2.hMSH2 aberrant expression in patients with sporadic colorectal cancer in Xinjiang.
Haiyan WANG ; Zhenqiang SUN ; Ligong YE ; Pei XU ; Yunxia ZHAO ; Lina TANG ; Xiaoyi DUN
Journal of Central South University(Medical Sciences) 2014;39(6):552-557
OBJECTIVE:
To explore the significance of hMSH2 aberrant expression in patients with sporadic colorectal cancer in Xinjiang Uygur Autonomous Region.
METHODS:
Clinicopathological parameters and postoperative samples of 327 patients with sporadic colorectal cancer were collected in Xinjiang Uygur Autonomous Region. Immunohistochemistry PV-9000 two-step method was performed to measure hMSH2 expression in the postoperative pathologic specimens. Prognostic value of hMSH2 expression was evaluated.
RESULTS:
Thirty-five (10.7%) patients showed aberrant nuclear staining of hMSH2 expression. The patients with aberrant expression of hMSH2 showed better prognosis than the normal expression group, with significant difference (P<0.05).
CONCLUSION
In Xinjiang, aberrant hMSH2 expression can be regarded as an independent prognostic factor in patients with sporadic colorectal cancer.
Colorectal Neoplasms
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genetics
;
metabolism
;
Humans
;
Immunohistochemistry
;
MutS Homolog 2 Protein
;
genetics
;
metabolism
;
Prognosis
3.Detection of free DNA septin 9 gene methylation in plasma.
Zhi LUO ; Yingbin HU ; Xiaoyun PU
Journal of Central South University(Medical Sciences) 2021;46(2):127-134
OBJECTIVES:
To explore the correlation between cytosine-phosphoric-guanylic (CpG) site of Septin 9 gene and colorectal cancer, and to develop a real-time PCR detection system in plasma in patients with colorectal cancer.
METHODS:
The methylation of training samples was detected by high-throughput sequencing technology, and the sites highly consistent with the clinical information of colorectal cancer were identified. Then the detection system of real-time PCR was designed to analyze the consistency of plasma and tissue based on methylationa sensitive enzyme digestion. Finally, 100 clinical trials were conducted to evaluate the performance of the detection system with the methylation sensitive enzyme digestion-real-time PCR.
RESULTS:
The highly consistent sites, which were selected by high-throughput sequencing from 71 training set samples, was the 38th CpG. Based on the detection region, the screened methylation sensitive enzymes were
CONCLUSIONS
The 38th CpG site of Septin 9 detected by the detection system of methylation sensitive enzyme digestion-real-time PCR can highly predict the occurrence of colorectal cancer with great clinical application value.
Colorectal Neoplasms/genetics*
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CpG Islands/genetics*
;
DNA
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DNA Methylation
;
Humans
;
Plasma/metabolism*
;
Septins/metabolism*
4.Expression, purification, and characterization of the histidine kinase CarS from Fusobacterium nucleatum.
Zhuting LI ; Xian SHI ; Ruochen FAN ; Lulu WANG ; Tingting BU ; Wei ZHENG ; Xuqiang ZHANG ; Chunshan QUAN
Chinese Journal of Biotechnology 2023;39(4):1596-1608
Fusobacterium nucleatum is an opportunistic pathogenic bacterium that can be enriched in colorectal cancer tissues, affecting multiple stages of colorectal cancer development. The two-component system plays an important role in the regulation and expression of genes related to pathogenic resistance and pathogenicity. In this paper, we focused on the CarRS two-component system of F. nucleatum, and the histidine kinase protein CarS was recombinantly expressed and characterized. Several online software such as SMART, CCTOP and AlphaFold2 were used to predict the secondary and tertiary structure of the CarS protein. The results showed that CarS is a membrane protein with two transmembrane helices and contains 9 α-helices and 12 β-folds. CarS protein is composed of two domains, one is the N-terminal transmembrane domain (amino acids 1-170), the other is the C-terminal intracellular domain. The latter is composed of a signal receiving domain (histidine kinases, adenylyl cyclases, methyl-accepting proteins, prokaryotic signaling proteins, HAMP), a phosphate receptor domain (histidine kinase domain, HisKA), and a histidine kinase catalytic domain (histidine kinase-like ATPase catalytic domain, HATPase_c). Since the full-length CarS protein could not be expressed in host cells, a fusion expression vector pET-28a(+)-MBP-TEV-CarScyto was constructed based on the characteristics of secondary and tertiary structures, and overexpressed in Escherichia coli BL21-Codonplus(DE3)RIL. CarScyto-MBP protein was purified by affinity chromatography, ion-exchange chromatography, and gel filtration chromatography with a final concentration of 20 mg/ml. CarScyto-MBP protein showed both protein kinase and phosphotransferase activities, and the MBP tag had no effect on the function of CarScyto protein. The above results provide a basis for in-depth analysis of the biological function of the CarRS two-component system in F. nucleatum.
