The two hybridoma cell lines secreting antikeratin monoclonal antibodies (McAb)were obtained from fusion between NS_1 myelema cells and spleen cells of BALB/cmice immunized with keratin prepared from human callus.The two cell lines(AF_2and AF_5)have a chromosome number of 97 and 96 respectively.By immunodiffusion,both AF_2 and AF_5 McAb belong to mouse IgG_1 subclass.They have been successivelypassaged for over six months in vitro,and can grow well in the peritoneal fluid ofBALB/c mice,both produce high titer of specific antibodies. To determine their specificity with keratin of epithelia,AF_2 and AF_2 McAbwere used to stain various human normal and neoplastic tissue sections by the ABC(Avidin-Biotin peroxidase complex)immunohistochemical techniques.The results showthat AF_2 and AF_5 McAb can stain a series of normal epithelial tissues(in epidermis,positive for cells above the basal layer)and epithelial tumor tissues.No staining wereobserved in nonepithelial normal and tumor tissues.These results suggest that the twoMcAb are quite specific for a definite number of cytokeratins and promise to be use-ful in the differential diagnosis of tumors.

AIM: To study the effects of (-)-epigallocatechin-3-gallate (EGCG), a tea extract, on the invasion and metastasis of breast cancer cell line MDA-MB-231 and the possible mechanisms in vitro. METHODS: The expression of MUC1 in breast cancer cells treated with or without EGCG was detected by immunohistochemistry. The effect of EGCG on invasion of MDA-MB-231 cells was evaluated using Transwell chambers attached with polycarbonate filters and reconstituted basement membrane (Matrigel). Gelatin zymography was performed to detect the secretion of collagenase-Ⅳ. RESULTS: EGCG reduced the expression of MUC1, significantly suppressed the invasion of tumor cells to basement membrane and reduced the secretion of collagenase-Ⅳ. CONCLUSION: In vitro, EGCG may suppress invasion, metastasis, and collagenase-Ⅳ secretion in MDA-MB-231 cells by inhibiting the production of MUC1.

Objective To investigate the correlation between the HSP90 expression and invasion and metastasis potential of human breast cancer cell line. Methods The mRNA expression level of HSP90 was detected by RT-PCR. The protein expression level of HSP90 was measured by Western blotting. The effects of HSP90 inhibitor Geldanamycin (GA) on the invasion and motility abilities of human breast cancer line MDA-MB-231 and MDA-MB-435s in vitro were explored by transwell chamber. Results The mRNA expression level of HSP90? in MDA-MB-435s cell was higher than that of MDA-MB-231 cell (P0.05) between the two cell lines. The protein expression level of HSP90 in MDA-MB-435s cell was higher than that of MDA-MB-231 cell (P0.05). After treated with GA, the invasive and metastatic potential in human breast cancer cell MDA-MB-435s and MDA-MB-231 was suppressed significantly (P

OBJECTIVETo investigate the effect of co-expression of bone morphogenetic protein 2 (BMP2) and Sox9 on chondrogenic differentiation of mesenchymal stem cells (MSCs) in vitro and provide experimental evidence for tissue engineering of cartilage.

METHODSMouse embryonic bone marrow MSC C3H10T1/2 cells were infected with recombinant adenovirus expressing BMP2, Sox9 and green fluorescent protein (GFP) for 3-14 days, with cells infected with the adenovirus carrying GFP gene as the control. The mRNA expression of the markers of chondrogenic differentiation, including collagen type II (Col2a1), aggrecan (ACAN), and collagen type X (Col10a1), were determined by real-time PCR. Alcian blue staining was used for quantitative analysis of sulfated glycosaminoglycan in the cellular matrix. The expression of Col2a1 protein was assayed by immunohistochemical staining and Western blot analysis.

RESULTSAdenovirus-mediated BMP2 expression induced chondrogenic differentiation of C3H10T1/2 cells. Overexpression of Sox9 effectively enhanced BMP2-induced expression of the chondrogenic markers Col2a1, aggrecan and Col10a1 mRNAs, and promoted the synthesis of sulfated glycosaminoglycan and Col2a1 protein in C3H10T1/2 cells.

CONCLUSIONCo-expression of BMP2 and Sox9 can promote chondrogenic differentiation of MSCs in vitro, which provides a new strategy for tissue engineering of cartilage.

