Development of NOG miceGeneral characteris5cs of NOG mice

1. T and B cell deﬁcient
2. NK cell deﬁcient
3. Reduced macrophage and dendri;c cell func;on
4. Complement ac;vity deﬁcient
5. No incidence of lymphoma
6. No T, B cell leakiness
7. Long life span
8. Sensi;ve for irradia;on
9. Sensi;ve against microbiological pathogens
10. High engraNment for xeno‐transplants

Basic characteris5cs ‐‐‐‐‐ Body weightBasic characteris5cs ‐‐‐‐‐ Organ weightfemalemale(g)
(g)

(g)

(g)

Histology of thymus from newborn miceNODNOD/SCIDNOG

Basic characteris5cs ‐‐‐‐‐ Irradia5on sensi5vity

Survival rates of NOG mice a7er irradia9on

Five mice in each group were irradiated with 2 to 4 Gy with using an X‐ray device(MBR‐1505R, Hitachi Medical Co., Tokyo) at age of 8 weeks.

Lymphocytes Leakiness in Immunodeﬁcient Mice
Mouse strain

CD19+

CD3+

No. of po;sive mice/ No. of mice*(range, µg/ml)
CD4+
CD8+

C.B‐17‐scid

6/10(0.5 ‐ 6.3)

1/10( 9.3)

1/10(6.5)

0/10‐

1/10(7.1)

1/10(2.1)

NOD‐scid

0/9‐

9/9(2.0 ‐ 13.8)

6/9(1.0 ‐ 12.5)

8/9(0.6 ‐ 7.8)

9/9(0.9‐12.4)

4/9(0.5 ‐ 1.1)

NOG

0/18‐

0/18‐

0/18‐

0/18‐

0/18
‐

0/18‐

IQI

4/4
(34.7 ‐ 59.9)

4/4(21.5 ‐ 34.9)

4/4(17.9 ‐ 25.5)

4/4(1.5 ‐ 3.5)

4/4
(20.0 ‐ 33.2)

4/4(0.7 ‐ 1.3)

TCRVb

TCRVgdMouse strain

Spleen test

Peripheral blood test

TCRVb

TCRVgd

CD19+

CD3+

No. of po;sive mice/ No. of mice*(range, µg/ml)
CD4+
CD8+

C.B‐17‐scid

4/10(0.5 ‐0. 8)

0/10
‐

0/10
‐

0/10
‐

0/10
‐

0/10
‐

NOD‐scid

0/9

2/9(2.4 ‐ 3.2)

2/9(1.53 ‐ 2.26)

2/9(0.8 ‐ 2.1)

4/9
(0.5 ‐ 4.4)

0/9

NOG

0/18
‐

0/18
‐

0/18
‐

0/18
‐

0/18
‐

0/18
‐

IQI

4/4
(28.1 ‐45.5)

4/4(14.4 ‐ 31.3)

4/4(10.4 ‐ 23.9)

4/4(2.6 ‐ 5.1)

4/4
(13.7 ‐ 30.0)

3/4(0.5 ‐ 1.4)

* No. of posi;ve mice means the number of mice with more than 0.5% of cells stained, since a value ofless than 0.5 % was considered as nonspeciﬁc staining.

No B cell leakiness in NOG mice

100%75%> 100 µg/mL
10 ~ 100 µg/mL

50%

1 ~ 10 µg/mL< 1 µg/mL

25%

0%

NOG
(n=71)

NOD-scid C.B-17-scid C57BL/6(n=50)(n=253)(n=5)

BALB/c(n=5)

NOG: 7-10 months oldNOD-scid: 6-7 months old
C.B-17-scid: 6-9 months old*
C57BL/6 & BALB/c: 12 weeks old*Ig M+G+A levels in sera of C.B-17- scid were measured in 1989.

Basic characteris5cs ‐‐‐‐‐ Hematology

Murine cells in bone marrow and spleen

Bone marrow and spleen were obtained from 12 weeks‐old NOG mice. Single cell suspensions prepared from them according
to the ordinal manner were stained with FITC‐ or PE‐labeled an;‐mouse CD45+, CD3+, CD4+, CD8+, CD19+, B220+, CD11b+,CD11c+, Gr‐1+ and analyzed with FACSCanto (BD sciences, CA).