Humans
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Histidine Kinase/metabolism*
;
Fusobacterium nucleatum/metabolism*
;
Automobiles
;
Protein Kinases/genetics*
;
Escherichia coli/metabolism*
;
Colorectal Neoplasms
5.Mutational studies of adenomatous polyposis coli gene in carcinomas from patients with hereditary non-polyposis colorectal cancers.
Jian HUANG ; Shen-hang JIN ; Shu-zhan ZHANG ; Shu ZHENG
Chinese Journal of Medical Genetics 2003;20(3):196-199
OBJECTIVETo analyze the mutational features of adenomatous polyposis coli (APC) gene and to explore the effect of mismatch repair (MMR) deficiency on its mutations in hereditary non-polyposis colorectal cancers (HNPCC).
METHODSPCR-based in vitro synthesized protein test (IVSP) assay and sequencing analysis were used to confirm somatic mutations of whole APC gene in 19 HNPCC patients.
RESULTSEleven cases with thirteen mutations were determined. The frequency of APC mutation was 58%(11/19). The exhibiting mutations consisted of 9 frameshift mutations and 4 nonsense ones, indicating the existence of more frameshift mutations (69%). All of frameshift mutations were deletion or insertion of 1-2 bp and most of them (7/9) happened at simple nucleotide repeat sequences, particularly within (A) n tracts (5/9). All of four nonsense mutations resulted from C to T transitions at CpG sites.
CONCLUSIONMutational inactivations of APC gene were detected in more than half of HNPCC patients in this study, indicating that APC mutation is a common molecular event in the tumorigenesis of HNPCC. According to the location of frameshift mutations at simple nucleotide repeat sequences and point mutations at CpG sites, it was suggested that endogenous mechanisms like MMR deficiency might exert an effect on the nature of APC mutations in most HNPCC.
Adenomatous Polyposis Coli ; genetics ; Adenomatous Polyposis Coli Protein ; genetics ; metabolism ; Carcinoma ; genetics ; Colorectal Neoplasms ; genetics ; pathology ; Colorectal Neoplasms, Hereditary Nonpolyposis ; genetics ; Genes, APC ; physiology ; Humans
6.Expression of NOB1 and its significance in colorectal cancer.
Journal of Southern Medical University 2012;32(3):420-422
OBJECTIVETo investigate the expression of NOB1 in colorectal cancer and its relationship with the clinicopathological characteristics.
METHODSThe expression of NOB1 was detected immunohistochemically in 60 primary colorectal cancer tissues and the corresponding normal epithelia (3.0 cm away from the cancer margin) and graded according to the staining intensity and the percentage of positively stained tumor cells.
RESULTSNOB1 overexpression was found in 32 of the 60 cases (53.3%). NOB1 overexpression in the adjacent non-neoplastic tissues was found in 10 of the cases (16.7%), a rate significantly lower than that in the cancer tissues (P<0.05). NOB1 expression was not correlated to such tumor characteristics as gender, age, histological differentiation grade, depth of invasion and lymph node metastasis (P>0.05).
