A study of soluble HLA-G1 protecting porcine endothelial cells against human natural killer cell-mediated cytotoxicity
Human natural killer (NK) cells, which can mediate direct lysis of porcine endothelial cells, play an important role in xenograft rejection. HLA-G, which is a critical molecule in maintaining maternal immune tolerance of semi-allogenic fetus, is able to protect susceptible target cells from lysis in...
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| creator | ZENG, M. H FANG, C. Y WANG, S. S ZHU, M XIE, L LI, R WANG, L WU, X. W CHEN, S |
| description | Human natural killer (NK) cells, which can mediate direct lysis of porcine endothelial cells, play an important role in xenograft rejection. HLA-G, which is a critical molecule in maintaining maternal immune tolerance of semi-allogenic fetus, is able to protect susceptible target cells from lysis induced by NK cells. In this study, we investigated whether soluble HLA-G1 (sHLA-G1) protected porcine xenogeneic cells against human NK cell-mediated lysis.
The human sHLA-G1 genomic DNA (pcDNA3-sHLA-G1) was transfected into a B lymphoblastoid cell line 721.221 (LCL721.221) by nucleofector. The sHLA-G1 expression of the transfected LCL721.221 cells was identified by RT-PCR and Dot-ELISA. The sHLA-G1 protein was purified by affinity chromatography on anti-HLA-ImAb W6/32 coupled to cyanogen-bromide-activated Sepharose 4B from culture supernates of transfectants. Various concentrations of sHLA-G(1) protein (0, 2, 4, 6, or 8 microg/mL) were added to a NK cell-mediated xenogenic cell lysis system with either NK92 cells or fresh human peripheral blood mononuclear cells (PBMCs) cocultured with the porcine endothelial cells line. A LDH release assay was used to evaluate NK cell-mediated cytotoxicity.
sHLA-G1 provided significant protection of porcine endothelial cells against human NK-mediated cytotoxicity in a dose-dependent manner. The rates of NK92 cell-mediated cytotoxicity were reduced to 83.4 +/- 5.7% (2 microg/mL), 56.6 +/- 9.3% (4 microg/mL), 39.3 +/- 10.2% (6 microg/mL), and 31.2 +/- 4.9% (8 microg/mL) versus 96.9 +/- 3.0% in the control group (P < .01). Similarly, adding 6 microg/mL sHLA-G1 reduced the mean rate of PBMC-mediated cytotoxicity (n = 4) to 5.8 +/- 1.6% from 23.9 +/- 1.3% in the control group (P < .01).
These results indicated that sHLA-G1 protected xenogeneic porcine endothelial cells against attack by human NK cells, thus providing a new approach to overcome NK-mediated immunity to xenografts. |
| doi_str_mv | 10.1016/j.transproceed.2006.10.179 |
| format | Article |
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The human sHLA-G1 genomic DNA (pcDNA3-sHLA-G1) was transfected into a B lymphoblastoid cell line 721.221 (LCL721.221) by nucleofector. The sHLA-G1 expression of the transfected LCL721.221 cells was identified by RT-PCR and Dot-ELISA. The sHLA-G1 protein was purified by affinity chromatography on anti-HLA-ImAb W6/32 coupled to cyanogen-bromide-activated Sepharose 4B from culture supernates of transfectants. Various concentrations of sHLA-G(1) protein (0, 2, 4, 6, or 8 microg/mL) were added to a NK cell-mediated xenogenic cell lysis system with either NK92 cells or fresh human peripheral blood mononuclear cells (PBMCs) cocultured with the porcine endothelial cells line. A LDH release assay was used to evaluate NK cell-mediated cytotoxicity.
sHLA-G1 provided significant protection of porcine endothelial cells against human NK-mediated cytotoxicity in a dose-dependent manner. The rates of NK92 cell-mediated cytotoxicity were reduced to 83.4 +/- 5.7% (2 microg/mL), 56.6 +/- 9.3% (4 microg/mL), 39.3 +/- 10.2% (6 microg/mL), and 31.2 +/- 4.9% (8 microg/mL) versus 96.9 +/- 3.0% in the control group (P < .01). Similarly, adding 6 microg/mL sHLA-G1 reduced the mean rate of PBMC-mediated cytotoxicity (n = 4) to 5.8 +/- 1.6% from 23.9 +/- 1.3% in the control group (P < .01).
