Animals ; Bufo bufo ; Caspase 3 ; genetics ; metabolism ; Female ; Immunohistochemistry ; Male ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; metabolism ; Proto-Oncogene Proteins c-kit ; genetics ; metabolism ; Spinal Cord ; metabolism ; Spinal Cord Injuries ; metabolism ; bcl-2-Associated X Protein ; genetics ; metabolism

Gastrointestinal stromal tumors (GISTs) of the gallbladder are representative of an extremely rare group of tumors. We have encountered a patient with a malignant GIST of the gallbladder and presented it with a review of some articles. A 72-yr-old woman initially presented with right upper quadrant abdominal pain, fever and chills. Emergency cholecystectomy was performed under the impression of gallbladder empyema. Liver metastasis was found at 7 months postoperatively and the patient expired 9 months after the surgery. At the time of cholecystectomy, the gallbladder showed a necrotic serosal surface with an irregular thickened wall. A mass, 6 cm in length and 3 cm in width, encircled the whole wall of the neck and upper body of the gallbladder. Microscopic findings revealed frequent mitotic figures (more than 20/50 HPF) and tumor necrosis. Hyperchromatic, pleomorphic and spindle shaped neoplastic cells that were arranged in a pattern of short fascicles infiltrated the entire layer of the gallbladder. The tumor cells were immunoreactive for CD117 antigen (c-kit protein) and vimentin. They were negative for desmin, smooth muscle actin and S-100 protein. Mutations of the c-kit proto-oncogene were not found in this case. These findings were sufficient to provide enough clinical, histopathological and immunohistochemicalevidence in diagnosing our case as a malignant GIST.
Aged ; Female ; Gallbladder Neoplasms/genetics/metabolism/*pathology ; Gastrointestinal Stromal Tumors/genetics/metabolism/*pathology ; Humans ; Proto-Oncogene Protein c-kit/genetics/metabolism

We report a case of a 38-yr-old man with a spontaneously ruptured gastric stromal tumor presenting as hemoperitoneum in outpatient clinic. He visited our hospital with generalized abdominal pain after abdominal CT scan for the evaluation of the asymptomatic palpable abdominal mass. Repeated abdominal CT scan showed a size decrement of cystic mass compared with the previous abdominal CT scan, and newly developed fluid collection in the left paracolic gutter. An emergency laparotomy revealed a ruptured gastric stromal tumor with bloody fluid in the peritoneal cavity. Immunohistochemical examination revealed positive reactivity to C-kit protein and CD34. The patient presented with hemoperitoneum due to spontaneous rupture of the tumor, which is an extremely rare complication.
Adult ; Antigens, CD34/biosynthesis ; Hemoperitoneum ; Human ; Immunohistochemistry ; Male ; Mitosis ; Proto-Oncogene Protein c-kit/metabolism ; Stomach Neoplasms/*diagnosis/pathology ; Stomach Rupture ; Tomography, X-Ray Computed

OBJECTIVETo compare the differentiation ability difference of hematopoietic, mesenchymal and endothelial potential between CD41⁺ cells derived from the mouse aorta-gonadmesonephros (AGM) region, yolk sac (YS) and embryonic circulating blood (CB).

METHODSCD41⁺ cells were sorted from AGM, YS and CB. The CD45 and c-kit expression were studied in CD41⁺ cells by flow cytometry. IL-3 and bone morphogenetic protein 4 (BMP-4) treatment together with semi solid culture were used to assess hematopoietic potential difference of CD41⁺ cells. Immunofluorescence staining of α-SMA was used to assess mesenchymal potential difference. The endothelial cell induction system was used to assess endothelial potential difference.

RESULTSThe proportions of CD45+ cells in CD41⁺ population were 51.9% (AGM), 45.8% (YS) and 22.2% (CB), respectively, while those of c-kit⁺ cells were 40.0% (AGM), 39.6% (YS) and 36.2% (CB), respectively. After stimulated by IL-3 factor, the number of total colonies increased in all three groups-derived CD41⁺ cells compared to that of unstimulated group[(14.1±1.9) vs (1.2±0.2), (32.4±1.1) vs (18.4±2.2) and (41.8±0.9) vs (10.4±1.8)], (P<0.01). After stimulated by BMP-4 factor, compared to unstimulated group, CFU-Mix colony number in CD41⁺ cells from AGM region and YS were significantly decreased[(0.5±0.6) vs (3.2±0.8), (1.3±0.7) vs (7.4±1.7)](P<0.01), but there was no difference in CB group[(2.5±0.5) vs (3.9±1.5)](P>0.01). The mesenchymal marker α-SMA was highly expressed in CD41⁺ cells from AGM region and YS, but lowly expressed in CD41⁺ cells from CB.

