JP2003038063A - Dermatitis-developed rat - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a dermatitis-developed rat serving as a pathological model for allergic dermatitis or the like and thus contributing to efficiently screening prophylactic and/or therapeutic agents for the dermatitis in question and carrying out their medicinal efficacy tests. SOLUTION: This dermatitis-developed rat is a hairless rat which lacks the ability to grow normal hair, having the following characteristics: (a) naturally developing slight skin erythema about 8 to 12 weeks after born, (b) naturally developing dermatitis at a rate of >=90% 24 weeks after born and thereafter, and (c) varying blood IgE antibody levels little. The 2nd objective method for creating the rat, and the other objective method for screening compounds effective for preventing and/or treating dermatitis by using the rat, are also provided respectively.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a hairless rat that spontaneously develops dermatitis.

[0002]

2. Description of the Related Art Generally, it is important to develop disease model animals to elucidate the mechanism of disease development and to develop therapeutic drugs and treatment methods, and various pathological model animals have been prepared.

As a model animal for skin diseases, PCA (passive cutaneou), which is a model for allergic dermatitis, is used.
s anaphylaxis) reaction in addition to mice, rats, rabbits, guinea pigs, etc., recently, animals that spontaneously develop dermatitis have been found, for example, Nc / Nga mice that develop atopic dermatitis (Int. Immunol., 9: 461,199
7), hairless mice that develop wet and dry skin lesions (WO98 / 05202) have been reported.

[0004] However, there is an inconvenience that treatment such as shearing and antigen administration must be performed in order to cause the PCA reaction in the animal, and in the spontaneously-occurring animal, the incidence rate, the site of onset and the inflammatory area are not limited. The current situation is that the points are not yet sufficient.

[0005]

SUMMARY OF THE INVENTION It is an object of the present invention to provide a disease state model animal that spontaneously develops dermatitis at a high incidence and enables efficient drug evaluation and the like.

[0006]

[Means for Solving the Problems] The inventors of the present invention have conducted extensive studies on hairless rats to solve the above-mentioned problems. We succeeded in establishing dermatitis-prone rats that spontaneously develop dermatitis at a high rate and in which the IgE antibody level in blood hardly fluctuates, and completed the present invention.

That is, the present invention is a hairless rat lacking the ability to grow general hair, and has the following characteristics: (a) Spontaneous skin erythema at about 8 to 12 weeks of age, (b)
Provided are a dermatitis-producing rat and a method for producing the dermatitis-producing rat, which spontaneously develops dermatitis with an incidence of 90% or more after 24 weeks of age, and (c) the IgE antibody level in blood hardly fluctuates. To do.

The present invention also provides a method for screening a compound effective for the prevention and / or treatment of dermatitis using the dermatitis-producing rat.

[0009]

BEST MODE FOR CARRYING OUT THE INVENTION The characteristics of the dermatitis-affected rats of the present invention (hereinafter, also referred to as "KSDR" Kowa Spontaneous Dermatitis Rats) are shown below. 1) Alopecia and dermatitis traits are transmitted to offspring by an autosomal dominant gene. 2) Although there are male and female differences and individual differences in the onset time of skin erythema, it is around 8 to 12 weeks of age (average male: 10 weeks old, female: 9 weeks old).
To mild erythema on the skin. As for the initial lesions, scratches-like erythema was observed in many rats in one to several places on the lower back (Fig. 1). Dermatitis progresses with age,
It expands to the entire back area at about 0 weeks of age. Around this time, some crusts also fell off (Fig. 2). When the skin area up to 21 weeks of age was measured for an individual who developed dermatitis at 15 weeks of age, the dermatitis area increased by about 30% in 2 weeks (Table 1). The incidence of dermatitis at 24 weeks of age was 90.
% Or more (male: about 94%, female: about 100%), and in females, onset occurs earlier and lesions tend to progress earlier than in males. In addition, the incidence is the breeding environment (barrier, open)
And is not affected by feed.