These results indicated that sHLA-G1 protected xenogeneic porcine endothelial cells against attack by human NK cells, thus providing a new approach to overcome NK-mediated immunity to xenografts.</description><identifier>ISSN: 0041-1345</identifier><identifier>EISSN: 1873-2623</identifier><identifier>DOI: 10.1016/j.transproceed.2006.10.179</identifier><identifier>PMID: 17175258</identifier><identifier>CODEN: TRPPA8</identifier><language>eng</language><publisher>New York, NY: Elsevier Science</publisher><subject>Animals ; B-Lymphocytes - immunology ; Biological and medical sciences ; Cell Line ; Cloning, Molecular ; Cytotoxicity, Immunologic ; DNA, Complementary - genetics ; Endothelium, Vascular - drug effects ; Endothelium, Vascular - immunology ; Fundamental and applied biological sciences. Psychology ; Fundamental immunology ; Histocompatibility Antigens Class I - genetics ; Histocompatibility Antigens Class I - pharmacology ; HLA Antigens - genetics ; HLA Antigens - pharmacology ; HLA-G Antigens ; Humans ; Killer Cells, Natural - immunology ; Medical sciences ; Models, Animal ; Reverse Transcriptase Polymerase Chain Reaction ; Surgery (general aspects). Transplantations, organ and tissue grafts. Graft diseases ; Swine ; Tissue, organ and graft immunology ; Transfection ; Transplantation, Heterologous - immunology</subject><ispartof>Transplantation proceedings, 2006-12, Vol.38 (10), p.3312-3314</ispartof><rights>2007 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c262t-c2e6d8c17899f2f5eb21455c380fdc4d0c1ea5cc4bc0d9b89df4e27cde5d136a3</citedby><cites>FETCH-LOGICAL-c262t-c2e6d8c17899f2f5eb21455c380fdc4d0c1ea5cc4bc0d9b89df4e27cde5d136a3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>309,310,314,780,784,789,790,23928,23929,25138,27922,27923</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=18456580$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/17175258$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>ZENG, M. H</creatorcontrib><creatorcontrib>FANG, C. Y</creatorcontrib><creatorcontrib>WANG, S. S</creatorcontrib><creatorcontrib>ZHU, M</creatorcontrib><creatorcontrib>XIE, L</creatorcontrib><creatorcontrib>LI, R</creatorcontrib><creatorcontrib>WANG, L</creatorcontrib><creatorcontrib>WU, X. W</creatorcontrib><creatorcontrib>CHEN, S</creatorcontrib><title>A study of soluble HLA-G1 protecting porcine endothelial cells against human natural killer cell-mediated cytotoxicity</title><title>Transplantation proceedings</title><addtitle>Transplant Proc</addtitle><description>Human natural killer (NK) cells, which can mediate direct lysis of porcine endothelial cells, play an important role in xenograft rejection. HLA-G, which is a critical molecule in maintaining maternal immune tolerance of semi-allogenic fetus, is able to protect susceptible target cells from lysis induced by NK cells. In this study, we investigated whether soluble HLA-G1 (sHLA-G1) protected porcine xenogeneic cells against human NK cell-mediated lysis.
The human sHLA-G1 genomic DNA (pcDNA3-sHLA-G1) was transfected into a B lymphoblastoid cell line 721.221 (LCL721.221) by nucleofector. The sHLA-G1 expression of the transfected LCL721.221 cells was identified by RT-PCR and Dot-ELISA. The sHLA-G1 protein was purified by affinity chromatography on anti-HLA-ImAb W6/32 coupled to cyanogen-bromide-activated Sepharose 4B from culture supernates of transfectants. Various concentrations of sHLA-G(1) protein (0, 2, 4, 6, or 8 microg/mL) were added to a NK cell-mediated xenogenic cell lysis system with either NK92 cells or fresh human peripheral blood mononuclear cells (PBMCs) cocultured with the porcine endothelial cells line. A LDH release assay was used to evaluate NK cell-mediated cytotoxicity.
sHLA-G1 provided significant protection of porcine endothelial cells against human NK-mediated cytotoxicity in a dose-dependent manner. The rates of NK92 cell-mediated cytotoxicity were reduced to 83.4 +/- 5.7% (2 microg/mL), 56.6 +/- 9.3% (4 microg/mL), 39.3 +/- 10.2% (6 microg/mL), and 31.2 +/- 4.9% (8 microg/mL) versus 96.9 +/- 3.0% in the control group (P < .01). Similarly, adding 6 microg/mL sHLA-G1 reduced the mean rate of PBMC-mediated cytotoxicity (n = 4) to 5.8 +/- 1.6% from 23.9 +/- 1.3% in the control group (P < .01).
