The Effects of N-acetyl-seryl-aspartyl-lysyl-proline on Expression of NF-κb and MCP-1 in Rats with Silicosis

doi:10.4236/ijcce.2013.22B006

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International Journal of Clean Coal and Energy, 2013, 2, 24-26

doi:10.4236/ijcce.2013.22B006 Published Online May 2013 (http://www.scirp.org/journal/ijcce)

The Effects of N-acetyl-seryl-aspartyl-lysyl-proline on

Expression of NF-κb and MCP-1 in Rats with Silicosis

Qian Li, Fang Yang*, Lijuan Zhang, Hong Xu, Wenli Zhang

Medical research center Hebei United University, Thangshan, China

Email: [email protected]

Received 2013

ABSTRACT

In the present study, we developed silicosis of rat model by bronchial perfusion SiO2 dust, and intervenes with AcSDKP,

immunohisto chemistry was used to detect NF-κb and MCP-1 expression in lung tissue, and positive cells were counted.

We found that compared with silicotic model group, the positive cells of NF-κb and MCP-1 were decreased signifi-

cantly in anti-fibrosis treatment of AcSDKP group. The findings suggest that AcSDKP could inhibit the expression of

NF-κb and MCP-1 in lung tissue of silicosos, this may be related to AcSDKP inhibit of macrophage infiltration in lung

tissue and reduced the degree of dust alveolitis.

Keywords: N-acetyl-seryl-aspartyl-lysyl-proline; Silicosis; Monocyte Chemotactic Protein-1; Nuclear Factor-κb

1. Introduction

The main pathological change of silicosis is the fibro-

blast proliferation and collagen deposition in lung tissue.

Recent studies have shown that cytokines play an impor-

tant role in the development of silicosis, they constitute a

complex signal transduction molecules network system

and the chain reaction system, leading to infiltration of

inflammatory cells, fibroblasts and mesenchymal con-

nective tissue deposition is characterized by a chronic

immune-mediated inflammation[1]. Rat silicosis model

adopted by the bronchial perfusion SiO2 dust, and inter-

venes with AcSDKP, study the therapeutic effects of

AcSDKP on silicotic fibrosis, provides new method for

silicosis preventing and controlling.

2. Materials and Methods

2.1. Ethics Statement

Male Wistar rats, weighting 180 ± 10 g, were purchased

from Vital River Laboratory Animal Technology Co. Ltd.

(Beijing, China). All animal experiments were reviewed

and approved by the Institutional Animal Care and Use

Committee at the Hebei United University. Animals were

given free access to food and water and were cared for

according to guidelines set by the National Institute of

Health (NIH)

2.2. Induction of Silicosis and AcSDKP

Treatment

Rats were anesthetized with Isoflurane and then received

either silica solution (50 mg/rat, 1ml) or 0.9% saline (as a

vehicle control) by trachea. Prior to instillation the 5 μm

silica particles (Sigma , St. Louis, MO, USA) were baked

at 180℃ for 6 hours. AcSDKP [800 μg/(kg d), Bachem

AG company, USA] or control (0.9% saline) was given

via a miniosmotic pump (Alzer 2 ml4, DURECT Co. Ltd,

USA) planted into the abdominal cavity. Rats were di-

vided into 3 groups (n = 10), 1) control group (instilled

with 0.9% saline, and then treated with 0.9% saline for 4

w) ; 2) silicotic model group (instilled of SiO2,, and then

treated with 0.9% saline for 4 w); 3) anti-fibrosis treat-

ment of AcSDKP group (instilled with SiO2, and treated

with AcSDKP for 4 w) [2].

2.3. Immunohistochemistry for NF-κb and

MCP-1

Take the lungs to organize 4% paraformaldehyde to be

fixed, conventional paraffin wax embedding, 6 μm serial

section, for immunohistochemical method dyeing. After

incubation with 5% horse serum, sections were incubated

with primary antibodies against NF-κb and MCP-1 (Santa

cruz Biotechnology, USA) followed by the biotinylated

secondary antibody and finally the ABC reagent (Wuhan

Boshide Biological Engineering Co. Ltd, China). Immu-

noreactivity was visualized with DAB. A brown color

staining was considered a positive result. Counts the

*Corresponding author.

Q. LI ET AL. 25

positive cells of NF-κb and MCP-1.