3) In the histopathological findings, the epidermis at the normal site of KSDR shows hyperplasia as compared with hairy rats. In the skin inflammation area, hyperplasia of the epidermis accompanied by hyperkeratosis is remarkable (Fig. 3). Furthermore, infiltration of eosinophils and mast cells is observed in the dermis (Fig. 4).

4) The number of peripheral blood leukocytes increases with the progress of dermatitis, and tends to increase as compared with normal (hairy) rats (Table 3). Further, the dermatitis area and the white blood cell count are correlated (Table 4). In addition, lymphocytes decrease and neutrophils (segmental nuclei) and eosinophils increase with the progress of dermatitis (Table 5).

5) The IgE antibody level in blood hardly changed even after the onset of dermatitis, and the IgE antibody level in blood at 15 weeks of age was about 65 ng / mL or less (Table 7).

6) The onset dermatitis is cured by steroid drugs (dexamethasone, prednisolone valerate acetate, etc.), but an immunosuppressant (tacrolimus, etc.) cures it.
It is not cured by antiseptic agents (chlorhexidine gluconate, povidone iodine, etc.), antibiotics (penicillin G, etc.), and antiallergic agents (ketotifen fumarate, etc.) (Table 8).

As described above, the dermatitis-affected rat of the present invention has the characteristics that it lacks the ability to grow general hair and spontaneously develops dermatitis at a high incidence. Although the IgE antibody level in blood hardly fluctuates, the rat with dermatitis of the present invention can be used as a pathological model of allergic skin disease.

The dermatitis-induced rat of the present invention develops dermatitis on the back, which was found during the process of mating hairless rats (WBN / Ila-Ht), and the dermatitis spreads with aging. It originates from a mutated individual. That is, the dermatitis-affected rat of the present invention has a dominant heritable character, and thus can be compared with a normal hairy rat or hairless rat and KSD.
Although it can be produced by mating with R, in principle, it is desirable to maintain it by sibling mating, like other inbred strains.

For breeding the above-mentioned generation rats, 1-2 rats are housed in a cage made of wire mesh, the temperature is 23 ± 3 ° C., and the humidity is 55 ± 15.
%, Under a barrier system environment where the lighting time was adjusted from 7 am to 7 pm, solid feed (CE-2, manufactured by CLEA Japan, Inc.) was autoclaved (121 ° C, 15 minutes) as a feed, and a beverage As for the water, ultraviolet sterilized tap water is freely given. Mating is possible from 10 weeks of age.

The applicant of the present invention is ready to sell the thus-established rat having dermatitis of the present invention in accordance with the provision of Article 27-3, paragraph 1 of the Patent Law Enforcement Regulation.

The dermatitis-affected rat of the present invention can be used as a pathological model of dermatitis of allergic skin disease as described above. Therefore, by using this, screening and drug efficacy test of a preventive and therapeutic drug for the dermatitis can be performed. be able to. That is, a suitable amount of a test substance is applied to the hairless rat of the present invention by an administration route such as transdermal, injection, oral, etc., and its time-course action and effect are observed and evaluated by a general means to obtain a therapeutic drug. Screening and efficacy tests can be performed. In this case, there is an advantage that the test can be performed efficiently because labor such as hair removal is not required.

[0019]

The present invention will be described in more detail with reference to the following examples.

Example 1 Dermatitis-induced rat of the present invention (KS
DR) production and maintenance Hairless rat (WBN / Ila-Ht, Ishikawa Laboratory for Animal Research)
After breeding (brother-sister mating with F ₁ ) based on S. cerevisiae, it was found that in F ₄ , skin erythema developed on the back and the dermatitis spreads with aging. Spontaneous dermatitis can be produced by obtaining hairless rats by sibling mating (between male and female with hairless or between male and female with hairless) using this individual as an ancestor. As with other inbred strains, sibling mating can maintain the strain. Thus, the dermatitis-causing rat of the present invention has been established as an inbred strain after 20s and has been established as KSDR (Kowa Spontaneous Dermatitis Rat).
s).

