Effects of He-Ne laser irradiation on chronic atrophic gastritis in rats

Abstract

AIM: To study the effects of He-Ne laser irradiation on experimental chronic atrophic gastritis (CAG) in rats.

METHODS: Sixty-three male adult Wistar rats were randomly divided into five groups including normal control group, model control group and three different dosages He-Ne laser groups. The chronic atrophic gastritis (CAG) model in rats was made by pouring medicine which was a kind of mixed liquor including 2% sodium salicylate and 30% alcohol down the throat for 8 wk to stimulate rat gastric mucosa, combining with irregular fasting and compulsive sporting as pathogenic factors; 3.36, 4.80, and 6.24 J/cm² doses of He-Ne laser were used, respectively for three different treatment groups, once a day for 20 d. The pH value of diluted gastric acid was determined by acidimeter, the histopathological changes such as the inflammatory degrees in gastric mucosa, the morphology and structure of parietal cells were observed, and the thickness of mucosa was measured by micrometer under optical microscope.

RESULTS: In model control group, the secretion of gastric acid was little, pathologic morphological changes in gastric mucosa such as thinner mucous, atrophic glands, notable inflammatory infiltration were found. After 3.36 J/cm² dose of He-Ne laser treatment for 20 d, the secretion of gastric acid was increased (P < 0.05), the thickness of gastric mucosa was significantly thicker than that in model control group (P < 0.01), the gastric mucosal inflammation cells were decreased (P < 0.05). Morphology, structure and volume of the parietal cells all recuperated or were closed to normal.

CONCLUSION: 3.36 J/cm² dose of He-Ne laser has a significant effect on CAG in rats.

Key Words: Chronic atrophic gastritis, Laser, He-Ne, Rat

INTRODUCTION

Chronic atrophic gastritis (CAG) is one of the most common digestive diseases worldwide[1-4]. The patient’s gastric mucosa atrophies, gastric sinus secret cells reduce, function is weakened, gastric acid secretion reduces, especially pathological changeable epithelium often contains intestine epithelium metaplasia and atypical hyperplasia, which are often seen as precancerous lesions of gastric carcinoma[5-7]. To the author’s knowledge, clinics still have no perfect and effective treatment projects[8]. So we successfully established an animal model with CAG in rats[9], used different radiation intensity of He-Ne laser to irradiate at the gastric projective area of rat with CAG, to study the therapeutic effects by examining changes of gastric acid and observing changes of pathologic histology in gastric mucosa.

MATERIALS AND METHODS

Animals

A total of 52 male adult Wistar rats weighing 180-230 g were purchased from Experimental Animal Center, Capital Medical University. They were housed in an air-conditioned room with 12 h dark/light cycle, and randomly divided into five groups: eight in normal control group, 11 in model control group, 11 in 3.36 J/cm² dose He-Ne laser group (small dose He-Ne laser group), 11 in 4.80 J/cm² dose He-Ne laser group (inside dose He-Ne laser group), 11 in 6.24 J/cm² dose He-Ne laser group (big dose He-Ne laser group). CAG model in rats were made by mixing 2% sodium salicylate and 30% alcohol and poured down their throats for 8 wk to stimulate their gastric mucosa, combined with irregular fasting and compulsive sporting.

He-Ne laser therapy

He-Ne laser therapy was carried out using the HJ-3DB He-Ne laser machine (wavelength 632.8 nm) which was made by Nanjing laser instrument factory, the He-Ne laser was amplified by convex lens, vertically radiated at the gastric projective area. The anterioposterior diameter of laser was 1 cm, its power density was 8 mW/cm². The following formed the course of treatment once a day for 20 d.

Small dose He-Ne laser treatment: irradiated 7 min each time, 3.36 J/cm² of dosage;

Inside dose He-Ne laser treatment: irradiated 10 min each time, 4.80 J/cm² of dosage;

Big dose He-Ne laser irradiation: irradiated 13 min each time, 6.24 J/cm² of dosage.

Process

The rats in each group were killed at the desired time-point. The animals were deprived of food but were offered enough water before being killed at 24 h. The rats were anesthetized with ether, the belly was opened immediately, cadre was ligature, the whole stomach was taken off, and the surface blood stains were washed by normal saline water, sucked dry with filter paper, then the gastric cavity was opened along greater curvature of stomach, flushed with distilled water, and the dilution gastric liquid (provided to measure the gastric acid pH value) was collected, then front and back gastric sinus and parts of stomach organized roughly into 3 mm × 10 mm were taken in parallel with lesser curvature of stomach, fixed in neutral formalin, embedded in paraffin wax, 6 µm sections, stained with hematoxylin-eosin (HE) for pathological examination.

Determination of gastric acid

The rat’s gastric acid (pH value) was determined five times for each example by acidimeter (PHS-3C) from Shanghai Instrument Factory, and took the average value as the pH value of this animal.

Pathology examinations

Take three sliced pieces for each example, and select three different visual fields for each slice such as the body of stomach, gastric sinus and the area of the body of stomach marked with gastric sinus. The whole gastric mucosa was observed under light microscope, which included the following aspects: (1) the immunity degree of the gastric mucosal ingluvitis cellular was divided into two grades such as negative (-) (no ingluvitis cells or some ingluvitis that jot spread at the gastric mucus) and positive (+) (more ingluvitis cell at the gastric mucosa or series of ingluvitis cells aggregated inside the mucus), counted the positive rate respectively by each visual field; (2) measured the thickness of the gastric mucosa with microscope, regarded µm as the unit, adopted the average of the mucosa thickness to show the reflection hyperplasia circumstance; (3) observed the parietal cell including the appearance, construction and arrangement.