3. Results

3.1. Morphological Observation

Surfaces of lung are smooth and no nodule formation in

control group. Canous nodules distribute sporadically on

the surface and cutting surface in silicotic model group.

Compared with silicotic mode group, the numbers of

canous nodules decreased on the surface and cutting sur-

face in the groups with AcSDKP treatment.

3.2. Expression of NF-κb and MCP-1 in Lung of

Rat with Silicosis

Immunohistochemistry staining shows that NF-κb and

MCP-1 only express in a small number of alveolar

macrophages and fibroblasts in control group (Figure 1).

NF-κb and MCP-1 expressions increase in silicotic nod-

ules and interstitial fibrotic area. The expressions of

NF-κb and MCP-1 in AcSDKP treatment are less than

that in silicotic model. Positive cells counting shows that

NF-κb and MCP-1 expressions in silicotic model group

are increases by 2.76 fold and 5.40 fold compared with

control group. AcSDKP treatment decreases the expres-

sions of NF-κb and MCP-1. The positive cells counting

of NF-κb and MCP-1 in anti-fibrotic treatment group is

70.12% and 65.37% of silicotic model group respectively.

Variance analysis reveals that differences are significant

(p < 0. 05).

a b c

d e f

control silicotic

model

AcSDKP

anti-fibrotic

ositive cells

NF-κb

MCP-1

(a) control group for NF-κb; (b) silicotic model group for NF-κb; (c)

AcSDKP anti-fibrotic group for NF-κb; (d) control group for MCP-1; (e)

silicotic model group for MCP-1; (f) AcSDKP anti-fibrotic group for MCP-1.

(A) The positive cells expression of NF-κb and MCP-1 in the lung of rat. *p

＜0.05 vs contro; #p ＜ 0.05 vs silicotic model.

Figure 1. Effect of Ac-SDKP on NF-κb and MCP-1 analyzed

by Immunohistochemistry in rats with silicosis. (× 400).

4. Discussion

N-acetyl-seryl-aspartyl-lysyl-proline(AcSDKP) is a the

physiological hematopoietic growth inhibition factor,

studies have shown that AcSDKP inhibit proliferation

and collagen synthesis of cardiac fibroblasts [3,4]. In

recent years NF-κB prepared in the silicosis fibrosis

pathogenesis research is paid attention, NF-κb is a tran-

scription activation function of proteins, with the wide

range of promoter or Enhancer combining parts of κb site

specificity and promote their transcription, NF-κB in-

volved in many of the former transcriptional regulation

of inflammatory mediators molecules promoting cyto-

kines, adhesion molecules, chemokines, inflammatory

factors, oxidative stress-related enzymes released in large

quantities, so as to promote organ inflammation and fi-

brosis[5]. MCP-1 is a chemokine CC subfamily, its main

function is the monocyte-macrophage cell chemotaxis, in

addition, MCP-1 except took one kind of front inflam-

mation cell factor participates in the adjustment white

blood cell outside the chemotaxy, it also has stimulates

the ciliary cell to multiply, the collagen synthesizes as

well as induces certain presses the function which the

fibrosis cell factor or the medium produce[6]. Our studies

found that positive cells of NF-κb and MCP-1 expres-

sions increase in silicotic model group compared with

control group, it is shown that in the stimulation of SiO2

dust, the production and expression of NF-κb and MCP-1

increased in silicosis lungs of rats. More important dis-

coveries found that in silicosis lungs of rats, the positive

cells of NF-κb and MCP-1 were significantly decreased

after given AcSDKP intervention. This indicated that,

AcSDKP can suppress the production and expression of

NF-κb and MCP-1 in the lung with the SiO2 dust stimu-

lation, reduced the inflammatory response in the lungs of

silicotic rats.

5. Conclusions

Our data indicate that AcSDKP possibly through inhibi-

tion of NF-κb and MCP-1 production and expression,

reduce the synthesis and release of pro-fibrotic cytokines,

thereby inhibiting or reducing extracellular matrix syn-

thesis and deposition of the lungs, reducing the extent of

silicosis fibrosis and inhibiting the progression of fibro-

sis.

6. Acknowledgements

This work was funded by National Natural Science

Foundation of China (No. 81072254) and Hebei Province

Natural Science Foundation of China (No. C 20114010

24).

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