Expressions of eukaryotic initiation factor-4B and eukaryotic initiation factor-5A in papillary thyroid carcinoma and their effects on cell proliferation in vitro / 肿瘤研究与临床

Objective:To investigate the expressions of eukaryotic initiation factor-4B (eIF4B) and eukaryotic initiation factor-5A (eIF5A) in papillary thyroid carcinoma tissues, and to analyze their regulatory effects on cell proliferation in vitro.Methods:The clinical data of 61 patients diagnosed with papillary thyroid carcinoma who received surgical resection at Yuncheng Central Hospital from January 2020 to October 2021 were retrospectively analyzed. The postoperative tumor tissues and paracancerous normal thyroid tissues (>1 cm from the margin of the mass) were retained. Immunohistochemistry was used to detect the expressions of eIF4B, eIF5A and proliferating cell nuclear antigen (PCNA) in different tissues. The correlation of eIF4B, eIF5A expressions with the clinicopathological characteristics of patients, and the relationship between eIF4B, eIF5A and PCNA were analyzed. The thyroid cancer cell line SW1736 and normal thyroid cell line HT-ori3 were selected. The expressions of eIF4B mRNA and eIF5A mRNA were detected by using real-ime quantitative polymerase chain reaction (qRT-PCR). After the small interfering RNA (siRNA) of siRNA-eIF4B and siRNA-eIF5A were synthesized, the interfering plasmids were constructed, and SW1736 cells were transfected, siRNA-eIF4B group and siRNA-eIF5A group were obtained; the empty plasmid transfection group and the blank control group without transfection intervention were established. The cell proliferation activity was detected by using CCK-8 assay, and the expression of PCNA mRNA was detected by using qRT-PCR.Results:The positive rates of eIF4B and eIF5A in papillary thyroid cancer tissues were higher than those in paracancerous normal thyroid tissues [65.57% (40/61) vs. 29.51% (18/61), 57.38% (35/61) vs. 9.84% (6/61), P < 0.001]. The positive rates of eIF4B and eIF5A were statistically different in patients with different tumor diameter [>3 cm vs. ≤3 cm: 88.89% (16/18) vs. 55.81% (24/43),77.78% (14/18) vs. 48.84% (21/43), all P < 0.05], lymph node metastasis [with vs. without: 85.00% (17/20) vs. 56.10% (23/41), 80.00% (16/20) vs. 46.34% (19/41), all P < 0.05] and the number of different nodes [multiple vs. single: 86.67% (13/15) vs. 58.70% (27/46), 86.67% (13/15) vs. 47.83% (22/46), all P < 0.05]; there were no statistically significant differences in the positive rates of eIF4B and eIF5A among patients with different age and gender (all P > 0.05). Positive correlation was found between eIF4B score and the positive cell proportion of PCNA ( r = 0.66, P = 0.0324), eIF5A score and the positive cell proportion of PCNA ( r = 0.62, P = 0.024), eIF4B score and eIF5A score ( r = 0.63, P = 0.021). The expression levels of eIF4B mRNA and eIF5A mRNA in thyroid cancer cell line SW1736 cell was higher than that of HT-ori3 cell in normal thyroid (all P < 0.05). The cell proliferation activity of SW1736 and PCNA mRNA expression level in siRNA-eIF4B group and siRNA-eIF5A group were lower than those in the empty vector transfected group and the blank control group (all P < 0.05). Conclusions:eIF4B and eIF5A are expressed elevated in papillary thyroid carcinoma, and both are involved in tumor development and progression. The role of eIF4B and eIF5A may be related to promoting the proliferation of tumor cells.

