Effect of Jasinwhalhyul-tang on MRL/MpJ-Ipr/Ipr Mouse Model with Systemic Lupus Erythematosus

(1)

서 론

(systemic lupus erythematosus, SLE)

1,2)

. SLE

, ,

, , ,

, , , , ,

1-3)

. 20-40

3,4)

,

5)

.

1,6,7)

,

T, B

2007 1 17

2008 3 4

,

(Tel:+82-42-229-6880, Fax:+82-42-254-3403, E-mail:[email protected])

滋腎活血湯全身性紅斑性狼瘡影響

최훈섭, 조충식, 김철중 Original Article

Effect of Jasinwhalhyul-tang on MRL/MpJ-Ipr/Ipr Mouse Model with Systemic Lupus Erythematosus

Hoon-Seob Choi, Chung-Sik Cho, Cheol-Jung Kim

Department of Internal Medicine, College of Oriental Medicine, Daejeon University, Daejeon, Korea.

Objective : The main purpose of this study was to evaluate the effect of Jasinwhalhyul-tang (Zishenhuoxue-tang, JWT) on MRL/MpJ-Ipr/Ipr mouse model with systemic lupus erythematosus.

Methods: The effect of JWT on MRL/MpJ-Ipr/Ipr mice that have autoimmune disease similar to SLE in humans was evaluated after JWT per oral in the present study. Mice were administered with Jasinwhalhyul-tang (Zishenh- uoxue-tang, JWT) (80 or 400mg/kg) or distilled water for control group from experimental week 10 for 22 weeks.

Results : The amount of erythematosus skin lesion and proteinuria were significantly decreased. The size and weight of cervical lymph nodes and spleen were significantly reduced. The ratio between activated CD3

⁺

CD69

⁺

T-cells and undifferentiated CD3

⁺

CD4

^-

CD8

^-

T-cells in lymph nodes, spleen and kidney was effectively reduced. The gene expression of TGF-β in spleen and kidney was increased. The amount of anti-dsDNA IgG in blood was decreased.

The gene expression of TGF-β in normal mouse spleen cells was increased depending on concentration by treatment of with T cell stimulating agent. In the histological examination of skin and kidney, the amount of infiltration of immune cells involved in the inflammatory response was decreased.

Conclusions : According to the above results, JWT should be considered as an applicable therapeutic agent to SLE in clinical practice. Further research is required to investigate other efficacies of JWT on SLE.

Key Words : SLE, Jasinwhalhyul-tang ( Zishenhuoxue-tang , JWT), MRL/MpJ-Ipr/Ipr mouse model

(2)

1)

, T

8,9)

. SLE

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, , , ,

10,11)

. ,

, , , ,

, , ,

,

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.

12)

SLE 1 ,

13-15)

SLE , ,

.

16)

SLE

, ,

, , ,

, , , , , ,

SLE .

, SLE

MRL/MpJ-lpr/lpr

17-19)

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,

, cytokine,

, .

실 험

1.

재료 1)

MRL/MpJ-lpr/lpr 30g 7 8

SLC Inc.(Hamamatsu, Japan)

, ( )

22±2 1

.

20g 6 8 Balb/c (

)

. kg

(Table 1).

2)

diethyl pyrocarbonate(DEPC), 3-4,5- dimethyl-thiazol-2,5-carboxymethoxyphenyl-2,4-s ulfophenyl-2H-tetrazolim(MTS), Freund’s complete adjuvant, chloroform, collagenase, RPMI -1640

IMDM medium, isopropanol,

(RBC lysis solution), ethidium bromide(EtBr), Dulbecco’s phosphate buffered saline(D-PBS), calf thymus DNA, formaldehyde, lamide, mag- nesium chloride(MgCl

2

) Sigma (U.S.A),

(fetal bovine serum, FBS) Hyclone (U.S.A), anti-CD3-PE(phycoerythrin), anti- CD4- FITC(fluorescein isothiocyanate), anti-Gr1-PE, anti- CD8-FITC, anti-CD25-PE, anti-CD28-PE, anti-

조단백질 22.1%

조지방 8.0%

조섬유 5.0%

조회분 8.0%

칼슘 0.6%

인 0.4%

Table 1.

