**1. Introduction**

Recent studies have demonstrated the immunomodulatory effects of heat-killed lactic acid bacteria. The aim of this study was to evaluate the protective effect of

heat-killed *Enterococcus faecalis EF-2001* (EF-2001) on a radiation protective and immunopotentiating Effect.

Drugs that suppress the radiosensitizer that enhances the biological action of ionizing radiation are collectively called radioprotector. Drugs that show protective effects by radical scavenging or hypoxic action, such as WR-2721, need to be administered before exposure [1]. Interleukins and the like are also effective with prior administration. Drugs that enhance *in vivo* production of hematopoietic cell growth factors, such as hematopoietic cell growth factors such as G-CSF and immunostimulants such as 432OK, are effective even after administration [2]. Chelating agents and iodine agents that prevent radionuclides from being absorbed from the skin and gastrointestinal tract and deposited in tissues and promote excretion are broadly considered protective agents [3].

Protective agents for radical scavengers have been studied as an adjunct to cancer radiotherapy, and WR-2721 (S-2-(3-Aminopropylamino) ethylphosphorothioic acid) is the most promising among many compounds. It is thought to show a protective action by eliminating radicals caused by indirect action of radiation, hydrogenation, reduction of oxygen effect, etc. A drug that can reduce radiation damage when administered immediately before radiation exposure is called a radioprotective drug. Typical examples are aminothiol derivatives such as cysteamine (mercaptoethylamine) and WR-2721. At the time of tumor radiotherapy, clinical application has been considered for protecting normal tissues around the tumor, but it has not been put into practical use. In addition, there is a possibility that these radiation prevention drugs may be used to protect rescue workers and decontamination workers in the event of a nuclear facility accident, but all currently known drugs are stamina, judgment, agility. Side effects such as decreased sex and vomiting are strong and have not been put to practical use [4].

Mn, Zn, and Cd ions, as activators of biological defense mechanisms, induce protective effects by inducing biosynthesis of metallothionein with radical scavenging ability in cells. In addition, gelen, interleukin 1 (IL-1), lipopolysaccharide (lipopolysaccharide, LPS), muramyl dipeptide (MDP) derivatives, ginseng extract, etc. Effective when administered prior to exposure. Leucons and innocities are nucleic acid precursors that are approved as drugs for leukopenia and have radiation protection but are not strong [5].

G-CSF (granulocyte colony-stimulating factor), a hematopoietic cell growth factor, is a growth factor for leukocyte progenitor cells such as neutrophils. Prevents infection and bleeding and provides lifesaving effect. GM-CSF (granulocyte/macrophage colony-stimulating factor) and interleukin 6 (IL-6) have similar effects, but G-CSF is most expected in terms of fewer side effects. These proteins are natural substances collectively called cytokines, but can be produced as recombinants by genetic engineering techniques. In addition, picibanil (OK-432), glucan, lactic acid bacteria preparations, etc. are said to enhance in vivo production of hematopoietic cell growth factor and are effective when administered after exposure. Hematopoietic hormones, such as GM-CSF, are actually post-accidents in Brazil and San Salvador used for exposed people [6].

New Protective Agents Stable nitroxide radicals have SOD activity and protect animal cells from oxidative stress caused by superoxide and hydrogen peroxide. Tempol, one of the stable nitroxide radicals, has been shown to exhibit radioprotective effects on C3H mice both in vitro and in vivo. It has also been shown to prevent radiation-induced bone marrow damage and appears promising as a protective agent in cancer radiotherapy [7].

**103**

*New Screening for the Development of Radioprotectors: Radioprotection and Anti-Cancer Effect…*

promote excretion as agents for preventing absorption and deposition of radionuclides in the body and promoting elimination these are protective agents in a broad sense. Prussian blue is used to prevent absorption of 137Cs. To promote 90Sr excretion, calcium citrate and sodium alginate are recommended. In addition, sodium citrate and a low phosphorus diet are effective. Geralmine (aluminum hydroxide gel) has the same effect as a low phosphorus diet because it inhibits the absorption of phosphoric acid. Zn-DTPA or Ca-DTPA is used to promote plutonium excretion. An iodine agent (potassium iodide agent) is effective for preventing radioactive

β-glucan is an edible mushroom, belonging to the β-glucan, and named β-glucan. It particularly resembles *L.shimeji* Hongo in its taste and touch. It is a stump mushroom, which grows in forest, having a grayish brown-shaped umbrella about 4–9 cm in diameter [11]. Glucan (β-*glucan decastes* Sing) was called the β-glucan aggregate and has highly been valued for a long time, and its artificial cultivation was difficult till now. Recently, there has been artificial cultivation using bacterial strains as seedlings [12]. Now you can see it with general partial exposure. The authors studied glucan for antitumor, angiotensin converting enzyme inhibition, and serum cholesterol-lowering activity [13]. Glucan was found to have higher activity than the same class of shiitake Enteroccous Faecalis in anti-tumor activity, β-1,6 D-glucan and β-1,3D-glucan, and their active β-1, 6 D-glucan contains β-1,3Dglucan [14]. Here, we report the effect of glucan extract on radiation therapy in

