Limits...
An improved protocol for efficient engraftment in NOD/LTSZ-SCIDIL-2Rγ mice allows HIV replication and development of anti-HIV immune responses.

Singh M, Singh P, Gaudray G, Musumeci L, Thielen C, Vaira D, Vandergeeten C, Delacroix L, Van Gulck E, Vanham G, de Leval L, Rahmouni S, Moutschen M - PLoS ONE (2012)

Bottom Line: In the present study, we further ameliorated the busulfan myeloablation regimen with fresh CB-CD34+cell transplantation in 3-4 week old NSG mice.Optimal engraftment promoted early and increased CD3+T cell levels, with better lymphoid tissue development and prolonged human cell chimerism over 300 days.We also saw a gradual decline of the CD4+T cell count, widespread immune activation, up-regulation of inflammation marker and microbial translocation after HIV-1 infection.

View Article: PubMed Central - PubMed

Affiliation: Immunology and infectious diseases unit GIGA-I3, University of Liege, Liege, Belgium.

ABSTRACT
Cord blood hematopoietic progenitor cells (CB-HPCs) transplanted immunodeficient NOD/LtsZ-scidIL2Rγ() (NSG) and NOD/SCID/IL2Rγ() (NOG) mice need efficient human cell engraftment for long-term HIV-1 replication studies. Total body irradiation (TBI) is a classical myeloablation regimen used to improve engraftment levels of human cells in these humanized mice. Some recent reports suggest the use of busulfan as a myeloablation regimen to transplant HPCs in neonatal and adult NSG mice. In the present study, we further ameliorated the busulfan myeloablation regimen with fresh CB-CD34+cell transplantation in 3-4 week old NSG mice. In this CB-CD34+transplanted NSG mice engraftment efficiency of human CD45+cell is over 90% in peripheral blood. Optimal engraftment promoted early and increased CD3+T cell levels, with better lymphoid tissue development and prolonged human cell chimerism over 300 days. These humanized NSG mice have shown long-lasting viremia after HIV-1JRCSF and HIV-1Bal inoculation through intravenous and rectal routes. We also saw a gradual decline of the CD4+T cell count, widespread immune activation, up-regulation of inflammation marker and microbial translocation after HIV-1 infection. Humanized NSG mice reconstituted according to our new protocol produced, moderate cellular and humoral immune responses to HIV-1 postinfection. We believe that NSG mice reconstituted according to our easy to use protocol will provide a better in vivo model for HIV-1 replication and anti-HIV-1 therapy trials.

Show MeSH

Related in: MedlinePlus

Human cell differentiation in humanized NSG mice.(A) Absolute number of human cells in lymph nodes, bone marrow and spleens of group 1 and group 7 mice. (B) Mesenteric and brachial lymph nodes from male mice obtained after 24 weeks of CD34+ cell transplantation in group 7 mice. (C) Flow cytometric analysis of engraftment percentage of CD3+ T cells and CD19+ B cells at 12 and 22 weeks after transplantation of CD34+ cells in representative mice from group 1 and group 7. (D) Flow cytometric analysis of CD4+CD8+, CD4+ and CD8+ population in thymus of 22 weeks engrafted mice. (E) Flow cytometric analysis of CD45RA+ and CD45RO+ positive CD3 cells 22 weeks after engraftment in mice from group 7. (F) Representative profile CD11c+ cells in spleen and CD14+ population in bone marrow and blood of group 7 mice 22 weeks after CD34+ cell transplantation.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3366932&req=5

pone-0038491-g002: Human cell differentiation in humanized NSG mice.(A) Absolute number of human cells in lymph nodes, bone marrow and spleens of group 1 and group 7 mice. (B) Mesenteric and brachial lymph nodes from male mice obtained after 24 weeks of CD34+ cell transplantation in group 7 mice. (C) Flow cytometric analysis of engraftment percentage of CD3+ T cells and CD19+ B cells at 12 and 22 weeks after transplantation of CD34+ cells in representative mice from group 1 and group 7. (D) Flow cytometric analysis of CD4+CD8+, CD4+ and CD8+ population in thymus of 22 weeks engrafted mice. (E) Flow cytometric analysis of CD45RA+ and CD45RO+ positive CD3 cells 22 weeks after engraftment in mice from group 7. (F) Representative profile CD11c+ cells in spleen and CD14+ population in bone marrow and blood of group 7 mice 22 weeks after CD34+ cell transplantation.

Mentions: We next evaluated if the overall improvement of human cell engraftment associated with group 7 protocol was observed in all lymphoid sites and specifically correlated with a better or faster differentiation of a given cell subset (i.e. CD19, CD3, CD4, CD8). We compared engraftment achieved in groups 1 and 7, in blood, spleen, lymph nodes and bone marrow at 12 and 22 weeks after transplantation of CB-CD34+ cells. At 22 weeks, high percentages of human CD45+ cells were observed in the lymph nodes (95.51±3.23%), bone marrow (83.41±2.65%) and spleen (74.31±2.92%) (n = 20) (Table 3). Humanized mice of group 7 also had higher absolute numbers of human cells in lymph nodes, bone marrow and spleen compared with group 1 (Fig. 2A). Twenty-four weeks after transplantation of fresh CB-CD34+ cells, lymph nodes of sizes from 3 to 7 mm could be observed in NSG mice from group 7 (Fig. 2B).