(3)

CD11b-FITC, anti-IgE-FITC anti-B220-PE, anti- CD69-FITC, goat anti-mouse IgG HRP conjugated, goat anti-TGFβ, propidium iodide(PI) RNase

Pharmingen (U.S.A) .

3)

( , Korea), rotary vaccum evaporator(Büchi B-480, Switzerland), freeze dryer(EYELA FDU-540, Japan), CO

2

incubator (Forma scientific Co., U.S.A), clean bench(Vision scientific Co., Korea), autoclave(Sanyo, Japan), micro-pipet(Gilson, France), water bath(Vision scientific Co., Korea), vortex mixer(Vision scientific Co., Korea), spectrophotometer(Shimazue, Japan), centrifuge (Sigma, U.S.A), deep-freezer(Sanyo, Japan), thermocycler system (MWG Biotech., Germany), ice-maker(Vision scientific Co., Korea), homogenizer(OMNI, U.S.A), plate shaker(Lab-Line, U.S.A) ELISA reader(Molecular Devices, U.S.A)

.

4)

, 1

(Table 2).

2.

방법 1)

1 5,000

2 rotary vacuum evaporator .

freeze dryer 1

25g .

.

MRL/MpJ-lpr/lpr 10

( )

80mg/kg (JWT- ), 400mg/kg

(JWT- ) 3 . 10

22 13

0.2ml

韓藥名生藥名用量(g)

生地黃 Rehmanniae Radix 7.50

牧丹皮 Moutan Cortex 7.50

赤芍藥 Paeonia Radix Rubra 7.50

桃仁 Persicae Semen 15.00

白花蛇舌草 Oldenlndiae Diffusae Herba 15.00

半枝蓮 Scutellariae Barbatae Herba 15.00

白朮 Atractylodis Macrocephalae Rhizoma 6.00

茯 Poria cocos 6.00

山藥 Dioscorea Rhizoma 7.50

Smilax Chinae Rhizoma 15.00

薏苡仁 Coicic Semen 15.00

丹參 Salvia Miltriorrhizae Radix 7.50

黨參 Codonopsis Pilosulae Radix 7.50

Total amount 132.00

Table 2.

(4)

, . 2)

MRL/MpJ-lpr/lpr JWT

10 22

. 3)

MRL/MpJ-lpr/lpr SLE

. ,

20)

1 .

0 :

1 : 1 0.5cm

2 : 2 0.5cm

3 : 2 0.5cm

4)

enzyme- linked immuno- sorbent assay(ELISA)

.

96well-ELISA plate 0.1M carbonate buffer goat anti- murine albumin(Bethyl, U.S.A.)

24 , 0.1%

Bovine serum albumin(BSA)

(PBS) 2 blocking

plate . plate well

2000 20,000 100

2 3

, 50,000 anti-murine

albumin/HRP conjugated(Bethyl, U.S.A.) 100 2

. well 5 , tetram-

ethylbenzidine(TMB) 100

15 30 1N

, ELISA reader

450nm .

0 500ng/ml murine

albumin .

4

20,21)

. grade 0 : 0 0.3mg/ml grade 1 : 0.3 1mg/ml grade 2 : 1 3mg/ml grade 3 : 3 20mg/ml

grade 4 : 20mg/ml , death

5) , ,

10 22 MRL

/MpJ-lpr/lpr ethyl ether , ,

.

6) , ,

MRL/MpJ-lpr/lpr Normal

Balb/c spleen, lymph node, kidney

100mesh (kidney

collagenase 30 2 )

D-PBS 5 (1700rpm) 2

cell strainer (FALCON)

. ACK (8.3g NH4Cl, 1g

KHCO3, in 1L of demineralized water+0.1mM

EDTA) 5

D-PBS 2 0.04%

trypan blue .