[0021] WBN / Ila-Ht was propagated by randomly mating closed colonies JCL: Wistar of CLEA Japan, Inc. at the Ishikawa Institute for Experimental Animals in 1979. A male with hair was discovered, and this rat was subsequently backcrossed with WBN / Kob (Wistar Bonn / Kobori), one of the inbred strains, and was born in 1985 as F ₈ (N ₈ ). WBN / Ila-Ht homozygotes (Ht / Ht) are almost hairless when mature, and heterozygotes (Ht / +) are intermediate hair types between homozygous and normal hair (+ / +). (Hairless) is shown.

Each rat is kept in a cage made of wire mesh.
~ 2 animals, temperature 23 ± 3 ℃, humidity 55 ± 15%, under the barrier system environment adjusted lighting time from 7 am to 7 pm, solid feed as feed (CE-2, manufactured by CLEA Japan Co., Ltd. ) High-pressure steam sterilization (121 ° C., 15 minutes), and drinking water was provided with ultraviolet sterilized tap water freely.

Example 2 The characteristics of the KSDR obtained in Example 1 were examined. (1) The onset time and incidence of dermatitis F ₁₄ males (n = 16) and females (n = 12) had a postnatal age of 6
Up to 24 weeks of age, the occurrence and degree of lesion macroscopically once / week (none, mild: total dermatitis area is about 10 m
m ² , moderate: total dermatitis area about 100 mm ² ) was observed. As a result, mild lesions were observed in males at 10.1 ±
The age was 0.6 weeks, the female was 8.9 ± 0.5 weeks, and the moderate lesions were the male 13.9 ± 0.5 weeks and the female 11.2 ± 0.5 weeks. In addition, the incidence at 24 weeks of age was 9 males.
It was 3.8% and 100% for females, and it was observed that dermatitis develops earlier in females and lesions progress earlier than in males.

(2) As for the dermatitis-prone site and early-stage lesion, scratches-like erythema was observed in many rats in one to several places on the lower back (FIG. 1). Eventually,
Dermatitis spread throughout the back (Fig. 2).

(3) Histopathological Findings of Inflammatory Skin Area The epidermis at the normal site of KSDR is hyperplastic compared to hairy rats. Hyperplasia of the epidermis accompanied by hyperkeratosis was prominent in the skin inflammation area (Fig. 3). Furthermore, infiltration of eosinophils and mast cells was observed in the dermis.

(4) Transition of dermatitis area F ₁₄ males who developed dermatitis at 15 weeks of age (n =
Regarding 14), the skin area was measured once every 2 weeks until 21 weeks of age. The area is transparent paper (OH
Area line meter (Super paper)
It was measured using PLANIX β, manufactured by Mataya Measurement System).
The results are shown in Table 1.

[0027]

[Table 1]

As a result, the dermatitis area was about 30% in 2 weeks.
Expanded.

(5) Transition of body weight For F ₁₃ males (n = 16) and hairy rats (n = 5), body weights from 9 to 21 weeks of age were measured. The results are shown in Table 2.

[0030]

[Table 2]

From these results, the weight of KSDR tended to be slightly lighter than that of hairy rats.

(6) Blood and blood chemical findings Blood characteristics F _19-23 male and hairy rats were anesthetized and blood was collected from the posterior vena cava (EDTA added blood), automatic blood cell counter (Sysmex F-800,
It was measured by Toa Medical Electronics. The results are shown in Table 3 and Table 4.
Shown in.

[0033]

[Table 3]

[0034]

[Table 4]

From this, the white blood cell count of KSDR tended to increase as compared with that in normal (hairy) rats, and increased with aging (progress of dermatitis). Moreover, the dermatitis area and the white blood cell count were correlated.

White blood cell classification Table 5 shows the white blood cell classification of KSDR. As a result, lymphocytes decreased and neutrophils (segmental nuclei) and eosinophils increased with aging (progress of dermatitis).