An improved protocol for efficient engraftment in NOD/LTSZ-SCIDIL-2Rγ mice allows HIV replication and development of anti-HIV immune responses.

Singh M, Singh P, Gaudray G, Musumeci L, Thielen C, Vaira D, Vandergeeten C, Delacroix L, Van Gulck E, Vanham G, de Leval L, Rahmouni S, Moutschen M - PLoS ONE (2012)

Human cell differentiation in humanized NSG mice.(A) Absolute number of human cells in lymph nodes, bone marrow and spleens of group 1 and group 7 mice. (B) Mesenteric and brachial lymph nodes from male mice obtained after 24 weeks of CD34+ cell transplantation in group 7 mice. (C) Flow cytometric analysis of engraftment percentage of CD3+ T cells and CD19+ B cells at 12 and 22 weeks after transplantation of CD34+ cells in representative mice from group 1 and group 7. (D) Flow cytometric analysis of CD4+CD8+, CD4+ and CD8+ population in thymus of 22 weeks engrafted mice. (E) Flow cytometric analysis of CD45RA+ and CD45RO+ positive CD3 cells 22 weeks after engraftment in mice from group 7. (F) Representative profile CD11c+ cells in spleen and CD14+ population in bone marrow and blood of group 7 mice 22 weeks after CD34+ cell transplantation.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3366932&req=5

pone-0038491-g002: Human cell differentiation in humanized NSG mice.(A) Absolute number of human cells in lymph nodes, bone marrow and spleens of group 1 and group 7 mice. (B) Mesenteric and brachial lymph nodes from male mice obtained after 24 weeks of CD34+ cell transplantation in group 7 mice. (C) Flow cytometric analysis of engraftment percentage of CD3+ T cells and CD19+ B cells at 12 and 22 weeks after transplantation of CD34+ cells in representative mice from group 1 and group 7. (D) Flow cytometric analysis of CD4+CD8+, CD4+ and CD8+ population in thymus of 22 weeks engrafted mice. (E) Flow cytometric analysis of CD45RA+ and CD45RO+ positive CD3 cells 22 weeks after engraftment in mice from group 7. (F) Representative profile CD11c+ cells in spleen and CD14+ population in bone marrow and blood of group 7 mice 22 weeks after CD34+ cell transplantation.
Mentions: We next evaluated if the overall improvement of human cell engraftment associated with group 7 protocol was observed in all lymphoid sites and specifically correlated with a better or faster differentiation of a given cell subset (i.e. CD19, CD3, CD4, CD8). We compared engraftment achieved in groups 1 and 7, in blood, spleen, lymph nodes and bone marrow at 12 and 22 weeks after transplantation of CB-CD34+ cells. At 22 weeks, high percentages of human CD45+ cells were observed in the lymph nodes (95.51±3.23%), bone marrow (83.41±2.65%) and spleen (74.31±2.92%) (n = 20) (Table 3). Humanized mice of group 7 also had higher absolute numbers of human cells in lymph nodes, bone marrow and spleen compared with group 1 (Fig. 2A). Twenty-four weeks after transplantation of fresh CB-CD34+ cells, lymph nodes of sizes from 3 to 7 mm could be observed in NSG mice from group 7 (Fig. 2B).

Bottom Line: In the present study, we further ameliorated the busulfan myeloablation regimen with fresh CB-CD34+cell transplantation in 3-4 week old NSG mice.Optimal engraftment promoted early and increased CD3+T cell levels, with better lymphoid tissue development and prolonged human cell chimerism over 300 days.We also saw a gradual decline of the CD4+T cell count, widespread immune activation, up-regulation of inflammation marker and microbial translocation after HIV-1 infection.

View Article: PubMed Central - PubMed

Affiliation: Immunology and infectious diseases unit GIGA-I3, University of Liege, Liege, Belgium.

ABSTRACT
Cord blood hematopoietic progenitor cells (CB-HPCs) transplanted immunodeficient NOD/LtsZ-scidIL2Rγ() (NSG) and NOD/SCID/IL2Rγ() (NOG) mice need efficient human cell engraftment for long-term HIV-1 replication studies. Total body irradiation (TBI) is a classical myeloablation regimen used to improve engraftment levels of human cells in these humanized mice. Some recent reports suggest the use of busulfan as a myeloablation regimen to transplant HPCs in neonatal and adult NSG mice. In the present study, we further ameliorated the busulfan myeloablation regimen with fresh CB-CD34+cell transplantation in 3-4 week old NSG mice. In this CB-CD34+transplanted NSG mice engraftment efficiency of human CD45+cell is over 90% in peripheral blood. Optimal engraftment promoted early and increased CD3+T cell levels, with better lymphoid tissue development and prolonged human cell chimerism over 300 days. These humanized NSG mice have shown long-lasting viremia after HIV-1JRCSF and HIV-1Bal inoculation through intravenous and rectal routes. We also saw a gradual decline of the CD4+T cell count, widespread immune activation, up-regulation of inflammation marker and microbial translocation after HIV-1 infection. Humanized NSG mice reconstituted according to our new protocol produced, moderate cellular and humoral immune responses to HIV-1 postinfection. We believe that NSG mice reconstituted according to our easy to use protocol will provide a better in vivo model for HIV-1 replication and anti-HIV-1 therapy trials.

Show MeSH
Related in: MedlinePlus