(5)

2×105 4 (immuno-fluorescence staining) . anti-CD3-PE(phycoerythrin), anti-CD4-FITC(fluorescein isothiocyanate), anti-Gr1 -PE, anti-CD8-FITC, anti-CD25-PE, anti-CD28- PE, anti-CD11b-FITC, anti-IgE-FITC anti-B220- PE, anti-CD69-FITC, fluorescein isothio- cyanate(FITC)-anti-mouse CD19 30

. 3

(flow cytometer, Becton Dickinson, U.S.A) MRL

/MpJ-lpr/lpr ,

CD3e, CD4, CD8, CD25, CD69,

CD19 .

CellQuest CD3

⁺

, CD19

⁺

, CD4

⁺

, CD8

⁺

, CD4

⁺

CD25

⁺

, CD3

⁺

CD69

⁺

, CD3

⁺

CD4

^-

CD8

^-

(gated, %) .

7) (1) RNA MRL/MpJ-lpr/lpr

RNAzolB .

chloroform(CHCl3) 15

. 15

13,000rpm 200

2-propanol 200

15 .

13,000rpm 80% EtOH

3 vaccum pump

RNA . RNA DEPC

20 heating block 75

first strand cDNA .

(2) -

(reverse transcription) total

RNA 3 75 5 ,

2.5 10mM dNTPs mix, 1 random sequence hexanucleotides(25pmole/25 ), RNA inhibitor

1 RNase inhibitor(20U/ ), 1 100mM DTT, 4.5 5×RT buffer (250mM Tris-HCl, pH 8.3, 375mM KCl, 15mM MgCl

2

) , 1

M-MLV RT(200U/ ) DEPC

20 . 20 2,000

rpm 5 37

60 first-strand cDNA

, 95 5 M-MLV

RT cDNA

polymerase chain reaction(PCR) . PCR iCycler(Biorad, U.S.A)

. 3 cDNA

, primer L32,

Foxp3, TGF-β, IL-10, IL-2, γ-IFN, IL-12, GITR,

CTLA-4, CD25

sense primer(20pmole/ ) antisense primer

(20pmole/ ) 1 , 3

2.5mM dNTPs, 3 10×PCR buffer(100mM Tris-HCl, pH 8.3, 500 mM KCl, 15mM MgCl

2

), 1X SyberGreen, 0.18 Taq polymerase

(5U/ ) 30

pre-denaturation; 95 , 5 , denaturation; 95 , 30 , annealing; 58 , 30 , elongation; 72 , 30 40cycles post-

elongation 72 3 PCR

. Real-time PCR iCycler

.

8) IgG Anti-dsDNA

cytokine ELISA

.

(6)

96well-ELISA plate 0.1M carbonate buffer goat anti- mouse IgG (total IgG ) calf thymus DNA (anti- dsDNA )

24 , 0.1%

BSA (Bovine serum albumin) PBS

2 blocking plate

. plate well 100 100,000 100

2 3 ,

1,000 100,000 anti-mouse IgG/HRP conjugated 100

2 . well 5

, TMB (tetramethyl-benzidine)

100 15 30

1N , ELISA

reader 450nm

. Total IgG 0

1,000ng/ml murine IgG

. Anti-dsDNA

. 9)

T-

, 2×106/ml

Balb/c PMA

(phorbol myristate acetate; 10ng/ml) IoM(iono-

mycin; 1μM) T-

JWT . 1

13,000rpm Realtime RT-

PCR .

10)

MRL/MpJ-lpr/lpr

10% formalin-

. 4-μm

, H&E . 11)

± , Student’s T-test

²²⁾

p<0.05 .

성 적

1.

체중에 미치는 영향

10 22

(Table 3) JWT

, JWT .

Group Mean Body Weight (g)

10 week 22 week Δ(22week∼ 10week)

Control 31.4 ± 2.1 41.2 ± 2.4 9.6 ± 2.7

JWT- Ⅰ 31.5 ± 1.6 40.9 ± 2.7 9.6 ± 2.9

JWT- Ⅱ 30.8 ± 2.6 40.0 ± 2.7 9.2 ± 3.6

The MRL/MpJ-lpr/lpr mice were administered with JWT (JWT- Ⅰ, 80mg/kg, p.o and JWT-Ⅱ, 400mg/kg, p.o) for 13weeks daily. Control group mice were administered with D.W. instead of JWT. Statistically significant differences in mean body weight of mice were not observed between the groups (n=10).

