JPH11255658A - Medicine for disease caused by verotoxin - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a medicine for treating mucomembranaceous enteritis-based diseases caused by a verotoxin produced by pathogenic Escherichia coli. SOLUTION: This medicine comprises creosote, its component or a mixture containing the same as an active ingredient. When orally or intestinally administered to a person, the creosote, its component or the mixture is administered at a daily dose of about 1-500 mg per kg of adult body weight. When administered as an injection, the creosote, its component or the mixture is administered at a daily dose of 2-300 mg per kg of adult body weight.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

[0001] The present invention relates to a therapeutic agent for a vero toxin-induced disease, and more particularly to a pharmaceutical agent for treating a disease induced by vero toxin produced by pathogenic Escherichia coli.

[0002]

2. Description of the Related Art Pathogenic Escherichia coli is classified into several types according to the type of toxin it produces, and one of the types is called "enterohemorrhagic Escherichia coli". . Among them, especially O15
7: H7 (Since it is sometimes called simply O157,
Pathogenic E. coli, also referred to herein as "O157", produces a very potent cytotoxin called verotoxin. Vero toxin enters epithelial cells in the intestinal tract of animals infected with the pathogenic Escherichia coli, thereby hydrolyzing N-glycosidic bonds between specific adenine and ribose residues of 28S ribosomal RNA. The function is destroyed, and thus the protein synthesis function of the cell is stopped, causing the cell to die. This results in shedding of intestinal epithelial cells, resulting in diarrhea and bleeding. Also, in some cases,
Vero toxin enters the blood vessels and attacks vascular endothelial cells, causing renal damage and causing serious diseases such as hemorrhagic uremic syndrome.

[0003] Diseases caused by pathogenic Escherichia coli are bacterial infections, but the effects of antibiotics and antibacterial agents are not clear. On the other hand, it has also been reported that administration of antibiotics causes further worsening of symptoms. And its use is questioned today. What is clear is that
At present, there is no drug for treating a disease caused by verotoxin, and the development of such a drug is eagerly desired.

[Object of the Invention] The present invention has been made in view of the above circumstances, and an object of the present invention is to provide a medicament for treating a disease induced by verotoxin produced by pathogenic Escherichia coli. It is in.

[0005]

DISCLOSURE OF THE INVENTION The present inventors have made intensive studies in search of a substance having a protective effect on the intestinal mucosa which may cause damage to verotoxin. It was found that they possessed, and led to the present invention. That is, as a result of examining whether or not intestinal mucosal damage caused by the vero toxin is suppressed by the simultaneously administered drug, various drugs are simultaneously administered to the inside of the intestine of the animal. It has been discovered that creosote successfully suppresses intestinal mucosal injury caused by verotoxin, and has led to the present invention.

[0006] The remedy for vero toxin-induced diseases according to claim 1 is a remedy for diseases caused by vero toxin, and comprises creosote or a component thereof, or a mixture containing the same as an active ingredient (hereinafter simply referred to as " Creosote).

[0007] The therapeutic agent for verotoxin-induced disease according to claim 2 is the therapeutic agent for verotoxin-induced disease according to claim 1, wherein
The main symptom of the disease is intestinal mucosal disorder.

[0008]

DESCRIPTION OF THE PREFERRED EMBODIMENTS Creosote is widely used as a preservative, as is well known. This is based on its own bactericidal and antibacterial action, as can be seen from the fact that creosote is a Greek word meaning "preservation of meat". For this reason, for example, an adaptation method of immersing in a cotton ball and inserting it into a caries has been proposed and useful.

[0009] The drug for a bacterial toxin-induced disease of the present invention is mainly the same as the above creosote, but the subject is not a "bacterium" but a "toxin" produced by a bacterium.
The mechanism of action of the drug cannot be explained at all with the bactericidal action as described above. Therefore, it cannot be said that the drug of the present invention exerts its efficacy based on a pharmacological action that cannot be predicted at all from the previously recognized mechanism of action, but the detailed mechanism of action is currently unknown. It is unknown at this time.

Wood creosote, beechwood
creosote) is mainly obtained by fractionation of wood tar and is transparent and liquid at room temperature (The Merck Index, by S. Budavari, Me.
rck Publishing Co., 1989). It has a peculiar odor and is a mixture of mainly aromatic chemicals such as guaiacol and cresol (Ogata et al., N. Ogata, T. Baba, Res.
Commun. Chem. Pathol. Pharmacol. 66, 411-423, 198
9). Other components include cresol, phenol and xylenol. This "creosote" is often mixed because of the same name, but is completely different from the "coal tar creosote" obtained from coal tar.

