• Title/Summary/Keyword: RT-PCR detection

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Effect of Polydeoxyribonucleotide on Human Periodontal Ligament Cells as a Storage Medium for Avulsed Tooth (탈구치 저장 매체로서 치주인대 세포에 미치는 Polydeoxyribonucleotide의 효과에 대한 연구)

  • Sang Tae Ro;Yong Kwon Chae;Ko Eun Lee;Mi Sun Kim;Ok Hyung Nam;Hyoseol Lee;Sung Chul Choi
    • Journal of the korean academy of Pediatric Dentistry
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    • v.50 no.3
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    • pp.347-359
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    • 2023
  • Objective: This study aimed to evaluate the suitability of polydeoxyribonucleotides (PDRN) as a storage medium for avulsed teeth. Materials and Methods: The viability of human periodontal ligament (PDL) cells stored in Hank's balanced salt solution and PDRN solutions (concentrations, 10, 25, 50, and 100 ㎍/mL) and tap water was measured using the Cell Counting Kit-8 and Live/Dead assays. In addition, Nitric oxide detection and quantitative real-time polymerase chain reaction (qRT-PCR) were performed to evaluate the anti-inflammatory effect of PDRN. Results: The viability of PDL cells stored in a 100 ㎍/mL PDRN solution was significantly higher than that of cells stored in the other solutions (p < 0.01). Furthermore, cells stored in 100 ㎍/mL PDRN solution demonstrated a significantly reduced NO production (p < 0.0001), and cells stored in 50 and 100 ㎍/mL PDRN solutions expressed significantly lower levels of tumor necrosis factor α, interleukin (IL) -4, IL-6, and IL-10 (p < 0.01) compared to cells stored in HBSS. Conclusion: The PDRN solution exhibited cell-preserving and anti-inflammatory effects on the PDL cells. The findings of this study can serve as a basis for further experiments directed at the development of an effective storage medium for avulsed teeth.

Tumor Inhibition Effects and Mechanisms of Angelica sinensis and Sophorae flavescentis ait Decoction Combined with Cisplatin in Xenograft Mice

  • Yan, De-Qi;Liu, Yong-Qi;Li, Ying-Dong;Li, Dou;Cheng, Xiao-Li;Wu, Zhi-Wei
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.11
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    • pp.4609-4615
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    • 2014
  • Background: To investigate tumor inhibition effects and mechanisms of Angelica sinensis and Sophorae flavescentis ait decoction (ASSF) combined with diamine-dichloroplatinum (DDP). Materials and Methods: Bodyweight, tumor inhibition rate and q value were calculated for single ASSF or ASSF combined with DDP on H22 carcinoma xenograft KM mice. Biochemical methods for serum LDH, AST, ALT, and AKP, ELISA method for serum HIF-$1{\alpha}$, pathological assessemnt of thymus, immunohistochemistry detection of tumor tissue caspase3 and mutant p53 protein, and qRT-PCR detection of bax/ bcl-2 mRNA were applied. Results: Compared with DDP control group, the bodyweight increased in ASSF-DDP group (p<0.01). Tumor inhibition rates for DDP, ASSF, ASSF-DDP were 62.7%. 43.7% and 71.0% respectively, with a q value of 0.90. Compared with other groups, thymus of DDP control group had obvious pathological injury (p<0.01), serum LDH, AST, ALT, AKP increased significantly in DDP control group (p<0.01), while serum HIF-$1{\alpha}$ was increased in the model control group. Compared with this latter, the expression of mutant p53 protein and bcl-2 mRNA were decreased in all treatment groups (p<0.01), but there were no statistical difference between DDP control p and ASSF-DDP groups. The expression of caspase3 protein and bax mRNA was increased in all treatment groups, with statistical differences between the DDP and ASSF-DDP groups (p<0.01). Conclusions: ASSF can inhibit bodyweight decrease caused by DDP, can inhibit tumor growth synergistically with DDP mainly through increasing serum HIF-$1{\alpha}$ and pro-apoptotic molecules such as caspase 3 and bax, rather than through decreasing anti-apoptotic mutant p53 and bcl-2. ASSF can reduce DDP toxicity due to decreasing the release of LDH, AST, ALT, AKP into blood and enhancing thymus protection.

