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ABSTRACT: This study was aimed to investigate the relationship between immunophenotype of the background lymphocytes and histological subtype of Hodgkin's lymphoma (HL), and its significance. The relative protein expressions of background lymphocytes were detected in 37 HL specimens on the basis of instant-rapid MaxVision(TM) immunohistochemical method and assessed quantitatively with image analysis software IPP6.0. The adoptive antibody included anti-CD3/CD45RO, anti-CD20/CD79a, anti-CD4, anti-CD8, anti-GrB, anti-TIA-1. The results indicated that out of 37 cases 4 were NLPHL, 33 were CHL including 6 of MCHL, 14 of NSHL, 13 of LRHL. In addition, 10 cases (1 was NLPHL, 4 were NSHL, 5 were LRHL) were involved in the analysis of T/B ratios. The ratio of T/B in NLPHL was 0.28 +/- 0.07, in CHL 4.34 +/- 2.45 (p = 0.001), in CHL the ratio was LRHL > NSHL > MCHL (p = 0.649); CD4(+)/CD8(+) ratio in NLPHL was 4.55 +/- 1.28, in CHL 4.10 +/- 1.50 (p = 0.574), in CHL it was MCHL > NSHL > LRHL (p = 0.037); GrB(+)/TIA-1(+) ratio in NLPHL was 0.71 +/- 0.57, in CHL 0.74 +/- 0.39, it was MCHL > NSHL > LRHL (p > 0.05). It is conduced that immune cell composition of the diagnostic HL lymph node represents the immune microenvironment. It is different between NLPHL and CHL in terms of the T- and B-lymphocyte distribution. NLPHL is of unique feature. The subtypes of CHL are of peculiar. The T/B ratio is in order as LRHL > NSHL > MCHL, but CD4(+)/CD8(+) and GrB(+)/TIA-1(+) ratios are in the opposite order. Combining with prognosis of subtypes of CHL i.e, LRHL > NSHL > MCHL, these data suggest that low ratio of T/B with high ratios of CD4(+)/CD8(+) and GrB(+)/TIA-1(+) may represent biological markers predicting an unfavorable outcome of CHL subtypes.
Zhongguo shi yan xue ye xue za zhi / Zhongguo bing li sheng li xue hui = Journal of experimental hematology / Chinese Association of Pathophysiology 08/2009; 17(4):888-93.
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ABSTRACT: To construct a recombinant lentivirus vector of latent membrane protein 1 (LMP1) and detect the expression of LMP1 in vitro.
The LMP1 fragment including all the exons was amplified by PCR and inserted to the downstream of CMV promoter in the lentivirus vector pCDF. The three plasmids (packaging plasmid pFIV-34N, envelope plasmid pVSV-G and target plasmid pCDF-LMP1) were packaged into 293FT cells via liposome. The virus supernatant was harvested, concentrated and titrated. Mouse B lymphoma cell line A20 was transfected with the recombinant lentivirus vector of LMP1, and the expression of LMP1 in A20 cells was detected by RT-PCR and Western blotting.
DNA sequencing confirmed that the sequence of PCR-amplified LMP1 was consistent with the GenBank data. The LMP1 gene fragment was cloned into pCDF in the right direction, and the open reading frame of LMP1 was maintained. The 3 plasmids were effectively transferred into 293FT cells, which emitted green fluorescence in the cytoplasm and on the cell membrane under fluorescence microscope. The titer of the lentivirus vector reached 10(7) Tu/ml with a transfection efficiency 90% in A20 cells. LMP1 expression was detected by RT-PCR and Western blotting in transfected A20 cells.
The recombinant lentivirus vector of LMP1 constructed can be effectively transfected into A20 cells, which provides a basis for exploring the role of LMP1 in the pathogenesis of lymphoma.
Nan fang yi ke da xue xue bao = Journal of Southern Medical University 06/2009; 29(5):837-40.
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ABSTRACT: To construct a recombinant lentivirus harboring RNA interference sequence targeting mouse CD99 antigen-like 2 (mCD99L2) gene and observe its infection efficiency of 293FT cells.
Four pairs of small interfering RNAs (siRNAs) targeting mCD99L2 cDNA were designed, synthesized and linked to the lentivirus vector SD1259 to construct the lentivirus shuttle plasmids. After sequencing, the 4 lentivirus shuttle plasmids were transfected into 293FT cells in the presence of packaging plasmids. Forty-eight hours later, the supernatant was collected and the titer and infection efficiency of the recombinant lentivirus were determined according to the expression of the reporter gene enhanced green fluorescent protein (EGFP) under fluorescent microscope.
DNA sequencing demonstrated that mCD99L2 siRNAs were successfully cloned to the lentiviral vector SD1259. The titer of concentrated virus was 1x10(7)/ml in the supernatant of the infected cells.
The recombinant lentivirus containing siRNA targeting mCD99L2 gene has been successfully constructed, which provide the basis for future establishment of visualized cell model and animal model of Hodgkin's lymphoma.
Nan fang yi ke da xue xue bao = Journal of Southern Medical University 03/2009; 29(2):228-31.
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ABSTRACT: To compare the efficacy of nuclear microarray combined with fluorescence in situ hybridization (FISH) and immunohistochemistry in detecting ALK gene translocation and ALK fusion protein in anaplastic large cell lymphoma (ALCL).
ALK gene translocation and ALK fusion protein in 17 paraffin-embedded ALCL specimens were detected using nuclear microarray combined with FISH and immunohistochemical straining, respectively.
The expression of ALK fusion protein was detected immunohistochemically with ALK antibody in 8 of the 17 specimens of systemic ALCL, including 4 with both nuclear and cytoplasmic positivity and 4 with only cytoplasmic positivity. Dual-color FISH identified 6 positive specimens, including the 4 specimens with both nuclear and cytoplasmic positivity as identified immunohistochemically, and 2 with immunohistochemical cytoplasmic positivity. FISH yielded negative results for the 2 specimens with immunohistochemical cytoplasmic positivity.
Nuclear microarray combined with FISH eliminated the cytoplasmic interference of the results of conventional FISH and provides a high-throughput platform for clinical detection with greater specificity than immunohistochemistry.
Nan fang yi ke da xue xue bao = Journal of Southern Medical University 05/2008; 28(4):572-5.