EF-2001 is a commercially available probiotic that was originally isolated from healthy human infant feces. Nihon BRM Co. Ltd. (Tokyo, Japan) supplied it as a heat-killed, dried powder. One gram of dried EF-2001 is equivalent to over 7.5 × 1012 colony-forming units prior to being heat-killed. *Nihon BRM Co. Ltd. (Tokyo, Japan) supplied Enteroccous Faecalis 2001®, a glucan product, composed of yeast extract,* 

*Dry Enterococcus faecalis* contains 35–45 g of β-glycan per 100 g. *Enteroccous Faecalis* was orally administered using a zoned for 2 weeks before the start of the

The experimental animals used in this study are ICR / Slc mice. ICR/Slc mice (5 weeks old, male) were obtained from SLC, Japan. Once received, the mice were pre-bred for a week and only healthy mice were used for testing. Mice were fed with commercial feed (CA-1, CLEA Japan) and water ad libitum, with a 12-hour light cycle (8:00 lights, 20:00 lights off). The room humidity and temperature were

The experimental groups were divided into control group, tumor seeding group, tumor seeding +2 Gy exposure group, β-glucan treatment group, β-glucan + tumor

seeding group, β-glucan + tumor seeding +2 Gy exposure by 6 groups.

*DOI: http://dx.doi.org/10.5772/intechopen.95016*

iodine from collecting in the thyroid gland [8–10].

mice with cancer.

**2.1** β**-Glucan**

*dextrin and gelatin*.

experiment.

**2.2 Animal**

**2. Materials and methods**

60–65% and 22 ± 2° C, respectively.

**2.3 Experimental groups**

Chelating agents and iodine agents that prevent radionuclides from being absorbed from the skin and gastrointestinal tract, or deposited in tissues, or

*New Screening for the Development of Radioprotectors: Radioprotection and Anti-Cancer Effect… DOI: http://dx.doi.org/10.5772/intechopen.95016*

promote excretion as agents for preventing absorption and deposition of radionuclides in the body and promoting elimination these are protective agents in a broad sense. Prussian blue is used to prevent absorption of 137Cs. To promote 90Sr excretion, calcium citrate and sodium alginate are recommended. In addition, sodium citrate and a low phosphorus diet are effective. Geralmine (aluminum hydroxide gel) has the same effect as a low phosphorus diet because it inhibits the absorption of phosphoric acid. Zn-DTPA or Ca-DTPA is used to promote plutonium excretion. An iodine agent (potassium iodide agent) is effective for preventing radioactive iodine from collecting in the thyroid gland [8–10].

β-glucan is an edible mushroom, belonging to the β-glucan, and named β-glucan. It particularly resembles *L.shimeji* Hongo in its taste and touch. It is a stump mushroom, which grows in forest, having a grayish brown-shaped umbrella about 4–9 cm in diameter [11]. Glucan (β-*glucan decastes* Sing) was called the β-glucan aggregate and has highly been valued for a long time, and its artificial cultivation was difficult till now. Recently, there has been artificial cultivation using bacterial strains as seedlings [12]. Now you can see it with general partial exposure. The authors studied glucan for antitumor, angiotensin converting enzyme inhibition, and serum cholesterol-lowering activity [13]. Glucan was found to have higher activity than the same class of shiitake Enteroccous Faecalis in anti-tumor activity, β-1,6 D-glucan and β-1,3D-glucan, and their active β-1, 6 D-glucan contains β-1,3Dglucan [14]. Here, we report the effect of glucan extract on radiation therapy in mice with cancer.

## **2. Materials and methods**

#### **2.1** β**-Glucan**

*Translational Research in Cancer*

immunopotentiating Effect.

broadly considered protective agents [3].

protection but are not strong [5].

San Salvador used for exposed people [6].

agent in cancer radiotherapy [7].

heat-killed *Enterococcus faecalis EF-2001* (EF-2001) on a radiation protective and

Protective agents for radical scavengers have been studied as an adjunct to cancer radiotherapy, and WR-2721 (S-2-(3-Aminopropylamino) ethylphosphorothioic acid) is the most promising among many compounds. It is thought to show a protective action by eliminating radicals caused by indirect action of radiation, hydrogenation, reduction of oxygen effect, etc. A drug that can reduce radiation damage when administered immediately before radiation exposure is called a radioprotective drug. Typical examples are aminothiol derivatives such as cysteamine (mercaptoethylamine) and WR-2721. At the time of tumor radiotherapy, clinical application has been considered for protecting normal tissues around the tumor, but it has not been put into practical use. In addition, there is a possibility that these radiation prevention drugs may be used to protect rescue workers and decontamination workers in the event of a nuclear facility accident, but all currently known drugs are stamina, judgment, agility. Side effects such as decreased sex and