CONCLUSIONSNOB1 expression is higher in colorectal cancer than in normal colorectal tissues, suggesting its involvement in the tumorigenesis and progression of colorectal cancer.
Colorectal Neoplasms ; genetics ; metabolism ; pathology ; Female ; Humans ; Immunohistochemistry ; Male ; Middle Aged ; Nuclear Proteins ; genetics ; metabolism ; RNA-Binding Proteins ; genetics ; metabolism
7.Omics docking and molecular classification in colorectal cancer.
Journal of Central South University(Medical Sciences) 2010;35(11):1201-1206
Omics docking has become a hot spot in oncology research, which is used to screen specific biomarkers for the establishment of molecular classification in the treatment and/or prognosis of diseases, especially in cancer research. Colorectal cancer (CRC) is one of the common malignant tumors, the traditional diagnosis and treatment of which depends on clinical manifestations, classic pathological and imaging examination. In clinical pathology, completely different fates and prognoses were observed in CRC patients with the same type, at the same stage, and even with the same treatment. It is critical to use the omics docking strategy to select molecular biomarkers for early diagnosis and to assess the prognosis of CRC, to further standardize molecular classification model, and to guide individual treatment of CRC.
Animals
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Biomarkers, Tumor
;
genetics
;
metabolism
;
Colorectal Neoplasms
;
classification
;
diagnosis
;
genetics
;
Genomics
;
Humans
;
Proteomics
8.Clinicopathological features of rectal adenocarcinoma with enteroblastic differentiation.
J LIU ; X L LIU ; D L LIN ; H ZHAO ; Y J LI ; X M XING
Chinese Journal of Pathology 2023;52(8):797-801
Objective: To investigate the clinicopathological features, immunophenotype, and genetic alterations of rectal adenocarcinoma with enteroblastic differentiation. Methods: Four cases of rectal adenocarcinoma with enteroblastic differentiation were collected at the Affiliated Hospital of Qingdao University, Qingdao, China (three cases) and Yantai Yeda Hospital of Shandong Province, China (one case) from January to December 2022. Their clinical features were summarized. Hematoxylin and eosin stain and immunohistochemical stain were performed, while next-generation sequencing was performed to reveal the genetic alterations of these cases. Results: All four patients were male with a median age of 65.5 years. The clinical manifestations were changes of stool characteristics, bloody stools and weight loss. All cases showed mixed morphology composed of conventional adenocarcinoma and adenocarcinoma with enteroblastic differentiation. Most of the tumors consisted of glands with tubular and cribriform features. In one case, almost all tumor cells were arranged in papillary structures. The tumor cells with enteroblastic differentiation were columnar, with relatively distinct cell boundaries and characteristic abundant clear cytoplasm, forming fetal gut-like glands. Immunohistochemically, the tumor cells were positive for SALL4 (4/4), Glypican-3 (3/4) and AFP (1/4, focally positive), while p53 stain showed mutated type in 2 cases. The next-generation sequencing revealed that 2 cases had TP53 gene mutation and 1 case had KRAS gene mutation. Conclusions: Rectal adenocarcinoma with enteroblastic differentiation is rare. It shows embryonal differentiation in morphology and immunohistochemistry, and should be distinguished from conventional colorectal adenocarcinoma.
Humans
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Male
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Aged
;
Female
;
Biomarkers, Tumor/metabolism*
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Adenocarcinoma/pathology*
;
Colorectal Neoplasms
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Rectal Neoplasms/genetics*
;
Cell Differentiation
9.Topographic expression of p21WAF1/SDI1/CIP1, bcl2, and p53 is altered at the early stage of colorectal carcinogenesis.
