• Asian Pacific Journal of Cancer Prevention
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• v.17 no.8
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• pp.4089-4093
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• 2016

Purpose: This study aimed to explore the association of ${\beta}-catenin$ expression pattern with pathological response after neoadjuvant chemotherapy in breast cancer (BC) patients. Materials and Methods: In this retrospective exploratory study, data for 50 BC patients who received neoadjuvant chemotherapy were recorded. ${\beta}-catenin$ expression in tumours was assessed using immunohistochemistry and classified as either membranous or cytoplasmic according to the pattern of staining. Distributions of different clinico-pathological parameters according to ${\beta}-catenin$ expression were assessed using the Chi-square test. Logistic regression analysis was used to assess any relation of the pattern of ${\beta}-catenin$ expression with the pathological response. Results: Cytoplasmic ${\beta}-catenin$ expression was detected in 34% of BCs. Among our cases, 52% were hormonal receptor (HR)-positive, 24% were HER2-positive, 74% were clinical stage III and 74% received both anthracycline and taxane-based chemotherapy. Patients with cytoplasmic expression were more commonly younger than 40 years at diagnosis (cytoplasmic, 41.2% vs. no cytoplasmic expression, 12.1%, p=0.03). By doing t-test, cytoplasmic ${\beta}-catenin$ expression was linked with a higher body mass index compared to membranous-only expression ($mean{\pm}SD$ $33.0{\pm}4.47$ vs. $29.6{\pm}6.01$, respectively, p=0.046). No significant associations were found between ${\beta}-catenin$ expression and other parameters such as HR and HER2 status, or clinical stage. Complete pathological response (pCR) rate was twice as great in patients with membranous expression but without statistical significance (membranous-only, 33.3% vs. cytoplasmic, 17.6%, OR= 2.3, 95% CI= 0.55-9.87, p=0.24). Conclusions: This study suggests that cytoplasmic ${\beta}-catenin$ expression may be linked with lower probability of achieving pCR after neoadjuvant chemotherapy. These data need to be validated in a larger cohort of patients.

• Archives of Pharmacal Research
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• v.19 no.4
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• pp.275-279
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• 1996

To study the functional roles of dopamine receptors, we decided to raise antibodies against these proteins. To make antigen, we expressed the whole 3rd cytoplasmic loop of dopamine receptors in a fusion protein with glutathione-S-transferase (GST). $For D_2\; and\; D_3$ receptors, it was successful to express and purify fusion proteins for the whole 3rd cytoplasmic loops. However, we could not express the fusion protein for the whole 3rd cytoplasmic loop of $D_4$ dopamine receptor in the bacteria. To study the causes that prevent the bacterial expression of the GST-fusion protein of the 3rd cytoplasmic loop of $D_4$ dopamine receptor, we conducted more detailed studies on $D_4$ dopamine receptor. To locate the region which prevents bacterial expression, we made sequential constructs in the 3rd cytoplasmic loop decreasing the size step by step, and confirmed their expressions in the SDS PAGE. It was found that certain regions of 3rd cytoplasmic loop of $D_4$ dopamine receptor, located in N-terminal side of the 3rd cytoplasmic loop of $D_4$ dopamine receptor suppress the bacterial expression of fusion protein.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.5
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• pp.2805-2809
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• 2013

Objective: This study aimed at investigating whether the orphan nuclear receptor NR4A2 is significantly associated with clinicopathologic features and overall survival of patients with nasopharyngeal carcinoma (NPC). Methods: Immunohistochemistry was performed to determine NR4A2 protein expression in 84 NPC tissues and 20 non-cancerous nasopharyngeal (NP) tissues. The prognostic significance of NR4A2 protein expression was evaluated using Cox proportional hazards regression models and Kaplan-Meier survival analysis. Results: We did not find a significant association between total NR4A2 expression and clinicopathological variables in 84 patients with NPC. However, we observed that high cytoplasmic expression of NR4A2 was significantly associated with tumor size (T classification) (P = 0.006), lymph node metastasis (N classification) (P = 0.002) and clinical stage (P = 0.017). Patients with higher cytoplasmic NR4A2 expression had a significantly lower survival rate than those with lower cytoplasmic NR4A2 expression (P = 0.004). Multivariate Cox regression analysis analysis suggested that the level of cytoplasmic NR4A2 expression was an independent prognostic indicator for overall survival of patients with NPC (P = 0.033). Conclusions: High cytoplasmic expression of NR4A2 is a potential unfavorable prognostic factor for patients with NPC.