Expression of High Mobility Group Box - B1 (HMGB-1) and Matrix Metalloproteinase-9 (MMP-9) in Non-small Cell Lung Cancer (NSCLC)

  • Wang, Jing-Luan;Wu, Da-Wei;Cheng, Zhao-Zhong;Han, Wei-Zhong;Xu, Sheng-Wei;Sun, Ni-Na
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.12
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    • pp.4865-4869
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    • 2014
  • Objective: This study evaluated the expression level of high mobility group box-B1 (HMGB-1) and matrix metalloproteinase-9 (MMP-9) in non-small cell lung cancer (NSCLC) inmorder to reveal any relation with development and prognosis. Methods: NSCLC and normal tissues were selected from 30 patients at age of 30-73, and used for RT-PCR and Western blot analyses of HMGB-1. A total of 100 paraffin embedded NSCLC tissues were also isolated from patients through surgical resection, and used for detection of HMGB-1 by immunohistochemistry. In addition, 50 samples were also applied for MMP-9 detection, and 30 normal tissues were considered as controls. Correlation analysis of HMGB-1 and MMP-9 was carried out by Pearsons correlation coefficient. Results: The average expression level of HMGB-1 in NSCLC patients was significantly higher than in normal lung tissues. In addition, patients in III-IV period exhibit significantly higher positive rate of HMGB-1 when compared with I-II period cases. Furthermore, a positive correlation with HMGB-1 was found in the expression of MPP-9. Conclusion: HMGB-1 was highly expressed in NSCLC, which may become a prognostic and predictive marker for NSCLC. Besides, MPP-9 was positively correlated with HMGB-1.

Expression Analysis of Two Cancer-testis Genes, FBXO39 and TDRD4, in Breast Cancer Tissues and Cell Lines

  • Seifi-Alan, Mahnaz;Shamsi, Roshanak;Ghafouri-Fard, Soudeh;Mirfakhraie, Reza;Zare-Abdollahi, Davood;Movafagh, Abolfazl;Modarressi, Mohammad Hossein;Kazemi, Golnesa;Geranpayeh, Lobat;Najafi-Ashtiani, Mitra
    • Asian Pacific Journal of Cancer Prevention
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    • v.14 no.11
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    • pp.6625-6629
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    • 2013
  • Breast cancer accounts for one third of new cancer cases among women. The need for biomarkers for early detection is the stimulus to researchers to evaluate altered expression of genes in tumours. Cancer-testis (CT) genes are a group with limited expression in normal tissues except testis but up-regulation in a wide variety of cancers. We here evaluated expression of two CT genes named FBXO39 and TDRD4 in 32 invasive ductal carcinoma samples, 10 fibroadenomas and 6 normal breast tissue samples, in addition to two breast cancer cell lines, MCF-7 and MDA-MB-231, by the means of quantitative real time RT-PCR. FBXO39 showed significant up-regulation in invasive ductal carcinoma samples in comparison with normal samples. It also was expressed in both cell lines and after RHOXF1 gene knock down it was down-regulated in MCF-7 but up-regulated in the MDA-MB-231 cell line. TDRD4 was not expressed in the MCF-7 cell line and any of the tissue samples except testis. However, it was expressed in MDA-MB-231 and was up-regulated after RHOXF1 gene knock down. Our results show that FBXO39 but not TDRD4 can be used for cancer detection and if proved to be immunogenic, might be a putative candidate for breast cancer immunotherapy.

Development of the Large-Scale Oligonucleotide Chip for the Diagnosis of Plant Viruses and its Practical Use

  • Nam, Moon;Kim, Jeong-Seon;Lim, Seungmo;Park, Chung Youl;Kim, Jeong-Gyu;Choi, Hong-Soo;Lim, Hyoun-Sub;Moon, Jae Sun;Lee, Su-Heon
    • The Plant Pathology Journal
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    • v.30 no.1
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    • pp.51-57
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    • 2014
  • A large-scale oligonucleotide (LSON) chip was developed for the detection of the plant viruses with known genetic information. The LSON chip contains two sets of 3,978 probes for 538 species of targets including plant viruses, satellite RNAs and viroids. A hundred forty thousand probes, consisting of isolate-, species- and genus-specific probes respectively, are designed from 20,000 of independent nucleotide sequence of plant viruses. Based on the economic importance, the amount of genome information, and the number of strains and/or isolates, one to fifty-one probes for each target virus are selected and spotted on the chip. The standard and field samples for the analysis of the LSON chip have been prepared and tested by RT-PCR. The probe's specific and/or nonspecific reaction patterns by LSON chip allow us to diagnose the unidentified viruses. Thus, the LSON chip in this study could be highly useful for the detection of unexpected plant viruses, the monitoring of emerging viruses and the fluctuation of the population of major viruses in each plant.

