• The Korean Journal of Physiology and Pharmacology
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• 제23권3호
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• pp.171-179
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• 2019

Pituitary tumors are usually benign but can occasionally exhibit hormonal and proliferative behaviors. Dysregulation of the G1/S restriction point largely contributes to the over-proliferation of pituitary tumor cells. F-box protein S-phase kinase-interacting protein-2 (SKP2) reportedly targets and inhibits the expression of $p27^{Kip1}$, a well-known negative regulator of G1 cell cycle progression. In this study, SKP2 expression was found to be upregulated while $p27^{Kip1}$ expression was determined to be downregulated in rat and human pituitary tumor cells. Furthermore, SKP2 knockdown induced upregulation of $p27^{Kip1}$ and cell growth inhibition in rat and human pituitary tumor cells, while SKP2overexpression elicited opposite effects on $p27^{Kip1}$ expression and cell growth. The expression of microRNA-186 (miR-186) was reported to be reduced in pituitary tumors. Online tools predicted SKP2 to be a direct downstream target of miR-186, which was further confirmed by luciferase reporter gene assays. Moreover, miR-186 could modulate the cell proliferation and $p27^{Kip1}$-mediated cell cycle alternation of rat and human pituitary tumor cells through SKP2. As further confirmation of these findings, miR-186 and $p27^{Kip1}$ expression were downregulated, while SKP2 expression was upregulated in human pituitary tumor tissue samples; thus, SKP2 expression negatively correlated with miR-186 and $p27^{Kip1}$ expression. In contrast, miR-186 expression positively associated with $p27^{Kip1}$ expression. Taken together, we discovered a novel mechanism by which miR-186/SKP2 axis modulates pituitary tumor cell proliferation through $p27^{Kip1}$-mediated cell cycle alternation.

• 한국콘텐츠학회논문지
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• 제19권11호
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• pp.375-382
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• 2019

임신 초기 염증으로 인한 영양막세포의 기능 이상은 C-reactive protein (CRP)의 발현을 증가시켜 산모와 태아의 상호작용에 영향을 미침으로써, 조산 및 자간전증 등을 유발한다. 그러나, CRP 발현 조절과 관련된 생체표지자 발굴 및 개발은 미흡한 실정이다. 본 연구는 염증이 유발된 영양막세포에서 증가된 CRP 발현과 관련된 miRNA를 발굴 및 그 발현을 분석함으로써, miRNA를 통해 영양막세포 염증 조절 기전에 관여하는 생체표지자를 밝히고자 한다. miRNA 데이터베이스(mirna, TargetScan, MicroCosm)에서 공통적으로 CRP 유전자 발현을 조절할 것으로 예측되는 miR-7, miR-150, miR-186, 그리고 miR-424를 선별하여 HTR-8/SVneo에 LPS (20ng/mL)를 처리하여 in vitro 상에서 염증 반응을 유도하였다. 각각의 miRNAs의 발현을 qRT-PCR 방법으로 비교 분석하였다. 그 결과, LPS 처리된 영양막세포에서 CRP의 발현은 유의성 있게 증가되었다(p<0.001). miR-150와 miR-424는 발현이 유의성 있게 감소됨을 확인하였다(p<0.001). 따라서, 염증이 유도된 영양막세포에서의 CRP 발현을 조절하는 기전에 miR-150와 miR-424가 관여하는 것을 의미하며, 향후 염증성 산과질환의 산전 진단에 유용한 자료로 사용될 것으로 사료된다.

• Molecules and Cells
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• 제37권8호
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• pp.620-627
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• 2014

We have previously shown that microRNAs (miRNAs) miR-760, miR-186, miR-337-3p, and miR-216b stimulate premature senescence through protein kinase CK2 (CK2) downregulation in human colon cancer cells. Here, we examined whether these four miRNAs are involved in the replicative senescence of human lung fibroblast IMR-90 cells. miR-760 and miR-186 were significantly upregulated in replicatively senescent IMR-90 cells, and their joint action with both miR-337-3p and miR-216b was necessary for efficient downregulation of the ${\alpha}$ subunit of CK2 ($CK2{\alpha}$) in IMR-90 cells. A mutation in any of the four miRNA-binding sequences within the $CK2{\alpha}3^{\prime}$-untranslated region (UTR) indicated that all four miRNAs should simultaneously bind to the target sites for $CK2{\alpha}$ downregulation. The four miRNAs increased senescence-associated ${\beta}$-galactosidase (SA-${\beta}$-gal) staining, p53 and $p21^{Cip1/WAF1}$ expression, and reactive oxygen species (ROS) production in proliferating IMR-90 cells. $CK2{\alpha}$ overexpression almost abolished this event. Taken together, the present results suggest that the upregulation of miR-760 and miR-186 is associated with replicative senescence in human lung fibroblast cells, and their cooperative action with miR-337-3p and miR-216b may induce replicative senescence through $CK2{\alpha}$ downregulation-dependent ROS generation.