Animals ; Bone Morphogenetic Protein 2 ; genetics ; metabolism ; Cartilage ; cytology ; Cell Differentiation ; Cells, Cultured ; Chondrocytes ; cytology ; Humans ; Mesenchymal Stromal Cells ; cytology ; metabolism ; Mice ; SOX9 Transcription Factor ; genetics ; metabolism ; Tissue Engineering

Objective To investigate the effect of co-expression of bone morphogenetic protein 2 (BMP2) and Sox9 on chondrogenic differentiation of mesenchymal stem cells (MSCs) in vitro and provide experimental evidence for tissue engineering of cartilage. Methods Mouse embryonic bone marrow MSC C3H10T1/2 cells were infected with recombinant adenovirus expressing BMP2, Sox9 and green fluorescent protein (GFP) for 3-14 days, with cells infected with the adenovirus carrying GFP gene as the control. The mRNA expression of the markers of chondrogenic differentiation, including collagen type II (Col2a1), aggrecan (ACAN), and collagen type X (Col10a1), were determined by real-time PCR. Alcian blue staining was used for quantitative analysis of sulfated glycosaminoglycan in the cellular matrix. The expression of Col2a1 protein was assayed by immunohistochemical staining and Western blot analysis. Results Adenovirus-mediated BMP2 expression induced chondrogenic differentiation of C3H10T1/2 cells. Overexpression of Sox9 effectively enhanced BMP2-induced expression of the chondrogenic markers Col2a1, aggrecan and Col10a1 mRNAs, and promoted the synthesis of sulfated glycosaminoglycan and Col2a1 protein in C3H10T1/2 cells. Conclusion Co-expression of BMP2 and Sox9 can promote chondrogenic differentiation of MSCs in vitro, which provides a new strategy for tissue engineering of cartilage.

Objective To investigate the effect of co-expression of bone morphogenetic protein 2 (BMP2) and Sox9 on chondrogenic differentiation of mesenchymal stem cells (MSCs) in vitro and provide experimental evidence for tissue engineering of cartilage. Methods Mouse embryonic bone marrow MSC C3H10T1/2 cells were infected with recombinant adenovirus expressing BMP2, Sox9 and green fluorescent protein (GFP) for 3-14 days, with cells infected with the adenovirus carrying GFP gene as the control. The mRNA expression of the markers of chondrogenic differentiation, including collagen type II (Col2a1), aggrecan (ACAN), and collagen type X (Col10a1), were determined by real-time PCR. Alcian blue staining was used for quantitative analysis of sulfated glycosaminoglycan in the cellular matrix. The expression of Col2a1 protein was assayed by immunohistochemical staining and Western blot analysis. Results Adenovirus-mediated BMP2 expression induced chondrogenic differentiation of C3H10T1/2 cells. Overexpression of Sox9 effectively enhanced BMP2-induced expression of the chondrogenic markers Col2a1, aggrecan and Col10a1 mRNAs, and promoted the synthesis of sulfated glycosaminoglycan and Col2a1 protein in C3H10T1/2 cells. Conclusion Co-expression of BMP2 and Sox9 can promote chondrogenic differentiation of MSCs in vitro, which provides a new strategy for tissue engineering of cartilage.

Objective To study the predictive value of magnetic resonance imaging (MRI),positron emission computed tomography (PET)and PET/MRI coregistration in intractable epilepsy of children.Methods A retrospec-tively analysis was performed based on the surgery data at the Department of Children Epilepsy Center of Peking Univer-sity First Hospital from September 2015 to March 2016.The clinical data,surgery and follow－up study data,interictal and ictal electroencephalogram,MRI,PET and PET/MRI coregistration data were collected.By comparison with the epi-leptogenic zone designed by pre－surgical workup,the accuracy of MRI,PET and PET/MRI coregistration in detecting lesion was assessed.In the patients who had no seizure during≥1 year follow－up,their sensitivity,specificity,positive predictive value,negative predictive value of MRI,PET and PET/MRI coregistration were calculated.Results (1)A total of 62 patients underwent surgery,30 boys and 32 girls. The average age on epilepsy onset was 2. 50 years (2 days－11.70 years),and average age on surgery was 5.10 years old(0.75－15.60 years old).(2)Surgical treat-ment of 62 cases included the resection of the focal or lobar (32 cases,51.6%),and the multilobar (16 cases, 25. 8%).Hemispherotomy was done in 14 cases (22.6%).During ≥1 year follow－up,seizure outcome was Engel class Ⅰ in 57 cases (91.9%)out of the 62 patients,Engel classⅡto Engel classⅣin 1 case,3 cases,and 1 case, respectively.(3)Referred to epileptogenic zone designed by presurgical workup,MRI represented 64.5%(40/62 ca-ses)results with accordance,PET and PET/MRI coregistration was 72.5%(45/62 cases)and 85.5%(53/62 cases), respectively,and the difference was significant(χ2＝7.25,P＝0.03).(4)Based on the patients of Engel class Ⅰ, their sensitivity and specificity were 66.7%,60.0% in MRI,75.4%,60.0% in PET %,and 85.9%,80.0% in PET/MRI coregistration,respectively.(5)There were 11 ＂non－lesion＂ cases of all focal cortical dysplasia in patholo-gy,and subtle structural abnormalities were de tected in 9 cases by reviewing MRI.Conclusions PET/MRI coregistra-tion can improve lesion detection of intractable epilepsy in children.