Table 3.

(7)

2.

홍반에 미치는 영향

2 2.1±0.9

, JWT

, 0.5cm

1 .

JWT- 0.8±0.5 (p<0.05), JWT- 0.2±0.4(p<0.05)

(Fig. 1, 2).

3.

요단백에 미치는 영향

(100mg/ , grade 2

Fig. 1.

4

3

2

1

0

Clinical score

Control JWT-I JWT-II

*

100%

90%

80%

70%

60%

50%

40%

30%

20%

10%

0%

0 1 2 3 4

Fig. 2. Fig. 3.

(8)

) 60%

, JWT- 40%

. JWT- 20% (p<0.05) (Fig. 3).

4.

비장

,

경부림프절

,

신장의 중량 및 크기에 미 치는 영향

JWT- , JWT-

(Fig. 4, 6).

, JWT- 0.64±0.31g(p<0.05) (Fig. 5, 7).

JWT- , JWT- (Fig. 8).

1.4

1.2 1.0

0.8

0.6

0.4

0.2

0.0

g

2.0 1.8 1.6 1.4 1.2 1.0 0.8 0.6 0.4 0.2 0.0

g *

Fig. 6. Fig. 7.

Fig. 4. Fig. 5.

(9)

5.

비장

,

경부림프절

,

신장에서의 세포 조성비에 미치는 영향

, JWT-

CD4

⁺

CD25

⁺

(p<0.05) , JWT- CD3

⁺

, CD3

⁺

CD69

⁺

, CD3

⁺

CD4

^-

CD8

^-

(p<0.05) ,

CD4

⁺

CD25

⁺

(p<0.05)

(Table 4).

, JWT-

CD3

⁺

CD69

⁺

, CD4

⁺

, CD8

⁺

, JWT- CD3

⁺

CD69

⁺

, CD4

⁺

, CD8

⁺

, CD4

⁺

CD25

⁺

, CD3

⁺

CD4

^-

CD8

^-

(p<0.05)

(Table 5).

, JWT-

0.8

0.7

0.6 0.5

0.4 0.3 0.2

0.1

0.0

g

14

12

10

8

6

4

2

0

(mg/ml)

Fig. 8. Fig. 9.

Leucocytes

type Organ Normal

Balb/c(%)

MRL-lpr lupus model mice(%)

Control JWT- Ⅰ JWT- Ⅱ

CD3

⁺

(T) LN# 73.0±0.85 94.6±1.20 93.7±.0.58 91.8±1.30

^*

CD19

⁺

(B) LN 25.6±0.90 1.7±0.12 1.7±0.86 4.0±0.48

CD3

⁺

CD69

⁺

LN 4.1±0.29 35.2±1.35 28.1±4.15 14.9±2.06

^*

CD4

⁺

LN 66.2±2.00 9.6±2.15 8.7±2.05 7.6±0.65

CD8

⁺

LN 17.7±0.30 2.0±0.20 2.0±0.30 2.0±0.45

CD4

⁺

CD25

⁺

LN 5.0±0.44 2.1±0.15 2.6±0.06

^*

2.6±0.07

^*

CD3e

⁺

CD4

^-

CD8

^-

LN 3.6±0.45 76.1±2.25 73.2±0.20 61.0±2.45

^*

#LN : Lymphnode

The MRL/MpJ-lpr/lpr mice were administered with JWT (JWT- Ⅰ, 80mg/kg, p.o and JWT-Ⅱ, 400mg/kg, p.o) for 13weeks daily.

Results are mean ± SD (n=5 mice per group). *p<0.05 versus control group.

Table 4.