[0011] Creosote, or a component thereof or a mixture containing the same, is used in a general form in medical drugs. Common forms include, for example, tablets, pills, powders, capsules (soft capsules, hard capsules), granules, oral liquids, and injections (intravascular, intramuscular, subcutaneous, skin) Internal administration) or suppositories.

For the purpose of treating intestinal mucosal injury caused by verotoxin, the dose of creosote or its components varies depending on the type of animal, sex, age, and degree of symptoms, but cannot be generalized. , Oral or rectal administration in humans (suppository)
In the case of, approximately 1 to 500 mg, preferably 2 to 100 m
g. In the case of administration as an injection, the dose is 0.2 to 300 mg, preferably 0.2 to 50 mg, per 1 kg of adult body weight per day. These daily doses may be administered in 2 to 4 divided doses.

[0013]

The present invention will be described in more detail with reference to the following Examples, which by no means limit the present invention. Further, the dosage form, administration method, animal, and the like are not limited to this example.

In the following examples, verotoxin was administered into the intestine of rabbits, and the resulting intestinal mucosal damage was confirmed by microscopic examination. In parallel with this, in other rabbits, verotoxin and a test substance were simultaneously administered into the intestinal tract, and it was examined whether the test substance suppressed intestinal mucosal damage. The details will be described below.

Reference experiment (Rabbit small intestine against saline solution )
Absorption) First, the extent of absorption of the rabbit small intestine into the saline solution was examined.

An 8-week-old male NZW rabbit (obtained from Kitayama Labes, weighing about 2 kg) was fasted for 24 hours. However, water was given freely. Rabbits are placed in an incubator, and pentobarbital sodium (Abbott) weighs 1 in the ear vein.
Anesthetized by slow intravenous injection of 25-30 mg / kg. After anesthesia, sufficient attention was paid to vital signs such as respiration, and sufficient attention was paid to deterioration of the general condition due to excessive anesthesia. Next, the patient was placed in the supine position on the operating table, the abdomen was quickly shaved with an electric clipper, disinfected with an isodine solution (manufactured by Meiji Seika Co., Ltd.), and the abdomen was opened through a median incision. The bleeding on the way was appropriately stopped with a small hemostatic forceps (for example, mosquito forceps) to prevent massive bleeding. The operation was aseptic as possible. After laparotomy, the origin of the jejunum was found, and a silk thread was placed at a position of 10 cm therefrom to ligate the jejunum. If necessary, the laparotomy was maintained with a retractor or hook. The ligature was carried out by holding a needle for suturing through No. 3 silk thread with a needle holder and passing it through the portion of the mesentery attached to the intestinal tract. At this time, attention was paid to the degree of ligating, because too weak a substance may cause intestinal contents to pass over a ligated part, and too strong a substance may damage tissues. In addition, care was taken to minimize damage to the mesentery and its blood vessels. All organizations also avoided the damage by treating them lovingly. Next, the anal side 5c
A similar ligation was performed at the position of m to make a "small intestine segment" of 5 cm. The same ligation was sequentially performed on the anal side by 5 cm each to make a total of three “small intestine segments”. During the surgery, the rabbit's body temperature tends to fall, so it was necessary to keep it warm in some way, for example, by slightly warming the bottom of the operating table or by heating around the body with a disposable body warmer. .
However, careful attention was paid to overheating (a little warmer is better).

For the lumens of the first to third "small intestines",
The components shown in Table 1 were injected, and the intestinal fluid volume after 3 hours and 18 hours was measured.

[0018]

[Table 1]

As is clear from Table 1, in the presence of glucose, absorption of intestinal fluid is promoted (0.92 m
0.61 ml was significantly different from 1), and it was found that creosote had no effect on intestinal fluid absorption (does not interfere with intestinal fluid absorption) in such an experimental system (0.61 ml). 0.60 ml is not significantly different). By the way, in the measurement after 3 hours as described above, the absorption of the salt solution in the presence of glucose is promoted because the sodium ion and glucose via sodium-glucose cotransporter present in the intestinal epithelium are associated with glucose. This is due to the absorption promoting effect, which is the principle of the oral rehydration agent (generally referred to as ORS) recommended by WHO. In the above-described experiments, the concentrations of the components were totaled to make them isotonic. Otherwise, the absorption rate changes due to the difference in osmotic pressure.

In this experiment (intestinal mucosal damage caused by verotoxin and
Disability control by creosote) Specimen Vero toxin (VT) specimen (50 ng / ml, 2 ml / 1
vial, Strain ATCC 43890 culture culture) was stored frozen at the time of acquisition (stored at -20 ° C or lower). In addition, it thawed immediately before use, and after thawing, it stirred enough.