These results indicated that sHLA-G1 protected xenogeneic porcine endothelial cells against attack by human NK cells, thus providing a new approach to overcome NK-mediated immunity to xenografts.</description><subject>Animals</subject><subject>B-Lymphocytes - immunology</subject><subject>Biological and medical sciences</subject><subject>Cell Line</subject><subject>Cloning, Molecular</subject><subject>Cytotoxicity, Immunologic</subject><subject>DNA, Complementary - genetics</subject><subject>Endothelium, Vascular - drug effects</subject><subject>Endothelium, Vascular - immunology</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Fundamental immunology</subject><subject>Histocompatibility Antigens Class I - genetics</subject><subject>Histocompatibility Antigens Class I - pharmacology</subject><subject>HLA Antigens - genetics</subject><subject>HLA Antigens - pharmacology</subject><subject>HLA-G Antigens</subject><subject>Humans</subject><subject>Killer Cells, Natural - immunology</subject><subject>Medical sciences</subject><subject>Models, Animal</subject><subject>Reverse Transcriptase Polymerase Chain Reaction</subject><subject>Surgery (general aspects). Transplantations, organ and tissue grafts. Graft diseases</subject><subject>Swine</subject><subject>Tissue, organ and graft immunology</subject><subject>Transfection</subject><subject>Transplantation, Heterologous - immunology</subject><issn>0041-1345</issn><issn>1873-2623</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2006</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpNkF9r2zAUxcXoWNNuX2GIQvfmTH8sW-5bCFs7COxlezby1XWrVLFSSS7Lt5_SBtoXCXF-91ydQ8gVZ0vOePN9u8zRTGkfAyDapWCsWR61tvtAFly3shKNkGdkwVjNKy5rdU4uUtqy8ha1_ETOectbJZRekOcVTXm2BxpGmoKfB4_0brOqbjkt_hkhu-me7kMENyHFyYb8gN4ZTwG9T9TcGzelTB_mnZnoZPIci_bovMf4glQ7tM5ktBQOOeTwz4HLh8_k42h8wi-n-5L8_fnjz_qu2vy-_bVebSooEXI5sbEaeKu7bhSjwkHwWimQmo0WasuAo1EA9QDMdoPu7FijaMGislw2Rl6Sb6--JczTjCn3O5eO3zIThjn1jRaqlVoV8OYVhBhSijj2--h2Jh56zvpj6_22f996f2z9RWu7Mvz1tGUeSty30VPNBbg-ASaB8WMxApfeOF2rRmkm_wNcYpLW</recordid><startdate>200612</startdate><enddate>200612</enddate><creator>ZENG, M. H</creator><creator>FANG, C. Y</creator><creator>WANG, S. S</creator><creator>ZHU, M</creator><creator>XIE, L</creator><creator>LI, R</creator><creator>WANG, L</creator><creator>WU, X. W</creator><creator>CHEN, S</creator><general>Elsevier Science</general><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope></search><sort><creationdate>200612</creationdate><title>A study of soluble HLA-G1 protecting porcine endothelial cells against human natural killer cell-mediated cytotoxicity</title><author>ZENG, M. H ; FANG, C. Y ; WANG, S. S ; ZHU, M ; XIE, L ; LI, R ; WANG, L ; WU, X. 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Psychology</topic><topic>Fundamental immunology</topic><topic>Histocompatibility Antigens Class I - genetics</topic><topic>Histocompatibility Antigens Class I - pharmacology</topic><topic>HLA Antigens - genetics</topic><topic>HLA Antigens - pharmacology</topic><topic>HLA-G Antigens</topic><topic>Humans</topic><topic>Killer Cells, Natural - immunology</topic><topic>Medical sciences</topic><topic>Models, Animal</topic><topic>Reverse Transcriptase Polymerase Chain Reaction</topic><topic>Surgery (general aspects). Transplantations, organ and tissue grafts. Graft diseases</topic><topic>Swine</topic><topic>Tissue, organ and graft immunology</topic><topic>Transfection</topic><topic>Transplantation, Heterologous - immunology</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>ZENG, M. H</creatorcontrib><creatorcontrib>FANG, C. Y</creatorcontrib><creatorcontrib>WANG, S. S</creatorcontrib><creatorcontrib>ZHU, M</creatorcontrib><creatorcontrib>XIE, L</creatorcontrib><creatorcontrib>LI, R</creatorcontrib><creatorcontrib>WANG, L</creatorcontrib><creatorcontrib>WU, X. W</creatorcontrib><creatorcontrib>CHEN, S</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>Transplantation