Jeong Hee CHO ; Im Hwan ROE ; Chang Young LIM ; Dong Kook PARK ; Woo Ho KIM ; Yong Il KIM
Journal of Korean Medical Science 2000;15(6):667-674
We analyzed the expression of p21, bcl2, and p53 in normal and different pathologic mucosa of the human colorectum using immunohistochemistry and cold polymerase chain reaction-single strand conformation polymorphism. The topography of normal mucosa showed; bcl2 and p53 expression restricted to basal epithelial cells and p21 expressed only in superficial epithelial cells. This topographic expression was altered in hyperplastic polyps and adenomas. Hyperplastic polyps revealed absence of or weak bcl2 expression and strong p21 expression without topography. In adenomas, whereas bcl2 expression increased and extended to parabasal and superficial dysplastic epithelium, the increase of p21 expression was limited to surface dysplastic epithelium. p53 was weakly expressed throughout the full thickness of dysplastic epithelium. Bcl2 expression in adenomas was stronger than in carcinomas; p53 expression was converse and p21 expression was variable. In carcinomas, this topographic expression was largely abrogated but p53 mutation (36%) was more frequent than in adenomas (2%). In carcinomas, p21 and p53 expression correlated inversely, but there was no relationship with bcl2. These results suggest that there is precisely ordered topographic pattern of p21, bcl2, and wild p53 expression in normal colorectal cells, but this becomes disordered during the early stage of colorectal carcinogenesis.
Colorectal Neoplasms/physiopathology
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Colorectal Neoplasms/pathology
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Colorectal Neoplasms/metabolism*
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Colorectal Neoplasms/genetics
;
Cyclins/biosynthesis*
;
Human
;
Mutagenesis
;
Protein p53/genetics
;
Protein p53/biosynthesis*
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Proto-Oncogene Proteins c-bcl-2/biosynthesis*
;
Time Factors
10.Down regulation of multidrug resistance-associated protein 4 expression by RNA interference enhances radiosensitivity of colorectal carcinoma cell lines in vitro.
Zhi-qi YU ; Chang ZHANG ; Rui CAI ; Xin-yuan LAO ; Hao WANG ; Xian-hua GAO ; Yi-fang HAN ; Xiao-qing ZHANG ; Guang-wen CAO ; Chuan-gang FU
Chinese Journal of Gastrointestinal Surgery 2012;15(1):67-71
OBJECTIVETo investigate the effect of multidrug resistance-associated protein 4 (MRP4) expression on the radiosensitivity of colorectal carcinoma cell lines in vitro.
METHODSThe vector of shRNA for RNA interference was constructed and then transfected into HCT116 cell line to steadily down-regulate the expression of MRP4. HCT116 cells were divided into 3 groups including the CON group(non-transfected), NC group (negative control virus was added), and KD group (RNAi target was added for transfection). To test the effectiveness of RNA interference, real-time polymerase chain reaction and Western blot were used to measure the expression pattern of MRP4 at both mRNA and protein levels, respectively. For the examination of the effect of RNA interference of MRP4 on the radiosensitivity, flow cytometry was used to calculate the rate of apoptotic cells 24 h after 4 Gy radiation. Proliferation of the cells was measured via MTT assay at different time points.
RESULTSShRNA plasmid was successfully constructed. Transfection of this constructed vector into HCT116 cell line caused steady silencing of MRP4 expression (HCT116-KD). MRP4 mRNA and protein expression were significantly down-regulated following RNA interference(P<0.05). Twenty-four hours after radiation, the apoptosis rate of KD cell line was (71.7±0.8)%, significantly higher than that in the CON group [(56.1±0.9)%] and NC group[(59.8±0.8)%](P<0.05). Fourty-eight hours and 72 hours after radiation, the proliferation was significantly inhibited in KD cells compared to the control groups(P<0.05).
CONCLUSIONSExpression of MRP4 is closely related to radio-tolerance of colorectal carcinoma. Down-regulation of MRP4 expression by RNA interference enhances radiosensitivity of colorectal carcinoma cell lines in vitro. MRP4 may be an effective molecular marker for predicting the radiosensitivity of colorectal carcinoma.
Colorectal Neoplasms ; genetics ; metabolism ; Down-Regulation ; HCT116 Cells ; Humans ; Multidrug Resistance-Associated Proteins ; genetics ; RNA Interference ; Radiation Tolerance ; genetics