(10)

CD3

⁺

, CD4

⁺

, CD4

⁺

CD25

⁺

, CD3

⁺

CD4

^-

CD8

^-

, JWT- CD3

⁺

, CD3

⁺

CD69

⁺

, CD4

⁺

, CD8

⁺

, CD4

⁺

CD25

⁺

, CD3

⁺

CD4

^-

CD8

^-

(p<0.05) (Table 6).

6.

비장 및 신장에서의 유전자 발현에 미치는 영향

L32 , TNF-α,

TGF-β, IL-10, IL-2, IL-12, IL-13, γ-IFN

cytokines Foxp3, CD25, CTLA-4, GITR regulatory T-cell

, JWT

, TGF-β JWT

(p<0.05) (Table 7).

Leucocytes

type Organ Normal

Balb/c(%)

MRL-lpr lupus model mice(%)

Control JWT- Ⅰ JWT- Ⅱ

CD3

⁺

(T) SP

^#

49.6±5.10 86.4±2.45 84.9±1.25 84.4±3.10

CD19

⁺

(B) SP 40.9±5.75 9.6±1.20 12.2±0.15 12.4±2.35

CD3

⁺

CD69

⁺

SP 2.9±0.60 18.4±0.40 12.6±2.50

^*

13.5±0.60

^*

CD4

⁺

SP 25.5±3.75 12.2±0.25 9.2±0.50

^*

11.2±0.05

^*

CD8

⁺

SP 13.6±2.35 7.4±0.10 4.6±0.35

^*

4.9±0.65

^*

CD4

⁺

CD25

⁺

SP 2.6±0.26 2.3±0.17 1.9±0.37 1.4±0.20

^*

CD3e

⁺

CD4

^-

CD8

^-

SP 3.7±2.06 69.1±0.70 65.5±1.85 61.8±1.00

^*

#SP : Spleen B/T Rate 0.824 0.111 0.144 0.147

The MRL/MpJ-lpr/lpr mice were administered with JWT (JWT- Ⅰ, 80mg/kg, p.o and JWT-Ⅱ, 400mg/kg, p.o) for 13weeks daily.

Results are mean ± SD (n=5 mice per group). *p<0.05 versus control group.

Table 5.

Leucocytes

type Organ Normal

Balb/c(%)

MRL-lpr lupus model mice(%)

Control JWT- Ⅰ JWT- Ⅱ

CD3

⁺

(T) KD

^#

20.8±2.15 91.3±2.10 83.6±0.80

^*

82.7±2.55

^*

CD19

⁺

(B) KD 3.0±1.15 0.4±0.09 1.2±0.45 1.4±0.26

CD3

⁺

CD69

⁺

KD 6.0±0.44 57.7±1.30 56.9±1.55 52.5±2.20

^*

CD4

⁺

KD 6.7±0.43 67.9±0.60 56.5±2.25

^*

57.8±1.30

^*

CD8

⁺

KD 5.1±1.65 20.5±0.20 22.2±0.65 13.6±1.15

^*

CD4

⁺

CD25

⁺

KD 0.8±0.10 1.5±0.43 0.50±0.04

^*

0.9±0.14

^*

CD11b

⁺

Gr-1

⁺

KD 0.42±0.02 1.24±0.14 1.16±0.39 0.62±0.27

CD3e

⁺

CD4

^-

CD8

^-

KD 0.1±0.01 3.6±0.64 1.0±0.07

^*

1.6±0.19

^*

#KD : Kidney B/T Rate 0.144 0.004 0.014 0.017

The MRL/MpJ-lpr/lpr mice were administered with JWT (JWT- Ⅰ, 80mg/kg, p.o and JWT-Ⅱ, 400mg/kg, p.o) for 13weeks daily.

Results are mean ± SD (n=5 mice per group). *p<0.05 versus control group.

Table 6.

(11)

7.

혈액내

IgG

및

Anti-dsDNA

정량 결과 IgG

(Fig. 9), Anti-dsDNA IgG JWT- (p<0.05) (Fig. 10).

8.

세포의 유전자 발현에 미치는 영향

TGF-β , L32

TNF-α, IL-10, IL-2, IL-12, IL-13, γ-IFN

cytokines Foxp3, CD25, CTLA-4, GITR regulatory T-cell

, TGF-β

, JWT

(p<0.05) (Fig. 11).