• IMMUNE NETWORK
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• v.8 no.4
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• pp.130-136
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• 2008

Background: Human cytomegalovirus UL18, a MHC class I homologue, has been considered a natural killer (NK) cell decoy. It ligates LIR-1/ILT2 (CD85j), an NK inhibitory receptor, to prevent lysis of infected target cells. However, precise role of UL18 to NK cell cytotoxicity is yet elusive. Difficulty in clarifying the function of UL18 lies in complication in detecting UL18 mainly due to low level expression of UL18 on the surface and gradual loss of its expression. Methods: To overcome this hurdle, cDNA of cytoplasmic tail-less UL18 was constructed and expressed in swine endothelial cell (SEC). The expression level and its stability in the cell surface were monitored with FACS analysis. Results: Surface expression of UL18 is up-regulated by removing cytoplasmic tail portion from UL18F (a full sequence of UL18). SECs transfected with a cDNA of UL18CY (a cytoplasmic tail-less UL18) stably expressed UL18 molecule on the surface without gradual loss of its expression during 6 week continuous cultures. In the NK cytotoxicity assay, UL18 functions either inhibiting or activating NK cell cytotoxicity according to the source of NK cells. We found that there is individual susceptibility in determining whether the engagement of NK cell and UL18 results in overall inhibiting or activating NK cell cytotoxicity. Conclusion: In this study, we found that cytoplasmic tail is closely related to the regulatory function for controlling surface expression of UL18. Furthermore, by constructing stable cell line in which UL18 expression is up-regulated and stable, we provided a useful tool to clarify exact functions of UL18 on various immune cells having ILT2 receptor.

• Journal of Pathology and Translational Medicine
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• v.52 no.6
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• pp.369-377
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• 2018

Background: Chemokine receptor CXC chemokine receptor type 4 (CXCR4) and its ligand CXC motif chemokine 12 (CXCL12; stromal cell-derived factor-1) are implicated in tumor growth, metastasis, and tumor cell-microenvironment interaction. A number of studies have reported that increased CXCR4 expression is associated with worse prognosis in triple-negative breast cancer (TNBC), but its prognostic significance has not been studied in TNBC patients treated with adjuvant chemotherapy. Methods: Two hundred eighty-three TNBC patients who received adjuvant chemotherapy were retrospectively analyzed. Tissue microarray was constructed from formalin-fixed, paraffin-embedded tumor tissue and immunohistochemistry for CXCR4 and CXCL12 was performed. Expression of each marker was compared with clinicopathologic characteristics and outcome. Results: High cytoplasmic CXCR4 expression was associated with younger age (p=.008), higher histologic grade (p=.007) and lower pathologic stage (p=.045), while high CXCL12 expression was related to larger tumor size (p=.045), positive lymph node metastasis (p=.005), and higher pathologic stage (p=.017). The patients with high cytoplasmic CXCR4 experienced lower distant recurrence (p=.006) and better recurrence-free survival (RFS) (log-rank p=.020) after adjuvant chemotherapy. Cytoplasmic CXCR4 expression remained an independent factor of distant recurrence (p=.019) and RFS (p=.038) after multivariate analysis. Conclusions: High cytoplasmic CXCR4 expression was associated with lower distant recurrence and better RFS in TNBC patients treated with adjuvant chemotherapy. This is the first study to correlate high CXCR4 expression to better TNBC prognosis, and the underlying mechanism needs to be elucidated in further studies.

• Asian-Australasian Journal of Animal Sciences
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• v.27 no.4
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• pp.567-573
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• 2014

Hormonal and nutrient signals regulate leptin synthesis and secretion. In rodents, leptin is stored in cytosolic pools of adipocytes. However, not much information is available regarding the regulation of intracellular leptin in ruminants. Recently, we demonstrated that leptin mRNA was expressed in bovine intramuscular preadipocyte cells (BIP cells) and that a cytoplasmic leptin pool may be present in preadipocytes. In the present study, we investigated the expression of cytoplasmic leptin protein in BIP cells during differentiation as well as the effects of various factors added to the differentiation medium on its expression in BIP cells. Leptin mRNA expression was observed only at 6 and 8 days after adipogenic induction, whereas the cytoplasmic leptin concentration was the highest on day 0 and decreased gradually thereafter. Cytoplasmic leptin was detected at 6 and 8 days after adipogenic induction, but not at 4 days after adipogenic induction. The cytoplasmic leptin concentration was reduced in BIP cells at 4 days after treatment with dexamethasone, whereas cytoplasmic leptin was not observed at 8 days after treatment. In contrast, acetate significantly enhanced the cytoplasmic leptin concentration in BIP cells at 8 days after treatment, although acetate alone did not induce adipocyte differentiation in BIP cells. These results suggest that dexamethasone and acetate modulate the cytoplasmic leptin concentration in bovine preadipocytes.