Performance of HPV E6/E7 mRNA Genotyping Test on Paired Cervical Cancer Exfoliated Cells and Formalin Fixed Paraffin Embedded Tissues

  • Park, Sunyoung;Wang, Hyeyoung;Kim, Sunghyun;Kim, Geehyuk;Bong, Sungyoung;Jang, Hyoungsoon;Park, Sangjung;Hwang, Kooyeon;Lee, Dongsup
    • Biomedical Science Letters
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    • v.22 no.3
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    • pp.98-106
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    • 2016
  • Investigation of human papillomavirus (HPV) in archival formalin-fixed paraffin-embedded (FFPE) material is important for understanding cervical carcinogenesis. The objective of the present study was to identify the high risk HPVs (HR-HPVs) using HPV E6/E7 mRNA testing from archival tissues in cervical cancer and the relation to HR-HPVs genotypes in paired cervical exfoliated cells. HPV E6/E7 mRNA testing and DNA chip testing were performed in 79 paired cervical FFPE tissues and exfoliated cells from women with histologically confirmed squamous cell carcinoma and adenocarcinoma. Overall agreement in HR-HPVs detection from FFPE samples and cytology samples were 98.5% in HPV 16, 100% in HPV 18, HPV 31, HPV 33, HPV 58, HPV 66, and HPV 68. Type-specific agreement between FFPE samples and cytology samples was 89.1% in HPV positive, 93.5% in HPV 16 and more than 70% in the other HR-HPVs. In conclusion, HR-HPVs were reliably detected in paired FFPE and cytology samples with some variation in type-specific detection.

Development of sandwich enzyme-linked immunosorbent assay for a large-scale detection of porcine transmissible gastroenteritis virus in feces

  • Oh, Yeonsu;Lee, Sang-Joon;Cho, Ho-Seong;Tark, Dongseob
    • Korean Journal of Veterinary Service
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    • v.43 no.4
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    • pp.237-244
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    • 2020
  • Porcine transmissible gastroenteritis (TGE) has been a significant cause of economic losses in pig farming industry since 1950s. Although transmissible gastroenteritis virus (TGEV) has declined in recent years, it should not be excluded because of its characteristics; the frequency of gene mutation, the mortality in piglets, and the possibility for sudden incidence. Therefore, the herd-level monitoring of the virus is important to prevent further circulation of TGE. The aim of this study is to develop a large-scale sandwich enzyme-linked immunosorbent assay (ELISA) with high specificity to rapidly detect TGEV in feces by using monoclonal antibodies (Mabs). The TGEV specific Mabs were produced in hybridoma cells. Among the Mabs belonged to the IgG class developed by this study, the final selected 8H6, 1B7, 4G3, and 1F8 were identified to have the neutralization ability against TGEV. The sandwich ELISA was established using 8H6 as a reporter antibody and 1B7 and the reported 5C8 as a capture antibody. The developed sandwich ELISA was able to distinguish TGEV from other pathogenic diarrheal agents (porcine rotavirus, porcine reovirus, porcine epidemic diarrhea virus (PEDV), E. coli, and C. perfringens) in tissue culture as well as fecal samples. And the detection rate of TGEV in feces was 80% compared with RT-PCR. The results suggested that the developed sandwich ELISA may be useful in the herd-level monitoring for effective preventive measures due to the early diagnosis of TGEV using a large amount of samples.

Association between Respiratory Virus Infection and Pneumococcal Colonization in Children (소아에서의 호흡기바이러스 감염과 비인두 폐렴구균 보균의 연관성)

  • Lee, Hyeon Seung;Choe, Young June;Cho, Eun Young;Lee, Hyunju;Choi, Eun Hwa;Lee, Hoan Jong
    • Pediatric Infection and Vaccine
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    • v.21 no.3
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    • pp.207-213
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    • 2014
  • Purpose: This study aimed to investigate the association between respiratory virus infection and pneumococcal colonization in children. Methods: From May 2009 to June 2010, nasopharyngeal (NP) aspirates were obtained from patients under 18 years old who visited Seoul National University Children's Hospital for respiratory symptoms. NP samples were used to detect respiratory viruses (influenza virus A and B, parainfluenza virus 1, 2 and 3, respiratory syncytial virus A and B, adenovirus, rhinovirus A/B, human metapneumovirus, human coronavirus 229E/NL63 and OC43/HKU1) by RT-PCR and pneumococcus by culture. Results: Median age of the patients was 27 months old. A total of 1,367 NP aspirates were tested for respiratory viruses and pneumococcus. Pneumococcus was isolated from 228 (16.7%) of samples and respiratory viruses were detected from 731 (53.5%). Common viruses were rhinovirus (18.4%), respiratory syncytial virus (RSV) A (10.6%), adenovirus (6.9%), influenza virus A (6.8%). Pneumococcal isolation rate was significantly higher in the cases of positive virus detection than negative detection [21.3% (156/731) vs. 11.3% (72/636), P <0.001]. For individual viruses, pneumococcal isolation rate was positively associated with detection of influenza virus A [24.7% (23/93) vs 16.1% (205/1274), P=0.001], RSV A [28.3% (41/145) vs 15.3% (187/1222), P=0.001], RSV B [31.3% (10/32) vs 16.3% (218/1335), P=0.042], rhinovirus A/B [22.6% (57/252) vs 15.3% (171/1115), P=0.010]. Conclusion: The study revealed that pneumococcal isolation from NP aspirates is related with respiratory virus detection. The result of this study could be used to investigate how respiratory viruses and pneumococcus cause clinical diseases.