[0037]

[Table 5]

Plasma Biochemical Values Plasma biochemical values are shown in Table 6. Based on this, plasma biochemical values were within the normal range for all items.

[0039]

[Table 6]

(7) Immune function F ₁₄ KSDR, hairy rats and normal Wistar rats were used once every 9 to 15 weeks of age and blood IgE of the same individual at a frequency of once / two weeks was used as a specific antibody. It was measured by the ELISA method. Blood was collected from the jugular vein of the rat under anesthesia.
The results are shown in Table 7.

[0041]

[Table 7]

From this, it was shown that the blood IgE level of KSDR did not rise. Therefore, in KSDR, I
It is thought to develop skin lesions that are not involved in gE production.

(8) Therapeutic effect The effectiveness of various drugs on the developed dermatitis was examined.
The results are shown in Table 8 and FIG.

[0044]

[Table 8]

Healing tended to occur with steroid drugs and with immunosuppressive drugs, but not with bactericidal / antibacterial drugs, antibiotics and antiallergic drugs. (9) Others The development of dermatitis was not affected by the breeding environment (barrier, open) and feed (CE-2 and MR: manufactured by Oriental Yeast Co., Ltd.).

[0046]

INDUSTRIAL APPLICABILITY The dermatitis-induced rat of the present invention serves as a pathological model of allergic dermatitis, and by using it, screening and drug efficacy test of a preventive and / or therapeutic drug for the dermatitis can be efficiently performed. it can.

[Brief description of drawings]

FIG. 1 is a photograph showing the external characteristics of KSDR (8 weeks old, male).

FIG. 2 is a photograph showing the external features of KSDR (60 weeks old, female).

FIG. 3 is a histopathological photograph of normal (A), normal KSDR (B) and inflammatory skin site (C) of hairy rats (AC: HE staining (× 33)).

FIG. 4 is a histopathological photograph showing infiltration of eosinophils (A) and mast cells (B) in the skin inflammation part of KSDR (A: HE staining (× 132), B: Alcian blue). Staining (x132)).

FIG. 5 is a photograph showing a drug treatment effect of KSDR on dermatitis (control group = base application; top: before application,
Bottom: 2 weeks after application, arrow: application part, prednisolone valerate (1.0 mg / kg) administration group; top: before application, bottom: 2 weeks after application, arrow: application part).

Front page continuation (51) Int.Cl. ⁷ Identification code FI theme code (reference) G01N 33/50 G01N 33/50 Z // A61K 31/573 A61K 31/573 (72) Inventor Jiro Matsumoto Higashimurayama, Tokyo 2-17-43 Noguchi (72) Inventor Teruji Koga 332-1 Ohno-Nitta Fuji City, Shizuoka Prefecture F-term (reference) 2G045 AA29 AA40 CA01 CA15 CB01 CB09 CB17 FA18 FB03 GA01 GC22 JA01 4C086 DA10 ZA89 ZB13

Claims (5)

[Claims] 1. A hairless rat lacking the ability to grow general hair, which has the following characteristics: (a) spontaneously developing mild erythema at about 8 to 12 weeks of age, (b) 24 weeks of age. After that, spontaneously develop dermatitis with an incidence of 90% or more,
(C) A dermatitis-onset rat having an IgE antibody level in blood that hardly changes. 2. Blood IgE at 11 to 15 weeks of age
The rat having dermatitis according to claim 1, which has an antibody value of about 65 ng / mL or less. 3. The dermatitis-producing rat according to claim 1 or 2, wherein the dermatitis is characterized by erythema and edema of the skin accompanied by infiltration of eosinophils and increase of mast cells. 4. The hairless rat or hairless rat is mated with a dermatitis-producing individual obtained by mating WBN / Ila-Ht hairless rat with siblings. A method for producing a rat with dermatitis. 5. A method for screening a compound effective for the prevention and / or treatment of dermatitis, using the dermatitis-causing rat according to any one of claims 1 to 3.