CONCLUSIONThere are some differences between CD41⁺ cells in AGM region, YS and CB on hematopoietic cell surface marker expression, hematopoietic colony formation with IL-3 and BMP-4 stimulation.

Animals ; Aorta ; cytology ; Bone Morphogenetic Protein 4 ; pharmacology ; Cell Differentiation ; Gonads ; cytology ; Interleukin-3 ; pharmacology ; Mesonephros ; cytology ; Mice ; Platelet Membrane Glycoprotein IIb ; metabolism ; Proto-Oncogene Proteins c-kit ; metabolism ; Yolk Sac ; cytology

This study was aimed to investigate the expression level of SHP-1 and C-kit genes in acute leukemia HL-60 cells and effect of inhibitor As2O3 demethylation on SHP-1 and C-kit genes expression. RT-PCR was used to detect the expression level of SHP-1 and C-kit mRNA in drug-treated cell group and control group. The methylation specific PCR (MSP) was applied to measure the methylation status of SHP-1 gene in HL-60 cells. The results showed that after being treated with As2O3 the recovery of SHP-1 gene expression was observed in HL-60 cells in which SHP-1 mRNA originally did not expressed, meanwhile the expression level of C-kit mRNA in HL-60 cells with high expression decreased. When HL-60 cells were treated with As2O3 of 1.0, 2.5, 5.0 µmol/L, the demethylation effects was enhanced, the expression of SHP-1 mRNA displayed an ascending tendency, and expression of C-kit mRNA showed an descending tendency in dose-dependent manner (P < 0.05). It is concluded that the absence of SHP-1 mRNA expression in HL-60 cells and recovery of expression after treatment with As2O3 suggest the hypermethylation of SHP-1 gene related with pathogenesis of leukemia, and the abnormal increase of C-kit mRNA expression maybe exist in formation of leukemia. The effect of As2O3 on expression of SHP-1 and C-kit shows dose-dependency, the higher the As2O3 concentration, the higher the SHP-1 expression and the lower the C-kit expression, moreover, the effect of As2O3 shows time-dependency in specific concentration. The SHP-1 mRNA expression negatively relates with C-kit mRNA expression, suggesting that the decrease or absence of SHP-1 expression in leukemia cells weakens the negative regulation on C-kit signaling pathway, thus plays a role in the formation of leukemia.
Arsenicals ; metabolism ; pharmacology ; DNA Methylation ; Gene Expression Regulation, Leukemic ; HL-60 Cells ; Humans ; Oxides ; metabolism ; pharmacology ; Protein Tyrosine Phosphatase, Non-Receptor Type 6 ; genetics ; Proto-Oncogene Proteins c-kit ; genetics

OBJECTIVETo explore the effect of Imatinib mesylate on proliferation, differentiation and apoptosis of leukemic Kasumi-1 cells bearing c-kit mutation.

METHODSKasumi-1 cells were treated with Imatinib at different concentrations in culture. Cell proliferation was assayed by MTT assay, expressions of c-kit antigen, surface myeloid antigen and cell cycle by flow cytometry, cell apoptosis by annexin V staining and agarose gel electrophoresis. Western blot was used to analyze the level of c-kit protein tyrosine phosphorylation.

RESULTSImatinib treatment caused a time- and dose-dependent inhibition of the cell proliferation, with a 72 h IC50 of 4.45 micromol/L. Imatinib treatment induced a decrease in the mean fluorescence value of c-kit antigen, a progressive decline in S-phase cell fraction and an increase in G0/G1 cells. Treatment with 5.00 micromol/L of imatinib for 72 h induced an increase in expression of myeloid surface protein CD11, CD13 and CD15, and for 24 h induced an increase in early apoptosis cells [from 9.04% to 86.84% (P < 0.05)]. The apoptosis ladder was observed on agarose gel electrophoresis on 5-day treatment. Tyrosine phosphorylation level of c-kit protein was decreased by Imatinib treatment.

