• Asian-Australasian Journal of Animal Sciences
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• 제33권1호
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• pp.4-11
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• 2020

Objective: Puerarin has the potential of regulating the differentiation of preadipocytes, but its mechanism of action has not yet been elucidated. Adipocytes found in adipose tissue, the main endocrine organ, are the main sites of lipid deposition, and are widely used as a cell model in the study of in vitro fat deposition. This study aimed to investigate the effects of puerarin on adipogenesis in vitro. Methods: Puerarin was added to the culture medium during the process of adipogenesis. The proliferation and differentiation of bovine preadipocytes was measured through cell viability and staining with oil red O. The content of triacylglycerol was measured using a triglyceride assay kit. The mRNA and protein expression levels of adipogenic genes, peroxisome proliferator-activated receptor-γ (PPARγ) and CCAAT/enhancer-binding protein-α, were measured using quantitative real-time polymerase chain reaction and western blotting, respectively. Results: The addition of puerarin significantly increased adipogenesis of bovine preadipocytes and enhanced the mRNA and protein level expression of PPARγ (p<0.01). The expression of P-Akt increased after adipogenic hormonal induction, whereas puerarin significantly increased PPARγ expression by promoting the Akt signaling component, P-Akt. The mechanism of adipogenesis was found to be related to the phosphorylation level of Ser473, which may activate the downstream signaling of the Akt pathway. Conclusion: Puerarin was able to promote the differentiation of preadipocytes and improve fat deposition in cattle. The mechanism of adipogenesis was found to be related to the phosphorylation level of Ser473.

• The Korean Journal of Physiology and Pharmacology
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• 제23권2호
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• pp.103-111
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• 2019

The study is to investigate effects of andrographolide on experimental autoimmune myocarditis (EAM). Lewis rats were immunized on day 0 with porcine cardiac myosin to establish EAM. The EAM rats were treated with either andrographolide (25, 50, 100 mg/kg/day) or vehicle for 21 days. An antigen-specific splenocytes proliferation assay was performed by using the cells from control rats immunized with cardiac myosin. Survival rates, myocardial pathology and myocardial functional parameters (left ventricle end-diastolic pressure, ${\pm}dP/dt$ and left ventricular internal dimension) of EAM rats received andrographolide were significantly improved. Andrographolide treatment caused an decrease in the infiltration of $CD3^+$ and $CD14^+$ positive cells in myocardial tissue. Moreover, andrographolide treatment caused a reduction in the plasma levels of tumor necrosis factor-alpha, interleukin-17 (IL-17) and myosin-antibody, and an increase in the level of IL-10 in EAM rats. Oral administration of andrographolide resulted in the decreased expression of p-PI3K, p-Akt without any change of PI3K and Akt. Further results indicate andrographolide significantly inhibited myosin-induced proliferation in splenocytes, and this effect was inhibited by co-treatment of SC79 (Akt activator). Our data indicate andrographolide inhibits development of EAM, and this beneficial effect may be due to powerful anti-inflammatory activity and inhibitory effect on PI3K/Akt pathway.

• The Korean Journal of Physiology and Pharmacology
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• 제27권5호
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• pp.493-511
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• 2023

Hippo/YAP signaling hinders cancer progression. Inactivation of this pathway contributes to the development of esophageal cancer by activation of Akt. However, the possible interaction between Akt and Hippo/YAP pathways in esophageal cancer progression is unclear. In this study, we found that ursolic acid (UA) plus 3'3-diindolylmethane (DIM) efficiently suppressed the oncogenic Akt/Gsk-3β signaling pathway while activating the Hippo tumor suppressor pathway in esophageal cancer cells. Moreover, the addition of the Akt inhibitor LY294002 and the PI3K inhibitor 3-methyladenine enhanced the inhibitory effects of UA plus DIM on Akt pathway activation and further stimulated the Hippo pathway, including the suppression of YAP nuclear translocation in esophageal cancer cells. Silencing YAP under UA plus DIM conditions significantly increased the activation of the tumor suppressor PTEN in esophageal cancer cells, while decreasing p-Akt activation, indicating that the Akt signaling pathway could be down-regulated in esophageal cancer cells by targeting PTEN. Furthermore, in a xenograft nude mice model, UA plus DIM treatment effectively diminished esophageal tumors by inactivating the Akt pathway and stimulating the Hippo signaling pathway. Thus, our study highlights a feedback loop between the PI3K/Akt and Hippo signaling pathways in esophageal cancer cells, implying that a low dose of UA plus DIM could serve as a promising chemotherapeutic combination strategy in the treatment of esophageal cancer.