2. Experimental animals (1) Animals used Male rabbits produced by Kitayama Labes Co., Ltd. (kbl:
NZW) 8 animals were purchased and used at 8 weeks of age or older (body weight: 1.50 to 2.20 kg) after quarantine and acclimation for 7 days or more.

(2) Rearing conditions During the test period, the temperature is 19 to 25 ° C. and the humidity is 40 to 70.
%, Ventilation rate 16 times / hour and lighting 12 hours (7-1)
(9 o'clock) / day, the animals were individually housed and raised in aluminum cages [35W x 50D x 35H (cm)] in a breeding room. During that time, they were allowed to freely take solid feed (LRC4, Oriental Yeast Co., Ltd.) and tap water.

3. Experimental method (1) Preparation of small intestine division In the same manner as in the above "Preliminary experiment (absorption of rabbit small intestine to salt solution)", 5 cm was collected in the small intestine of each rabbit.
Eight "small intestine divisions" were made. These "small intestine divisions" are assigned No. 1 to No. And 8.

1.5 ml of each sample (see [Table 2] below) was administered to the intestinal lumen of each small intestine segment (see [Table 3] below for which sample was administered to which location). .

[0025]

[Table 2]

[0026]

[Table 3]

After the administration, 0.1 w /
The peritoneal cavity was washed with v% pentocillin physiological saline (pentocillin: Toyama Chemical, lot number: AF650),
The muscular layer and the skin were separately sutured. After suturing, only water was freely taken.

(2) Preparation of Specimen for Microscopic Examination A specimen for an optical microscope of the small intestine was prepared as follows.
That is, each small intestine segment was fixed with a 10% neutral buffered formalin aqueous solution. Further, each small intestine segment was embedded in paraffin according to a conventional method to prepare a hematoxylin-eosin stained specimen.

(3) Results As described above, in order to examine the inhibitory effect of creosote on intestinal mucosal damage induced by verotoxin,
1.5 ml of a culture supernatant of Escherichia coli O157, that is, a liquid containing 50 ng / ml of verotoxin and 0-100 μg / ml of creosote added thereto
Was injected into the small intestinal segment of a rabbit 5 cm in length. 8 after injection
Time and microscopic observation of mucosal damage in each small intestine segment showed that when creosote was 0 μg / ml (absence) (when using sample 1), a high degree of intestinal mucosal damage was observed due to verotoxin. Sorting 1μg / ml, 5μg
/ Ml, 10 μg / ml, 30 μg / ml, and 100
When the amount was contained at μg / ml, the intestinal mucosa was not significantly damaged, and the inhibitory effect of creosote on intestinal mucosal damage could be confirmed. Here, sample 1 (creosote 0 μg / ml), sample 2 (creosote 1 μg / m
l), sample 3 (creosote 5 μg / ml), sample 4
(Creosote 10 μg / ml), sample 5 (creosote 30 μg / ml) and sample 6 (creosote 100 μg / ml).
g / ml) are shown in FIG. 1 to FIG. 6 each showing a micrograph showing the state of the small intestinal mucosa (optional). As can be seen from FIGS. 1 to 6, creosote has an effect of suppressing intestinal mucosal damage caused by verotoxin.
This indicates that creosote can be used as a therapeutic to pathogenic E. coli. In addition, the intestinal mucosa protective effect of creosote described above appears most remarkably when sample 6 is used, and then sample 5, sample 4, sample 3, and sample 2
It is in order.

[0030]

According to the present invention, it is possible to provide a medicament for treating a disease induced by verotoxin produced by pathogenic Escherichia coli.

[Brief description of the drawings]

FIG. 1 is a photomicrograph showing the state of the intestinal mucosa when sample 1 is used.

FIG. 2 is a micrograph showing the state of the intestinal mucosa when a specimen 2 is used.

FIG. 3 is a photomicrograph showing the state of the intestinal mucosa when a specimen 3 is used.

FIG. 4 is a photomicrograph showing the state of the intestinal mucosa when a specimen 4 is used.

FIG. 5 is a photomicrograph showing the state of the intestinal mucosa when a specimen 5 is used.

FIG. 6 is a photomicrograph showing the state of the intestinal mucosa when the sample 6 is used.

Claims (2)

[Claims] 1. A therapeutic agent for a disease caused by verotoxin, which comprises creosote or a component thereof or a mixture containing the same as an active ingredient. 2. The therapeutic agent for vero toxin-induced disease according to claim 1, wherein the main symptom of the disease is intestinal mucosal damage.