Mn, Zn, and Cd ions, as activators of biological defense mechanisms, induce protective effects by inducing biosynthesis of metallothionein with radical scavenging ability in cells. In addition, gelen, interleukin 1 (IL-1), lipopolysaccharide (lipopolysaccharide, LPS), muramyl dipeptide (MDP) derivatives, ginseng extract, etc. Effective when administered prior to exposure. Leucons and innocities are nucleic acid precursors that are approved as drugs for leukopenia and have radiation

G-CSF (granulocyte colony-stimulating factor), a hematopoietic cell growth factor, is a growth factor for leukocyte progenitor cells such as neutrophils. Prevents infection and bleeding and provides lifesaving effect. GM-CSF (granulocyte/macrophage colony-stimulating factor) and interleukin 6 (IL-6) have similar effects, but G-CSF is most expected in terms of fewer side effects. These proteins are natural substances collectively called cytokines, but can be produced as recombinants by genetic engineering techniques. In addition, picibanil (OK-432), glucan, lactic acid bacteria preparations, etc. are said to enhance in vivo production of hematopoietic cell growth factor and are effective when administered after exposure. Hematopoietic hormones, such as GM-CSF, are actually post-accidents in Brazil and

New Protective Agents Stable nitroxide radicals have SOD activity and protect animal cells from oxidative stress caused by superoxide and hydrogen peroxide. Tempol, one of the stable nitroxide radicals, has been shown to exhibit radioprotective effects on C3H mice both in vitro and in vivo. It has also been shown to prevent radiation-induced bone marrow damage and appears promising as a protective

Chelating agents and iodine agents that prevent radionuclides from being absorbed from the skin and gastrointestinal tract, or deposited in tissues, or

vomiting are strong and have not been put to practical use [4].

Drugs that suppress the radiosensitizer that enhances the biological action of ionizing radiation are collectively called radioprotector. Drugs that show protective effects by radical scavenging or hypoxic action, such as WR-2721, need to be administered before exposure [1]. Interleukins and the like are also effective with prior administration. Drugs that enhance *in vivo* production of hematopoietic cell growth factors, such as hematopoietic cell growth factors such as G-CSF and immunostimulants such as 432OK, are effective even after administration [2]. Chelating agents and iodine agents that prevent radionuclides from being absorbed from the skin and gastrointestinal tract and deposited in tissues and promote excretion are

**102**

EF-2001 is a commercially available probiotic that was originally isolated from healthy human infant feces. Nihon BRM Co. Ltd. (Tokyo, Japan) supplied it as a heat-killed, dried powder. One gram of dried EF-2001 is equivalent to over 7.5 × 1012 colony-forming units prior to being heat-killed. *Nihon BRM Co. Ltd. (Tokyo, Japan) supplied Enteroccous Faecalis 2001®, a glucan product, composed of yeast extract, dextrin and gelatin*.

*Dry Enterococcus faecalis* contains 35–45 g of β-glycan per 100 g. *Enteroccous Faecalis* was orally administered using a zoned for 2 weeks before the start of the experiment.

#### **2.2 Animal**

The experimental animals used in this study are ICR / Slc mice. ICR/Slc mice (5 weeks old, male) were obtained from SLC, Japan. Once received, the mice were pre-bred for a week and only healthy mice were used for testing. Mice were fed with commercial feed (CA-1, CLEA Japan) and water ad libitum, with a 12-hour light cycle (8:00 lights, 20:00 lights off). The room humidity and temperature were 60–65% and 22 ± 2° C, respectively.

#### **2.3 Experimental groups**

The experimental groups were divided into control group, tumor seeding group, tumor seeding +2 Gy exposure group, β-glucan treatment group, β-glucan + tumor seeding group, β-glucan + tumor seeding +2 Gy exposure by 6 groups.

#### **2.4 Irradiation device**

The X-ray generator used was Philips MG226/4.5. For the pipe voltage, a dose of 200 kV was applied at a rate of 0.35 Gy/min. The total amount of X-rays exposed was 2 Gy. The exposed place of the mouse was fixed to the position of the front part of the left foot (place where cancer cells were seeded) with a holder, and the place not irradiated was covered with a lead container. X-rays were irradiated on the 1 and 3 days.

The specific radiation irradiation method in each experimental group is shown below. For changes in blood cells, single irradiation with 2 Gy was performed. However, in the radiation tumor effect experiment, 2 Gy was divided into 3 divided doses.

#### **2.5 Tumor inoculation**

In order to obtain reproducible experimental data, mice were inoculated on the 15th day after breeding for more than 1 month. Approximately 2 x 106 sarcoma 180 cancer cells were inoculated into the muscle of the left foot of an ICR male mouse.