• Asian Pacific Journal of Cancer Prevention
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• 제17권2호
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• pp.743-748
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• 2016

Background: $K^-Ras$ activation is an early event in colorectal carcinogenesis and associated mutations have been reported in about 40% of colorectal cancer patients. These mutations have always been responsible for enhancing malignancy and silencing them is associated with attenuation of tumorigenicity. Among downstream effectors are the RAF/MEK/ERK and the PI3K/Akt signaling pathways. PI3K/Akt signaling leads to reduction of apoptosis, stimulated cell growth and enhanced proliferation. Ellagic acid (EA), a naturally occurring antioxidant, has recently emerged as a promising anti-cancer agent. Purpose: To evaluate the impact of cellular genetic makeup of two colon cancer cell lines with different genetic backgrounds, HCT-116 ($K^-Ras^-/p53^+$) and Caco-2 ($K^-Ras^+/p53^-$), on response to potential anti-tumour effects of EA. In addition, the influence of $K^-Ras$ silencing in HCT-116 cells was investigated. Materials and Methods: Cellular proliferation, morphology and cell cycle analysis were carried out in addition to Western blotting for detecting total Akt and p-Akt (at Thr308 and Ser473) in the presence and absence of different concentrations of EA. Cell proliferation was also assessed in cells transfected with different concentrations of $K^-Ras$ siRNA or incubated with ellagic acid following transfection. Results: The results of the present study revealed that EA exerts anti-proliferative and dose-dependent pro-apoptotic effects. Cytostatic and cytotoxic effects were also observed. p-Akt (at Thr308 and Ser473) was downregulated. Moreover, EA treatment was found to (i) reduce $K^-Ras$ protein expression; (ii) in cells transfected with siRNA and co-treated with EA, pronounced anti-proliferative effects as well as depletion of p-Akt (at Thr308) were detected. Conclusions: Cellular genetic makeup ($K^-Ras^-/p53^-$) was not likely to impose limitations on targeting EA in treatment of colon cancer. EA had a multi-disciplinary pro-apoptotic anti-proliferative approach, having inhibited Akt phosphorylation, induced cell cycle arrest and showed an anti-proliferative potential in HCT-116 cells (expressing mutant $K^-Ras$).

• 대한임상검사과학회지
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• 제52권1호
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• pp.62-68
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• 2020

Cisplatin (CDDP)은 난소암 치료에 사용되는 화학 요법제로 암세포에 따라 그리고 치료 용량에 따라 다중 신호경로를 활성화하여 세포 반응을 다르게 일으킬 수 있다. Cisplatin이 세포에 작용하는 신호전달 기전은 분명하지 않아 더 많은 연구가 필요해 보인다. 이에 본 연구는 cisplatin을 난소암 세포(SKOV3)에 처리하여 세포사멸 유도 과정에서 나타나는 신호 단백질의 역할을 밝히고자 하였다. 결과는 cisplatin으로 처리한 난소암 세포에서 TUNEL assay와 유세포 분석을 통해 대조군과 비교하여 세포 사멸수가 증가하였다. 세포 사멸 단계에서 나타나는 PARP 및 caspase-3 활성도 증가하였다. 그러나 Bcl-2의 발현은 감소하였다. 신호 전달 경로에서 나타나는 단백질의 발현은 ERK1/2의 활성은 시간 의존적으로 감소하였으나 Akt 활성은 24시간에 감소하다 48시간에서의 활성은 일정하였다. p38과 p-JUN의 활성은 24시간에 증가하는 것으로 나타났으나 48시간에서 p38의 활성은 감소하였으며 p-JUN의 활성은 일정하였다. 이상의 결과들을 토대로 결론은 cisplatin이 SKOV3 세포에서 Akt 활성을 감소하여 세포 증식을 억제하고 MAPK의 p38 발현을 조절하여 세포사멸을 유도하는 것으로 판단된다. 향후, 암치료 전략에 도움이 되는 cisplatin을 포함한 백금기반 화학요법제의 신호전달 기전을 밝히기 위한 더 많은 연구가 필요할 것으로 생각된다. 본 실험을 통해 제시한 결과는 MAPK 신호 경로를 타겟으로 하는 암 치료 전략에 유용하게 사용 될 수 있기를 기대한다.