9.

피부 및 신장의 조직학적 소견

JWT

Genes Control JWT-I JWT-II

(Spleen)

TNF- α 0.15 ± 0.07 0.20 ± 0.05 0.21 ± 0.06

TGF- β 0.46 ± 0.09 0.76 ± 0.06

^*

0.66 ± 0.04

^*

IL-10 0.26 ± 0.15 0.31 ± 0.06 0.36 ± 0.10

IL-2 2.12 ± 0.55 2.53 ± 0.62 1.95 ± 0.33

IL-12 0.02 ± 0.01 0.02 ± 0.02 0.02 ± 0.01

IL-13 1.90 ± 0.49 2.24 ± 0.33 2.60 ± 0.93

γIFN 0.10 ± 0.03 0.15 ± 0.07 0.12 ± 0.03

Foxp3 0.21 ± 0.07 0.28 ± 0.10 0.27 ± 0.14

CD25 0.30 ± 0.23 0.29 ± 0.05 0.38 ± 0.06

CTLA-4 0.61 ± 0.08 0.59 ± 0.05 0.69 ± 0.11

GITR 0.02 ± 0.01 0.03 ± 0.02 0.05 ± 0.02

(Kidney)

TNF- α 0.74 ± 0.02 0.84 ± 0.24 0.68 ± 0.07

TGF- β 2.23 ± 052 6.19 ± 0.64

^*

3.56 ± 0.37

^*

IL-10 0.38 ± 0.19 0.37 ± 0.03 0.39 ± 0.08

IL-2 0.49 ± 0.05 0.58 ± 0.15 0.60 ± 0.11

IL-12 0.06 ± 0.02 0.05 ± 0.04 0.03 ± 0.03

IL-13 1.59 ± 0.16 2.46 ± 0.62 1.51 ± 0.12

γIFN 0.32 ± 0.05 0.24 ± 0.07 0.29 ± 0.06

Foxp3 0.21 ± 0.17 0.40 ± 0.17 0.39 ± 0.18

CD25 0.10 ± 0.04 0.12 ± 0.05 0.09 ± 0.05

CTLA-4 0.16 ± 0.07 0.19 ± 0.04 0.14 ± 0.06

GITR 0.05 ± 0.02 0.08 ± 0.03 0.03 ± 0.02

The MRL/MpJ-lpr/lpr mice were administered with JWT (JWT-I, 80mg/kg, p.o and JWT-II, 400mg/kg, p.o) for 13weeks daily. Gene expression levels were analyzed by real-time RT-PCR method as described in Methods section. Results are mean ± SD (n=5 mice per group). *p<0.05 versus control group. Results mean normalized fold increase in specific mRNA levels, that is expression levels of specific genes normalized against those of L32 gene.

Table 7.

(12)

,

(Fig. 12).

, JWT glomeruli

(Fig. 13).

고 찰

SLE

1,23)

, , , ,

4,23)

,

, , ,

6)

. SLE

, ,

1,6,15)

,

1,4,6,15)

. SLE

(ANA)

, DNA, , RNA

ANA

25)

. ANA

, ANA

*

10 9 8 7 6 5 4 3 2 1 0

Ratio to normal level

Normal

*

Control JWT-5 JWT-50

80 90

70 60 50 40 30 20 10 0

norm alized fold increase in TGF-mRNA levels

*

Fig. 10. Fig. 11. β

μ μ

β

μ

(13)

(Immune Complexes)

SLE

⁶⁾

.

,

, , , , ,

,

, , ,

ANA

²⁵⁾

.

SLE 1982

(American Rheumatism Association, ARA)

11 4

SLE

^1,3,24-27)

.

, DNA , LE

, , γ-globulin ,

, Extractable

, Reumatic Factor ,

,

, FDP , albumin ,

ESR CRP

^25,28)

.

SLE

,

7,29)

.

30)

,

31)

.

,

Fig. 12. Fig. 13.

(14)

,

32,33)

.

