis showed that miR-125a expression predicted favorable outcome for cervical cancer patients. Dual luciferase assays identified the STAT3 gene as a novel direct target of miR-125a. Functional studies showed that miR-125a overexpression significantly suppressed the growth, invasion and epithelial-mesenchymal transition (EMT) of cervical cancer cells both in vitro and in vivo via decreasing STAT3 expression. Moreover, miR-125a conferred to G2/M cell cycle arrest, accompanied by inhibition of several G2/M checkpoint proteins. Mechanistically, inactivation of miR-125a during cervical carcinogenesis was caused by HPV suppression of p53 expression. Clinically, STAT3, the expression of which, predicted poorer outcome, was inversely correlated with miR-125a in cervical cancer. These data highlight the importance of miR-125a in the cell proliferation and progression of cervical cancer, and indicate that miR-125a may be a useful therapeutic target for cervical cancer.

[FHL1 knockdown mediated by lentiviral shRNA promotes the growth of HeLa and HepG2 cells].

OBJECTIVE: To investigate the effect of four-and-a-half LIM domain 1 (FHL1) knockdown by lentiviral-mediated shRNA on the growth of HeLa and HepG2 cells. METHODS: pLenti-H1 FHL1 shRNA was cloned, and then transfered into HEK293T cells. The inhibitory effect of pLenti-H1 FHL1 shRNA on FHL1 gene was detected by Western blotting and real-time quantitative PCR (qRT-PCR). Lentivirus particles were packaged, added to HeLa and HepG2 cells, followed by puromycin treatment for 2-3 weeks to screen stable clones. The knockdown effect on FHL1 expression in these cells was checked by Western blotting and qRT-PCR. Cell growth and colony formation analysis were performed to investigate the effect of FHL1 down-regulation on tumor cell growth. Soft agar analysis was used to analyze its effect on anchorage-independent tumor cell growth. RESULTS: Western blotting and qRT-PCR revealed that the pLenti-H1 FHL1 shRNA apparently inhibited the expression of FHL1 gene. Cell growth and colony formation assay showed that the lentiviral-mediated shRNA for FHL1 gene significantly accelerated the tumor cell growth in HepG2 and HeLa cells. Soft agar analysis demonstrated that FHL1 shRNA increased the anchorage-independent growth of tumor cells. CONCLUSION: pLenti-H1 FHL1 shRNA could significantly accelerate tumor cell growth via inhibiting the expression of FHL1.

RSRC1 SUMOylation enhances SUMOylation and inhibits transcriptional activity of estrogen receptor ß.

The transcription factor estrogen receptor ß (ERß) plays roles in the central nervous, endocrine, cardiovascular, and immune systems. ERß can be SUMOylated. However, the underlying mechanism remains unclear. Here, we show that RSRC1/SRrp53 interacts with ERß and SUMOylation of RSRC1 is required for regulation of PIAS1-mediated ERß SUMOylation. RSRC1 promotes ERß SUMOylation through enhanced interaction between ERß and PIAS1. RSRC1 represses ERß transcriptional activity through regulation of ERß SUMOylation. By establishing RSRC1 as a novel cofactor for SUMOylation, our data provide insight into regulation of ERß SUMOylation and indicate that SUMOylation of one protein can regulate another protein SUMOylation.

Metformin inhibits thyroid cancer cell growth, migration, and EMT through the mTOR pathway.

Mammalian target of rapamycin (mTOR) signaling pathways have been shown to be activated in thyroid cancer. Recent evidences have demonstrated that the antidiabetic agent metformin, an activator of 5'-AMP-activated protein kinase, can impair the proliferation and migration of cancer cells via inhibition of mTOR. However, the underlying mechanisms remain unclear. In this study, we show that metformin can inhibit mTOR pathway to impair growth and migration of the thyroid cancer cell lines. Cyclin D1 and c-Myc are important regulators of cancer cell growth, and we observed that treatment of thyroid cancer cells with metformin reduced c-Myc and cyclin D1 expression through suppression of mTOR and subsequent inhibition of P70S6K1 and 4E-BP1 phosphorylation. Metformin reduced epithelial to mesenchymal transition (EMT) in thyroid carcinoma cells. Moreover, metformin regulated expression of the EMT-related markers E-cadherin, N-cadherin, and Snail. Additionally, knockdown of TSC2, the upstream regulatory molecule of mTOR pathway, or treatment of rapamycin, the mTOR inhibitor, could abolish the effects of metformin to regulate thyroid cancer cell proliferation, migration, EMT, and mTOR pathway molecules. These results indicate that metformin can suppress the proliferation, migration, and EMT of thyroid cancer cell lines by inhibiting mTOR signaling. These findings suggest that metformin and its molecular targets may be useful in thyroid carcinoma therapy.

[Evaluation of the 1 mg overnight dexamethasone suppression test in the diagnosis of the subclinical Cushing's syndrome in patients with Adrenal Incidentaloma].

OBJECTIVE: To evaluate efficacy of the 1 mg overnight dexamethasone suppression test (1 mg overnight DST) in the diagnosis of subclinical Cushing's syndrome, and to explore the best diagnostic cut-off value. METHODS: The clinical data of patients with adrenal incidentaloma in Chinese PLA General Hospital from 1995 to 2013 were gathered. The data of subclinical Cushing's syndrome (SCS) and non-functional adrenal adenoma (NFA) was retrospectively analyzed. The ROC curve was used to evaluate the efficacy of the 1 mg overnight DST and to explore the best cut-off value with high sensitivity and specificity. RESULTS: There were 447 patients with NFA (224 male and 223 female), and the mean age was 53±11 years old.49 patients were with SCS (19 male and 30 female), and the mean age was 47±12 years old. The area under the ROC of serum cortisol level after 1 mg overnight DST was 0.967 (95%CI: 0.942-0.993). The best cut-off value of serum cortisol after 1 mg overnight DST was 63.65 nmol/L, with a sensitivity of 100.0% and a specificity of 88.8%. The best cut-off value of the suppression ratio of serum cortisol was 85.64%, with a sensitivity of 83.3% and a specificity of 84.6%. CONCLUSIONS: The best criterion for 1 mg overnight DST in the diagnosis of SCS was serum cortisol level , and the recommend cut-off point was 63.65 nmol/L, with both a higher sensitivity and specificity. The suppression ratio of serum cortisol after 1 mg overnight DST was also considered as a suitable criterion in the diagnosis of SCS.

[SUMO-2/3 can covalently bind to progesterone receptor B to regulate its transcriptional activity].

OBJECTIVE: To investigate whether progesterone receptor B (PRB) can be sumoylated by SUMO-2/3 and the effect of sumoylation on PRB transcriptional activity. METHODS: SUMO-2/3 cDNA was amplified from MCF-7 cDNA and cloned into the eukaryotic expression vector pcDNA3-FLAG. The plasmid pXJ40-myc-PRB was cotransfected with pcDNA3FLAG-SUMO2, pcDNA3FLAG-SUMO3 or the mock control into 293T cells, and PRB sumoylation was detected by immunoprecipitation and Western blotting. The effect of PRB sumoylation on its transcriptional activity was determined using reporter luciferase assay. RESULTS: pcDNA3FLAG-SUMO2 and pcDNA3FLAG-SUMO3 vectors were successfully constructed. SUMO-2/3 could bind covalently to PRB and increase its transcriptional dependent on the presence of progesterone. CONCLUSION: PRB can be sumoylated by SUMO-2/3 and its function is regulated by this modification.