• 생명과학회지
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• 제26권6호
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• pp.663-672
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• 2016

벌 사상자[Cnidium monnieri (L.) Cusson]는 중국과 한국에 분포하는 일년생 초본으로, 화농성피부염 및 여성의 생식기 질환의 치료에 널리 사용되고 있다. 이 외에도 면역기능개선과 천식 등에 대한 효과는 보고된 바 있으나 아직까지 암과 관련된 연구는 많이 이루어지지 않았다. 이에 따라 본 연구에서는 인간 대장암 세포인 HCT116 세포주에서 벌 사상자 에탄올 추출물(CME)의 apoptosis 및 세포주기정지 유도 효과에 대하여 알아보고자 하였으며, 이러한 효과가 AKT/mTOR/GSK-3β 신호경로의 조절을 통하여 이루어지는지 확인하고자 하였다. MTT assay와 LDH assay 결과, 벌 사상자 에탄올 추출물에 의하여 HCT116 세포의 세포생존율이 감소하였으며, 세포독성효과가 나타났다. 또한 벌 사상자 에탄올 추출물의 농도의존적으로 apoptotic body의 수와 apoptosis 비율이 증가하였으며, G1기에서 세포주기정지 유도 효과가 관찰되었다. 세포의 성장과 증식 및 분열에 관련된 단백질인 Akt는 mTOR, p53, GSK-3β와 같은 신호단백질들의 발현을 조절하는 것으로 보고되었다. 벌 사상자 에탄올 추출물을 처리하였을 때, Akt와 mTOR 단백질의 인산화가 저해되었으며, 이에 따라 하위 신호조절 단백질인 GSK-3β, Bcl-2 family, Caspase-3, PARP의 발현이 조절되었다. 또한 Akt와 GSK-3β, mTOR 저해제 처리를 통하여 CME에 의한 apoptosis 효과가 AKT/mTOR/GSK-3β 신호경로를 통하여 이루어지는 것을 확인하였다. 결론적으로, 본 연구를 통하여HCT116 대장암 세포주에서 벌 사상자 에탄올 추출물이 암세포의 apoptosis 및 세포주기정지 유도에 효과적임을 확인하였다.

• KSBB Journal
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• 제30권5호
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• pp.223-229
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• 2015

In this study, extract from Artemisia annua in L. (AAE) is known as a medicinal herb that is effective against cancer. The cell cycle is regulated by the activation of cyclin-dependent kinase (CDK)/cyclin complex. We will focus on regulation of CDK2 by cyclin E. cyclin E is associated with CDK2 to regulate progression from G1 into S phase. Akt is known to play an important role in cell proliferation and cell survival. Activation of Akt increases mTOR activity that promotes cell proliferation and cancer growth. In this study, we investigated that AAE-induced cell cycle arrest at G1/S phase in HCT116 colon cancer. Treatment of AAE shows that reduced activation of Akt decreases mTOR/Mdm2 activity and then leads to increase the activation of p53. The active p53 promotes activation of p21. p21 induces inactivation of CDK2/cyclin E complex and occurs cell cycle arrest at G1/S phase. We treated LY294002 (Akt inhibitor) and Rapamycin (mTOR inhibitor) to know the relationship between the signal transduction of proteins associated with cell cycle arrest. These results suggest that AAE induces cell cycle arrest at G1/S phase by Akt/mTOR pathway in HCT116 colon cancer cell.