proceedings</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>ZENG, M. H</au><au>FANG, C. Y</au><au>WANG, S. S</au><au>ZHU, M</au><au>XIE, L</au><au>LI, R</au><au>WANG, L</au><au>WU, X. W</au><au>CHEN, S</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>A study of soluble HLA-G1 protecting porcine endothelial cells against human natural killer cell-mediated cytotoxicity</atitle><jtitle>Transplantation proceedings</jtitle><addtitle>Transplant Proc</addtitle><date>2006-12</date><risdate>2006</risdate><volume>38</volume><issue>10</issue><spage>3312</spage><epage>3314</epage><pages>3312-3314</pages><issn>0041-1345</issn><eissn>1873-2623</eissn><coden>TRPPA8</coden><abstract>Human natural killer (NK) cells, which can mediate direct lysis of porcine endothelial cells, play an important role in xenograft rejection. HLA-G, which is a critical molecule in maintaining maternal immune tolerance of semi-allogenic fetus, is able to protect susceptible target cells from lysis induced by NK cells. In this study, we investigated whether soluble HLA-G1 (sHLA-G1) protected porcine xenogeneic cells against human NK cell-mediated lysis.
The human sHLA-G1 genomic DNA (pcDNA3-sHLA-G1) was transfected into a B lymphoblastoid cell line 721.221 (LCL721.221) by nucleofector. The sHLA-G1 expression of the transfected LCL721.221 cells was identified by RT-PCR and Dot-ELISA. The sHLA-G1 protein was purified by affinity chromatography on anti-HLA-ImAb W6/32 coupled to cyanogen-bromide-activated Sepharose 4B from culture supernates of transfectants. Various concentrations of sHLA-G(1) protein (0, 2, 4, 6, or 8 microg/mL) were added to a NK cell-mediated xenogenic cell lysis system with either NK92 cells or fresh human peripheral blood mononuclear cells (PBMCs) cocultured with the porcine endothelial cells line. A LDH release assay was used to evaluate NK cell-mediated cytotoxicity.
sHLA-G1 provided significant protection of porcine endothelial cells against human NK-mediated cytotoxicity in a dose-dependent manner. The rates of NK92 cell-mediated cytotoxicity were reduced to 83.4 +/- 5.7% (2 microg/mL), 56.6 +/- 9.3% (4 microg/mL), 39.3 +/- 10.2% (6 microg/mL), and 31.2 +/- 4.9% (8 microg/mL) versus 96.9 +/- 3.0% in the control group (P < .01). Similarly, adding 6 microg/mL sHLA-G1 reduced the mean rate of PBMC-mediated cytotoxicity (n = 4) to 5.8 +/- 1.6% from 23.9 +/- 1.3% in the control group (P < .01).
These results indicated that sHLA-G1 protected xenogeneic porcine endothelial cells against attack by human NK cells, thus providing a new approach to overcome NK-mediated immunity to xenografts.</abstract><cop>New York, NY</cop><pub>Elsevier Science</pub><pmid>17175258</pmid><doi>10.1016/j.transproceed.2006.10.179</doi><tpages>3</tpages></addata></record> |
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| subjects | Animals B-Lymphocytes - immunology Biological and medical sciences Cell Line Cloning, Molecular Cytotoxicity, Immunologic DNA, Complementary - genetics Endothelium, Vascular - drug effects Endothelium, Vascular - immunology Fundamental and applied biological sciences. Psychology Fundamental immunology Histocompatibility Antigens Class I - genetics Histocompatibility Antigens Class I - pharmacology HLA Antigens - genetics HLA Antigens - pharmacology HLA-G Antigens Humans Killer Cells, Natural - immunology Medical sciences Models, Animal Reverse Transcriptase Polymerase Chain Reaction Surgery (general aspects). Transplantations, organ and tissue grafts. Graft diseases Swine Tissue, organ and graft immunology Transfection Transplantation, Heterologous - immunology |
| title | A study of soluble HLA-G1 protecting porcine endothelial cells against human natural killer cell-mediated cytotoxicity |
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