Immunological characteris5cs ‐‐‐‐‐ NK cell defect

1.7%

25.2%

30.1%

9.3%

0.8%

NK cell Ac;vity of NOG mice
The NK cell ac;vity was determined by a cytotoxicity test using NK sensi;ve YAC‐1 cells as a target cell. Mice were intraperitoneally inoculated with
100 mg of polyinosinic‐polycy;dylic acid (poly I:C, SIGMA Chemical Co., St. Louis, MO) to s;mulate NK cell ac;vity for 48 hr before assay. Spleen cells
were separated from 4 mice of each strain of mice, pooled and co‐cultured with 51Cr‐labeld YAC‐1 cells as target cells for 4 hr at 37 oC in 5% CO2. The
supernatants harvested were assayed on a gamma counter. The present speciﬁc 51Cr release was calculated using the following formula, where X is themean experimental release from triplicate wells. Total release (T) was determined from wells with 51Cr labeled YAC‐1 cells and 1H HCl, and spontaneous
release (S) was determined from wells with 51Cr –labeled YAC‐1 cells and medium: % speciﬁc release = [(X‐S)/(T‐S)] x 100.

Immunological characteris5cs‐‐‐‐‐ Macrophage and Complement ac5vity

Defect of hemoly;c complement ac;vity in serum% 51Cr release

Reduced IL‐1 produc;on from macrophages

Stimulation of bone-marrow cells
IL‐1 produc;on from bone marrow cells
IL‐1 produc;on from bone marrow cells s;mulated with IFN‐g and LPS wasdetermined. Bone marrow cells were cultured with 500 U/ml human rM‐CSF, with and without 10 U/ml rat rIFN‐g and cultured for 4 days at 37 oC in
5% CO2. ANer 4 days, the medium was replaced with fresh medium alone orwith medium containing Escherichia coli LPS at 10 mg/ml. ANer anaddi;onal 24 hr incuba;on period, the culture supernatants were harvested
and assayed for IL‐1a levels using ELISA kits. The amount of IL‐1a in thesupernatants was expressed as the absorbance at 405 nm.

Serum dilution
Complement‐dependent hemoly;c ac;vityComplement‐dependent hemoly;c ac;vity in sera were determined bymeasurement of 51Cr released in the supernatants aNer 30 min incuba;onof mouse sera and 51Cr labeled SRBC/an; SRBC an;body conjugates.
Spontaneous release (S) was determined from wells with 51Cr SRBC‐Ab
conjugate in media. Total release (T) was determined from wells with 51Cr
SRBC‐Ab conjugates and 100 ml 2% SDS. Percent speciﬁc release = [(X‐S)/(T‐S)] x 100.

from Ito, M. et al. NOD/SCID/gamma(c)(null) mouse: an excellent recipientmouse model for engraNment of human cells. Blood 100, 3175‐3182. (2002).

No produc5on of IFNγ in NOG mice

In vitro cytokine produc;on of L. monocytogenes‐s;mulated
spleen cells from 3 strains of mice.
Spleen cells were separated aNer injec;on of 1 ml of 1 mg/ml
Collagenase D solu;on into the spleen. CD11c+ cells weredepleted from spleen cells from NOD/Shi‐scid mice treated withan;‐asialo GM1 an;serum, using an;‐CD11c an;body labeledmagne;c beads, by magne;c cell sorter (MACS). The cellsuspension were co‐cultured with 107 of heat‐killed L.
monocytogenes for 8 hours at 37 oC. The IFN‐g and IL‐6 levels inthe supernatants were determined using ELISA kits. Asterisk
indicates a signiﬁcant diﬀerence (* vs **: P<0.01).

Collabora5ve studies with Japanese Academic Society

2008.8.25/CIEA

GVHD/Others

Immunology

CancerOrgan/Tissue
Transplanta;onHematology
Infec;ousdiseases

Applica;on
1. Infec;ous disease model• HIV‐1 infec;on• ATL infec;on• EBV infec;on model• Hodgkin’s disease model
2. Cancer model• Liver metastasis• Mul;ple myeloma• Acute myeloid leukemia
3. Human ;ssue or organ model• Model with human ovary• Model with human liver• Model with human endometrium
4. Other models• GVHD model• Eﬃcacy test model for thrombopoie;c drugs• Safety test for human cell (ES, iPS, gene‐manipulated cells) transplanta;on