• Biomolecules & Therapeutics
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• 제31권6호
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• pp.661-673
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• 2023

Treatment of colorectal cancer (CRC) has always been challenged by the development of resistance. We investigated the antiproliferative activity of licochalcone H (LCH), a regioisomer of licochalcone C derived from the root of Glycyrrhiza inflata, in oxaliplatin (Ox)-sensitive and -resistant CRC cells. LCH significantly inhibited cell viability and colony growth in both Ox-sensitive and Ox-resistant CRC cells. We found that LCH decreased epidermal growth factor receptor (EGFR) and AKT kinase activities and related activating signaling proteins including pEGFR and pAKT. A computational docking model indicated that LCH may interact with EGFR, AKT1, and AKT2 at the ATP-binding sites. LCH induced ROS generation and increased the expression of the ER stress markers. LCH treatment of CRC cells induced depolarization of MMP. Multi-caspase activity was induced by LCH treatment and confirmed by Z-VAD-FMK treatment. LCH increased the number of sub-G1 cells and arrested the cell cycle at the G1 phase. Taken together LCH inhibits the growth of Ox-sensitive and Ox-resistant CRC cells by targeting EGFR and AKT, and inducing ROS generation and ER stress-mediated apoptosis. Therefore, LCH could be a potential therapeutic agent for improving not only Ox-sensitive but also Ox-resistant CRC treatment.

• Asian Pacific Journal of Cancer Prevention
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• 제14권9호
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• pp.5117-5122
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• 2013

Purpose: To explore the association between PIK3CA and AKT single nucleotide polymorphisms(SNP) and osteosarcoma susceptibility. Methods: TaqMan polymerase chain reaction(PCR) was used to detect the genotypes of SNPs (rs7646409, rs6973569 and rs9866361) in peripheral blood samples from 59 patients with osteosarcoma and from 63 healthy controls. Unconditional logistic regression was used to analyze the correlation between SNPs and osteosarcoma risk. Results: No statistically significant difference was found between osteosarcoma patients and healthy controls in the genotype of AKT rs6973569 (P=0.7). However, after stratified analysis, the genotype AA of AKT rs6973569 carried a higher risk of osteosarcoma metastasis (OR:2.94, 95%CL:1.00-8.59); the difference of rs7646409 genotype distributions between the case and control groups was statistically significant (P=0.032). Taking genotype TT as a reference, the risk of osteosarcoma increased three fold in patients with genotype CC (OR:3.47, 95%CL:1.26-9.56). A statistically significant difference was found between the alleles C and T (P=0.005). Further analysis showed that the risk factor was more pronounced in male patients with Enneking's stage IIB and osteoblastic osteosarcoma. PIK3CA rs9866361 did not fit Hardy-Weinberg equilibrium (P<0.05). Conclusions: Genotype CC in locus PIK3CA rs7646409 may increase the risk of osteosarcoma in the Chinese population.

• Journal of Ginseng Research
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• 제42권4호
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• pp.496-503
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• 2018

Background: Korean ginseng (Panax ginseng) plays an anti-inflammatory role in a variety of inflammatory diseases such as gastritis, hepatitis, and colitis. However, inflammation-regulatory activity of the calyx of the P. ginseng berry has not been thoroughly evaluated. To understand whether the calyx portion of the P. ginseng berry is able to ameliorate inflammatory processes, an ethanolic extract of P. ginseng berry calyx (Pg-C-EE) was prepared, and lipopolysaccharide-activated macrophages and HEK293 cells transfected with inflammation-regulatory proteins were used to test the anti-inflammatory action of Pg-C-EE. Methods: The ginsenoside contents of Pg-C-EE were analyzed by HPLC. Suppressive activity of Pg-C-EE on NO production, inflammatory gene expression, transcriptional activation, and inflammation signaling events were examined using the Griess assay, reverse transcription-polymerization chain reaction, luciferase activity reporter gene assay, and immunoblotting analysis. Results: Pg-C-EE reduced NO production and diminished mRNA expression of inflammatory genes such as cyclooxygenase-2, inducible NO synthase, and tumor necrosis factor-${\alpha}$ in a dose-dependent manner. This extract suppressed luciferase activity induced only by nuclear factor-${\kappa}B$. Interestingly, immunoblotting analysis results demonstrated that Pg-C-EE reduced the activities of protein kinase B (AKT)1 and AKT2. Conclusion: These results suggest that Pg-C-EE may have nuclear-factor-${\kappa}B$-targeted anti-inflammatory properties through suppression of AKT. The calyx of the P. ginseng berry is an underused part of the ginseng plant, and development of calyx-derived extracts may be useful for treatment of inflammatory diseases.