CONCLUSIONTyrosine kinase inhibitor Imatinib mesylate treatment could inhibit proliferation of Kasumi-1 cells which bear a c-kit mutation, induce differentiation, apoptosis and G0/G1 cells accumulation.

Apoptosis ; drug effects ; genetics ; Benzamides ; Cell Differentiation ; drug effects ; genetics ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Humans ; Imatinib Mesylate ; Mutation ; Piperazines ; pharmacology ; Protein-Tyrosine Kinases ; antagonists & inhibitors ; Proto-Oncogene Proteins c-kit ; genetics ; metabolism ; Pyrimidines ; pharmacology

This study was aimed to investigate the expression level of SHP-1 and C-kit genes in acute leukemia HL-60 cells and effect of 5-aza-CdR demethylation on expression of SHP-1 and C-kit genes. RT-PCR was used to detect the mRNA expression level of SHP-1 and C-kit mRNA in HL-60 cells of the drug-treated group and control group.The methylation specific PCR (MSP) was applied to measure the methylation status of SHP-1 and C-kit genes in HL-60 cells.The results showed that after being treated with 5-aza-CdR, the recovery of SHP-1 gene expression was observed in HL-60 cells in which SHP-1 mRNA originally was not expressed. Meanwhile, the high expression level of C-kit mRNA in HL-60 cells was decreased. When HL-60 cells were treated with 0, 0.5, 1.0, 2.0 µmol/L 5-aza-CdR, the demethylation effect was enhanced, the expression of SHP-1 mRNA displayed an ascending tendency, and the expression of C-kit mRNA showed an descending tendency in dose-dependent manner (P < 0.05) . It is concluded that the absence of SHP-1 mRNA expression in HL-60 cells and recovery of expression after treatment with 5-aza-CdR suggest that the hypermethylation of SHP-1 gene relates with pathogenesis of leukemia, and the abnormal increase of C-kit mRNA expression maybe exist in formation of leukemia. The effect of 5-aza-CdR on expression of SHP-1 and C-kit shows dose-dependency, the higher the 5-aza-CdR concentration, the higher the SHP-1 expression and the lower the C-kit expression, moreover, the effect of 5-aza-CdR shows time-dependency in specific concentration.The SHP-1 mRNA expression negatively correlates with C-kit mRNA expression, suggesting that the decrease or absence of SHP-1 expression in leukemia cells weakens the negative regulation on C-kit signaling pathway, thus plays a role in the formation of leukemia.
Azacitidine ; pharmacology ; DNA Methylation ; drug effects ; HL-60 Cells ; Humans ; Leukemia ; genetics ; metabolism ; Protein Tyrosine Phosphatase, Non-Receptor Type 6 ; genetics ; Proto-Oncogene Proteins c-kit ; genetics ; RNA, Messenger

In carcinoma ex pleomorphic adenoma (CXPA), pleomorphic adenoma (PA) and diverse carcinoma components showing luminal (ductal) or non-luminal (myoepithelial) differentiation coexist. To elucidate the clinicopathological implications of cellular differentiation in CXPA and the potential role of p53, vascular endothelial growth factor (VEGF), c-erbB-2, c-kit, and glucose transporter 1 (Glut-1) in carcinogenesis, we analyzed 11 CXPAs with luminal differentiation (CXPAs-LD) and 6 CXPAs with non-luminal differentiation (CXPAs-NLD) and compared protein expressions in residual PAs and carcinomas by immunohistochemistry. Among the CXPAs-LD, 5 were invasive and 8 were histologically high-grade tumors. The 5-year survival rate was 72.7%. P53, c-erbB-2, VEGF, and Glut-1 were more immunoreactive in carcinoma components than in PAs (P = 0.008, 0.004, 0.002, and 0.024, respectively); c-erbB-2 overexpression was associated with high histological grade (P = 0.024). Carcinoma components frequently lacked c-kit expression (P = 0.009). CXPAs-NLD were all low-grade and invasive with a larger mean tumor size (5.2 cm) than CXPAs-LD (3.3 cm) (P = 0.040). The patients remained disease-free without significant immunohistochemical expression. The immunoprofiles and clinical course of CXPA differed according to cellular differentiation. Therefore, it is important to report the histological subtype and to assess potential biomarkers in diagnostic and therapeutic trials.
Adenoma, Pleomorphic/*immunology/metabolism/*pathology ; Adult ; Aged ; Carcinoma/*immunology/metabolism/*pathology ; Cell Differentiation ; Female ; Glucose Transport Proteins, Facilitative/metabolism ; Humans ; Male ; Middle Aged ; Proto-Oncogene Proteins c-kit/metabolism ; Receptor, erbB-2/metabolism ; Salivary Gland Neoplasms/*immunology/metabolism/*pathology ; Tumor Markers, Biological/*analysis ; Tumor Suppressor Protein p53/metabolism ; Vascular Endothelial Growth Factor A/metabolism