34)

,

SLE

³⁵⁾

.

SLE

35 90%

^24,36)

, ARA

0.5gm 3

⁺

( ,

, , )

3,12,24)

.

SLE 2/3 ,

globulin

3,12,24,36)

.

SLE , , ,

, , , ,

10-12)

, SLE

, ,

^11,29)

.

SLE

, , , ,

, ,

^10-12,37)

. , ,

, , , , ,

11-12)

,

, , ,

37)

.

,

, SLE

, , ,

, , , , ,

12)

.

MRL/MpJ-lpr/lpr SLE

, fas

³⁸⁾

, fas apoptosis

38)

. Fas

SLE

³⁸⁾

.

T ,

. 3

75 ,

anti-dsDNA

. 22

17-19)

. SLE

MRL/MpJ-lpr/lpr

^17-19)

. , JWT

, JWT

.

JWT- , JWT-

(Fig. 1, 2).

. , (100mg/dL, grade 2 )

60% , JWT-

40%, JWT- 20%

(Fig. 3).

.

(15)

(Fig. 4, 5), JWT

, JWT-

.

JWT ,

,

JWT- CD3

⁺

, CD3

⁺

CD69

⁺

, CD3

⁺

CD4

^-

CD8

^-

, CD4

⁺

CD25

⁺

, JWT- CD4

⁺

CD25

⁺

(Table 4), JWT- CD3

⁺

CD69

⁺

, CD4

⁺

, CD8

⁺

, JWT-

CD3

⁺

CD69

⁺

, CD4

⁺

, CD8

⁺

, CD4

⁺

CD25

⁺

, CD3

⁺

CD4

^-

CD8

^-

(Table 5),

JWT- CD3

⁺

, CD4

⁺

, CD4

⁺

CD25

⁺

, CD3

⁺

CD4

^-

CD8

^-

, JWT- CD3

⁺

, CD4

⁺

, CD4

⁺

CD25

⁺

, CD3

⁺

CD4

^-

CD8

^-

, CD3

⁺

CD69

⁺

, CD8

⁺

(Table 6). CD4

⁺

CD25

⁺

regulatory T-

, CD

⁺

CD69

⁺

, CD3

⁺

CD4

^-

CD8

^-

SLE T-

T- , JWT-

. T- ,

. JWT

, L32 , TNF-

α, TGF-β, IL-10, IL-2, IL-12, IL-13, γ-IFN cytokines Foxp3, CD25, CTLA-4, GITR

regulatory T-cell

JWT

, TGF-β

JWT . TGF-

β pleiotropic cytokine

,

³⁹⁾

.

TGF-β ,

,

DNA chip analysis

. TGF

-β

⁴⁰⁾

,

. ,

IgG , SLE

anti-dsDNA IgG

⁴¹⁾

JWT-

. FACS

CD3

⁺

CD4

^-

CD8

^-

T- CD3

⁺

CD69

⁺

T- ,

MRL/MpJ-lpr/lpr

20,21)

. TGF-β

, JWT

. TGF-β

, TGF-β

membrane-bound TGF-β

41)

. JWT MRL/MpJ-lpr/lpr

TGF-β in vitro

, L32

(16)

TNF-α, IL-10, IL-2, IL-12, IL-13, γ -IFN cytokines Foxp3, CD25, CTLA-4, GITR regulatory T-cell

. , TGF-β JWT

PMA/ionomycin

50 μg/ml 2 .

T-

.

, JWT

,

.

. ,

, JWT glomeruli

.

MRL/MpJ-

lpr/lpr SLE ,

, , CD3

⁺

CD4

^-

CD8

^-

T- CD3

⁺

CD69

⁺

T- , anti-dsDNA IgG,

, TGF-β , SLE

, .

결 론

MRL/MpJ-lpr/lpr

1. .

2. .

3. , , CD3

⁺

CD69

⁺

T- CD3

⁺

CD4

^-

CD8

^-

T-

.

4. TGF-β

.

5. anti-dsDNA IgG

.

6. TGF-β

. 7.

. SLE

, .

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