PHA-Induced Peripheral Blood Cytogenetics and Molecular Anslysis : a Valid Diagnostic and Follow-up Modality For Acute Primyelocytic Leukemia Patients Treated With ATRA and/or Arsenic Tri-oxide

  • Baba, Shahid M;Azad, Niyaz A;Shah, Zaffar A;Afroze, Dil;Pandith, Arshad A;Jan, Aleem;Aziz, Sheikh A;Dar, Fayaz A
    • Asian Pacific Journal of Cancer Prevention
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    • v.17 no.4
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    • pp.1999-2006
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    • 2016
  • Background: Acute promyelocytic leukemia (APML) is characterized by the reciprocal translocation t(15;17) (p22;p12) resulting in the PML-$RAR{\alpha}$ fusion gene. A dual diagnostic and follow up approach was applied including cytogenetic demonstration of the t(15;17) translocation and detection dg PML-$RAR{\alpha}$ chimeric transcripts by molecular means. Purpose: Conventional cytogenetics involving bone marrow is beset with high probability of poor metaphase index and was substituted with phytohemagglutinin (PHA)-induced peripheral blood culture based cytogenetic analysis as a diagnostic & follow up modality in APML patients of Kashmir (North India). Both qualitative (RT-PCR) and quantitative (Q-PCR) tests were simultaneously carried out to authenticte the modified cytogenetics. Materials and Method: Patient samples were subjected to the said techniques to establish their baseline as well as follow-up status. Results: Initial cytogenetics revealed 30 patients (81%) Positive for t(15;17) whereas 7 (19%) had either cryptic translocation or were negative for t(15;17). Two cases had chromosome 16q deletion and no hallmark translocation t(15;17). Q-PCR status for PML-$RAR{\alpha}$ was found to be positive for all patients. All the APML patients were reassessed at the end of consolidation phase and during maintenance phase of chemotherapy where 6 patients had molecular relapse, wherein 4 also demonstrated cytogenetic relapse. Conclusions: It was found that PHA-induced peripheral blood cytogenetics along with molecular analysis could prove a reliable modality in the diagnosis and assessment of follow up response of APML patients.

Pandemic Novel Influenza A (H1N1) Virus in Korea: The Experience from August to September 2009

  • Lee, Kyung-Ok;Park, Min-Young;Kim, Lyoung-Hyo;Seong, Hye-Soon;Park, Bo-Hyun;Jeong, Su-Jin
    • Korean Journal of Clinical Laboratory Science
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    • v.41 no.4
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    • pp.145-152
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    • 2009
  • Novel influenza A virus, subtype H1N1 of swine-lineage, has been transmitted rapidly to many regions of the world. Rapid detection of the virus is essential to instigate appropriate patient care and public health management and for disease surveillance. The aim of this study is to determine the prevalence of novel influenza A (H1N1) virus in Korea using reverse-transcription real time polymerase chain reaction (rRT-PCR). Novel H1N1 virus was detected in a total of 8,948 nasopharyngeal samples from patients with influenza-like illness throughout Korea from August to September 2009. RNA was extracted from $300{\mu}l$of sample using an RNA extraction kit (Zymo Research, CA, USA). In the present study, Genekam kit (Genekam, Duisburg, Germany) was used to detect novel H1N1 virus. Novel H1N1 virus was found in 1,130 samples from a total of 8,948 samples (12.6%). The highest frequency was found in 10- to 19-year-olds (M: 29.3% vs. F: 16.4%), followed by 20- to 29-year-olds (M: 17.9% vs. F: 15.4%), 40- to 49-year-olds (M: 6.5% vs. F: 8.1%), 50- to 59-year-olds (M: 6.0% vs. F: 5.5%), and 30- to 39-year-olds (M: 4.6% vs. F: 3.8%). The mean positive rate was higher in men than in women (M: 14.7% vs. F: 7.4%). Novel H1N1 virus showed the lowest prevalence in patients over 60 years old. The positive rate increased daily and showed a significant high peak in mid-September 2009. In 19 provinces of Korea, Cheonan (41.1%), Busan (37.3%), Gangneung (33.3%), Jinju (32.1%), Ulsan (24.6%), Deajeon (23.7%) areas showed high frequencies and other provinces were found less than 10% of novel H1N1 virus. Since reverse-transcription real time PCR assay is rapid, accurate, and convenient, it may assist public health laboratories in detecting novel H1N1 virus. Moreover, these data could be useful for the management of patients with influenza-like illness.

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