Cyclin-Dependent Kinase 6 ; metabolism ; Gene Expression Regulation, Neoplastic ; Genes, p53 ; Humans ; Killer Cells, Natural ; pathology ; Lymphoma, T-Cell ; genetics ; metabolism ; pathology ; Mutation ; Nose Neoplasms ; genetics ; metabolism ; pathology ; Protein Tyrosine Phosphatase, Non-Receptor Type 6 ; Proto-Oncogene Proteins c-kit ; metabolism ; fas Receptor ; genetics ; metabolism

BACKGROUNDThe addition of anti-human epidermal growth factor receptor 2 (HER2)-targeted drugs, such as trastuzumab, lapatinib, and trastuzumab emtansine (T-DM1), to chemotherapy significantly improved prognosis of HER2-positive breast cancer patients. However, it was confused that metastatic patients vary in the response of targeted drug. Therefore, methods of accurately predicting drug response were really needed. To overcome the spatial and temporal limitations of biopsies, we aimed to develop a more sensitive and less invasive method of detecting mutations associated with anti-HER2 therapeutic response through circulating-free DNA (cfDNA).

METHODSFrom March 6, 2014 to December 10, 2014, 24 plasma samples from 20 patients with HER2-positive metastatic breast cancer who received systemic therapy were eligible. We used a panel for detection of hot-spot mutations from 50 oncogenes and tumor suppressor genes, and then used targeted next-generation sequencing (NGS) to identify somatic mutation of these samples in those 50 genes. Samples taken before their first trastuzumab administration and subsequently proven with clinical benefit were grouped into sensitive group. The others were collected after disease progression of the trastuzumab-based therapy and were grouped into the resistant group.

RESULTSA total of 486 single-nucleotide variants from 46 genes were detected. Of these 46 genes, phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA), proto-oncogene c-Kit (KIT), and tumor protein p53 (TP53) were the most common mutated genes. Seven genes, including epidermal growth factor receptor (EGFR), G protein subunit alpha S (GNAS), HRas proto-oncogene (HRAS), mutL homolog 1 (MLH1), cadherin 1 (CDH1), neuroblastoma RAS viral oncogene homolog (NRAS), and NOTCH1, that only occurred m utations in the resistant group were associated with the resistance of targeted therapy. In addition, we detected a HER2 S855I mutation in two patients who had persistent benefits from anti-HER2 therapy.

CONCLUSIONTargeted NGS of cfDNA has potential clinical utility to detect biomarkers from HER2-targeted therapies.

Adolescent ; Adult ; Aged ; Biomarkers, Tumor ; genetics ; Breast Neoplasms ; genetics ; metabolism ; Cadherins ; genetics ; Chromogranins ; genetics ; Class I Phosphatidylinositol 3-Kinases ; Drug Resistance, Neoplasm ; genetics ; Female ; GTP-Binding Protein alpha Subunits, Gs ; genetics ; Humans ; Male ; Middle Aged ; Mutation ; genetics ; Phosphatidylinositol 3-Kinases ; genetics ; Proto-Oncogene Proteins c-kit ; genetics ; Receptor, ErbB-2 ; metabolism ; Receptor, Notch1 ; genetics ; Tumor Suppressor Protein p53 ; genetics ; Young Adult