• Asian Pacific Journal of Cancer Prevention
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• 제14권1호
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• pp.139-143
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• 2013

Patients at the same pathological stage of esophageal cancer (EC) that received the same surgical therapy by the same surgeon may have distinct prognoses. The current study aimed to explore the possibility of differentially-expressed microRNAs (miRNAs) underlying this phenomenon. Samples were collected from EC patients at the same tumor node metastasis (TNM) stage but with different prognoses. Paracancerous normal tissues were taken as controls. The specimens were histopathologically analyzed. Differentially-expressed miRNAs were analyzed using real-time quantitative reverse transcription polymerase chain reaction. Compared with patients with poor prognosis, those with good prognosis exhibited 88 two-fold or more than two-fold increased miRNA fragments and 4 half-decreased miRNAs. The most noticeably up-regulated miRNAs included hsa-miR-31, hsa-miR-196b, hsa-miR-652, hsa-miR-125a-5p, hsa-miR-146b, hsa-miR-200c, hsa-miR-23b, hsa-miR-29a, hsa-miR-186, hsa-miR-205, hsa-miR-376a, hsa-miR-410, hsa-miR-532-3p, and hsa-miR-598, whereas the most significantly-downregulated miRNAs were hsa-let-7e, hsa-miR-130b, and hsa-miR-103. EC patients at same TNM stage but with different prognoses show differentially-expressed miRNAs.

• 한국환경농학회지
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• 제40권3호
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• pp.186-193
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• 2021

BACKGROUND: Before generating transgenic plant using the CRISPR-Cas9 system, the efficiency test of sgRNAs is recommended to reduce the time and effort for plant transformation and regeneration process. The efficiency of the sgRNA can be measured through the transient expression of sgRNA and Cas9 gene in tomato cotyledon; however, we found that the calculated efficiency showed a large variation. It is necessary to increase the precision of the experiment to obtain reliable sgRNA efficiency data from transient expression. METHODS AND RESULTS: The cotyledon of 11^th, 15^th, 19^th, and 23^rd-day-old tomato (Solanum lycopersicum cv. Micro-Tom) were used for expressing CRISPR-Cas9 transiently. The agrobacterium harboring sgRNA for targeting ALS2 gene of tomato was injected through the stomata of leaf adaxial side and the genomic DNA was extracted in 5 days after injection. The target gene edition was identified by amplifying DNA fragment of target region and analyzing with Illumina sequencing method. The target gene editing efficiency was calculated by counting base deletion and insertion events from total target sequence read. CONCLUSION: The CRISPR-Cas9 editing efficiency varied with tomato cotyledon age. The highest efficiency was observed at the 19-day-old cotyledons. Both the median and mean were the highest at this stage and the sample variability was also minimized. We found that the transgene of CRISPR-Cas9 system was strongly correlated with plant leaf development and suggested the optimum cotyledon leaf age for Agrobacterium-mediated transfection in tomato.

• Journal of Oral Medicine and Pain
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• 제44권1호
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• pp.25-30
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• 2019

Purpose: The exact causes of burning mouth syndrome (BMS) is unclear so far. There are many studies to elucidate the relation between oral disease and genetic predisposition. In this study, we first tried to investigate salivary exosomal genetic components that could play an important role for diagnosing and elucidating the progression of BMS. Methods: We compared salivary exosomal micro RNAs (miRNAs) of BMS Patients to those of control using next generation sequencing (NGS). Unstimulated whole saliva from 15 patients with BMS and 10 control subjects were divided into two sets. Isolated exosomes and their total RNAs were subject to NGS for the screening of miRNAs. Results: There were up-regulated 10 exosomal miRNAs (hsa-miR-1273h-5p, hsa-miR-1273a, hsa-miR-1304-3p, hsa-miR-4449, hsa-miR-1285-3p, hsa-miR-6802-5p, hsa-miR-1268a, hsa-miR-1273d, hsa-miR-1273f, and hsa-miR-423-5p) and down-regulated 18 exosomal miRNAs (hsa-miR-27b-3p, hsa-miR-16-5p, hsa-miR-186-5p, hsa-miR-142-3p, hsa-miR-141-3p, hsa-miR-150-5p, hsa-miR-374a-5p, hsa-miR-93-5p, hsa-miR-29c-3p, hsa-miR-29a-3p, hsa-miR-148a-3p, hsa-miR-22-3p, hsa-miR-27a-3p, hsa-miR-424-5p, hsa-miR-19b-3p, hsa-miR-99a-5p, hsa-miR-548d-3p, and hsa-miR-19a-3p) in BMS patients comparing with those of control subjects. Conclusions: We show that there are 28 differential expression of miRNAs between the patients with BMS and those of control subjects. The specific function of indicated miRNAs should be further elucidated.