• Journal of Plant Biotechnology
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• v.32 no.4
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• pp.269-273
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• 2005

Toll-like receptors (TLR) are important components of innate immunity in the defense against pathogens. TLRs recognize pathogen-associated common molecular patterns. TLRs are similar to the receptors involved in defense responses in plants. TLR protein is a type 1 membrane protein, consisting of an extracellular domain containing leucine-rich repeats and a cytoplasmic domain. The cytoplasmic domain delivers ligand recognition signals that result in production of anti-microbial agents. The cytoplasmic domain (amino acid 858-1032) of toll-like receptor 9 has been expressed using methylotrophic yeast Pichia pastoris. The protein expression was confirmed by Western-blot, N-terminal sequencing and MALDl-TOF mass spectrometry. The proteins have been purified by nickel affinity, cation exchange and gel-filtration chromatography.

• Bulletin of the Korean Chemical Society
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• v.28 no.9
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• pp.1549-1552
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• 2007

The cytoplasmic domain of syndecan-4, a type I transmembrane heparan sulfate proteoglycan, was overexpressed as a fused form with the ubiquitin molecule in Escherichia coli, and the fusion protein was purified using immobilized metal affinity chromatography (IMAC). The cytoplasmic domain was released from its fusion partner by using yeast ubiquitin hydrolase (YUH), and subsequently purified by reverse phase chromatography. The integrity of the resulting peptide fragment was checked by MALDI-TOF and NMR spectroscopy. The yield of the peptide was 3.0-1.5 mg per liter in LB or minimal medium, respectively. The recombinant expression and purification of this domain will enable us its structural and functional studies using multidimensional NMR spectroscopy.

• Fisheries and Aquatic Sciences
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• v.14 no.4
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• pp.303-310
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• 2011

We characterized a cytoplasmic actin gene locus in greenling Hexagrammos otakii (Scorpaeniformes). Genomic clones isolated from the greenling DNA library contained two homologous cytoplasmic actin gene copies (HObact2.1 and HObact2.2) in a tail-to-head orientation. Their gene structure is characterized by six translated exons and one non-translated exon. Exon-intron organization and the nucleotide sequences of the two actin gene isoforms are very similar. However, only the HObact2.1 isoform contains microsatellite-like, dinucleotide repeats in the 5'-flanking region (named HOms2002) and intron 1 following the non-translated exon 1 (named HOms769). One microsatellite locus (HOms769) was highly polymorphic while the other (HOms2002) was not. Based on bioinformatic analysis, different transcription factor binding motifs are related to stress and immune responses in the two actin isoforms. Semiquantitative and real-time reverse transcription-PCR assays showed that both isoform transcripts were detectable ubiquitously in all the tissues examined. However, the basal expression levels of each isoform varied across tissues. Overall, the two isoforms showed a similar, but not identical, expression pattern. Our data suggest that the cytoplasmic actin genes may be the result of a recent duplication event in the greenling genome, which has not experienced significant subfunctionalization in their housekeeping roles.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.5
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• pp.1943-1948
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• 2012

Introduction: Prostate cancer in Indonesia is the $3^{rd}$ ranking cancer among males and the $5^{th}$ rank for their cancer mortality. Prognostic markers that can identify aggressive prostate cancer in early stages and help select appropriate therapy to finally reduce the mortality are therefore urgently needed. It has been suggested that stem cells in the prostate gland have a role in initiation, progression, and metastasis of cancer, although controversy continues to exist. Maintenance of normal stem cell or reserve cell populations in several epithelia including prostate has been shown to be regulated by p63 and alteration of p63 expression is considered to have an oncogenic role in prostate cancer. We hypothesize that the expression of cytoplasmic aberrance of p63 is associated with high ALDH1A1 expression as a cancer stem cell marker, thus leading to progression of prostate cancer. Methods: Using a cross-sectional study during two years (2009-2010), a total of 79 paraffin embedded tissues of benign prostatic hyperplasia, PIN prostatic intraepithelial neoplasia, low and high Gleason score prostate cancer were investigated using immunohistochemistry. Associations between cytoplasmic p63 and ALDH1A1, as well as with pathological diagnosis, were analyzed by Chi-Square test using SPSS 15.0. Links of both markers with cell proliferation rate (KI-67) and apoptotic rate (cleaved caspase 3) were also analyzed by Kruskal-Wallis test. Results: The mean age of patient at the diagnosis is 70.0 years. Cytoplasmic aberrance of p63 was associated with ALDH1A1 expression (p<0.001) and both were found to have significant relationships with pathological diagnosis (including Gleason score), (p=0.006 and p<0.001 respectively). Moreover, it was also found that higher levels of cytoplasmic p63 were significantly associated with the frequency of proliferating cells and cells undergoing apoptosis in prostate cancers (p=0.001 and p=0.016 respectively). Conclusion: p63 cytoplasmic aberrance is associated with high ALDH1A1 expression. These components are suggested to have an important role in prostate cancer progression and may be used as molecular markers.