Overexpression of eukaryotic initiation factor 5A (eIF5A) affects susceptibility to benznidazole in Trypanosoma cruzi populations

Eukaryotic initiation factor 5A (eIF5A) is a conserved protein with an essential role in translation elongation. Using one and two-dimensional western blotting, we showed that the eIF5A protein level was 2-fold lower in benznidazole (BZ)-resistant (BZR and 17LER) Trypanosoma cruzi populations than in their respective susceptible counterparts (BZS and 17WTS). To confirm the role of eIF5A in BZ resistance, we transfected BZS and 17WTS with the wild-type eIF5A or mutant eIF5A-S2A (in which serine 2 was replaced by alanine). Upon overexpressing eIF5A, both susceptible lines became approximately 3- and 5-fold more sensitive to BZ. In contrast, the eIF5A-S2A mutant did not alter its susceptibility to BZ. These data suggest that BZ resistance might arise from either decreasing the translation of proteins that require eIF5A, or as a consequence of differential levels of precursors for the hypusination reactions (e.g., spermidine and trypanothione), both of which alter BZ's effects in the parasite.

Effect of DHPS on Metastasis of Gastric Cancer and its Possible Mechanism / 胃肠病学

Background:Deoxyhypusine synthase(DHPS)is a key factor in post-translational modification of the precursor of eukaryotic initiation factor 5A(eIF-5A),and eIF-5A is closely related to the regulation of proliferation and invasion of tumor cells. Aims:To investigate the expression of DHPS in gastric cancer and its clinical significance,and to explore the possible mechanism of its effect on metastasis of gastric cancer. Methods:Tissue microarray containing 92 gastric cancer tissues and paired adjacent cancerous tissues was employed to detect the DHPS expression by using immunohistochemical staining,and the correlation of DHPS expression with the clinicopathological characteristics of gastric cancer was analyzed. DHPS-siRNA and GC7,an inhibitor of DHPS were used,respectively to intervene human gastric cancer cell line MGC803. The invasive ability of MGC803 cells was assessed by cell invasion assay,and the expressions of metastasis-related proteins including vascular endothelial growth factor(VEGF),matrix metalloproteinase 2(MMP2)and MMP9 were detected by ELISA. Results:In 62(67.4%)cases of gastric cancer,DHPS was highly expressed,and its expression was closely related to tumor diameter,TNM stage and depth of invasion(P <0.05),but not related to gender,age,lymph node metastasis and distant metastasis(P >0. 05). Both DHPS-siRNA and GC7 could down-regulate the invasiveness of MGC803 cells,while the former could also reduce the expressions of VEGF,MMP2 and MMP9 proteins(P <0.05). Conclusions:DHPS is highly expressed in gastric cancer and associated with tumor invasion and progression. DHPS is expected to be a new target for diagnosis and treatment of gastric cancer because of its regulatory effect on invasion and metastasis of tumor cells.

Protein kinase CK2 phosphorylates and interacts with deoxyhypusine synthase in HeLa cells

Deoxyhypusine is a modified lysine and formed posttranslationally to be the eukaryotic initiation factor eIF5A by deoxyhypusine synthase, employing spermidine as butylamine donor. Subsequent hydroxylation of this deoxyhypusine-containing intermediate completes the maturation of eIF5A. The previous report showed that deoxyhypusine synthase was phosphorylated by PKC in vivo and the association of deoxyhypusine synthase with PKC in CHO cells was PMA-, and Ca(2+)/phospholipid-dependent. We have extended study on the phosphorylation of deoxyhypusine synthase by protein kinase CK2 in order to define its role on the regulation of eIF5A in the cell. The results showed that deoxyhypusine synthase was phosphorylated by CK2 in vivo as well as in vitro. Endogenous CK2 in HeLa cells and the cell lysate was able to phosphorylate deoxyhypusine synthase and this modification is enhanced or decreased by the addition of CK2 effectors such as polylysine, heparin, and poly(Glu, Tyr) 4:1. Phosphoamino acid analysis of this enzyme revealed that deoxyhypusine synthase is mainly phosphorylated on threonine residue and less intensely on serine. These results suggest that phosphorylation of deoxyhypusine synthase is CK2-dependent cellular event as well as PKC-mediated effect. However, there were no observable changes in enzyme activity between the phosphorylated and unphosphorylated forms of deoxyhypusine synthase. Taken together, besides its established function in hypusine modification involving eIF5A substrate, deoxyhypusine synthase and its phosphorylation modification may have other independent cellular functions because of versatile roles of deoxyhypusine synthase.