Statistical analysis

Software SPSS 10.0 was used in the statistical analysis, parameters were expressed as mean±SD, and compared using One-way ANOVA analysis of variance, followed by χ² tests and differences were considered significant at P < 0.05.

RESULTS

Gastric acid

The determination results of gastric acid showed that the secret function of gastric mucosa in model control rats was more decreased than that in both normal control group and small dose He-Ne laser group (P < 0.05, Table 1).

Table 1 Detection of gastric results of pH value in Wistar rats.

Group	n	pH value (mean±SD)
Normal control	6	3.72 ± 1.02a
Model control	7	5.86 ± 1.45
Small dose He-Ne laser	7	4.43 ± 1.18a
Inside dose He-Ne laser	6	4.66 ± 1.00
Big dose He-Ne laser	7	4.57 ± 1.48

^aP < 0.05 vs control group.

Pathological findings

The pathological histologies were observed under microscope. In normal control group, the gastric mucosa was thick, the epithelium was complete, only fell ingluvitis were spread, there was no obvious ingluvitis cellular immunity (Table 2). The parietal cells in normal control group were as follows: large volume, pyramid form, cell edge tactful, full, numerous, were arranged neatly and above 2/3 at a fundic gland (Figure 1A). In model control group, the gastric mucosa was thinner than that in normal control group (P < 0.01), there were large quantities of immunity ingluvitis (P < 0.01), most of the cells were lymphocytes and plasma cells. The parietal cells in model control group were as follows: volume sterigma, cell edge anomaly polygon, afterbirth syrup was wrinkled and arranged foul-up, intercellular space was enlarged, the ratio in syrup and nucleosidase increased the breadth of the gland antrum, and large quantity of cells were put through empty transformation (Figure 1B). In He-Ne laser groups, the gastric mucosa was thicker than that in model group (P < 0.01), ingluvitis cells were less than that in model control group, particularly the small dose He-Ne laser group (P < 0.05). The parietal cells in small dosage He-Ne laser group were as follows: cell appearance, volume and construction were all near to normal, and arranged neatly (Figure 1C). Besides, there was no obvious ingluvitis edema in normal control group, model control group and small dosage He-Ne laser group, but it was obvious in inside and large dose He-Ne laser groups, and the cells were transformed in the two groups.

Figure 1 Pathology changes of parietal cells. A: parietal cells of gastric in model control group (HE ×400); C: parietal cells of gastric mucosa in small mucosa in normal control group (HE ×400); B: parietal cells of gastric mucosa dose He-Ne laser group (HE ×400).

Table 2 Changes of gastric mucosa thickness and the observations of ingluvitis cellular immunity degree.

Group	n	The thickness of gastric mucosa(µm) (mean±SD)	Ingluvitis cellular immunity degree			Ingluvitis edema
			-	+	Positive rate (%)	Yes	No
Normal control	8	525.17 ± 57.52b	7	1	13b		√
Model control	11	387.21 ± 51.60	1	10	91		√
Small dose He–Ne laser	11	499.06 ± 57.25b	6	5	45a		√
Inside dose He–Ne laser	11	456.77 ± 47.56b	1	10	91	√
Big dose He–Ne laser	11	475.62 ± 53.75b	2	9	82	√

^aP < 0.05,

^bP < 0.01 vs model control group.

DISCUSSION

The cause of disease and pathogenesis of chronic atrophic arthritis (CAG) are not yet completely clean, generally speaking, it is a synthesis factor with more effects and actions such as immunity, gall back streaming, pyloric infection, food, wine, smoke, and drugs[2,8,9]. Research expressed that sodium salicylate could hurt gastric mucosa and enzyme could inhibit gastric cell growth[10], alcohol could incite gastric mucosa. In our experiment, we poured a mixture of 2% sodium salicylate and 30% alcohol down the rat’s throat for 8 wk, which could affect the natural cover and organize variety in gastric mucus, combined with fatigue from overwork, hunger and dissatisfaction in order to make the pathological changes of CAG appear on the gastric mucosa appear[9]. Since the parietal cells reduced and degenerated, there was disorder both in gastric acid secretion and digestion enzyme weakness, the performance was a series of weak symptoms of the function of digestion and absorption such as the reduction of gastric acid secretion, diminishing of the gland and atrophy, and metaplasis of the intestinal epithelium[9].

He-Ne laser, its characteristics such as 632.8 nm of wavelength, good directivity , high intensity, good monochro-maticity, is a low-level laser[11-14]. The low-level laser has some biology effects such as cell vitality[15-18], phagocytosis[19-21], immune responses[22-25], but as far as its usage for the treatment of digestive disease, there were only few reports[26-28]. On one hand, there was no difference between treating gastric ulcer through bark and by endoscope[29]. On the other hand, external irradiation is easy, and patients can bear some pain with ease. We treated rats with CAG by external irradiation with He-Ne laser as amplified by a convex mirror, radiated at the gastric projective area. The results implied that the gastric acid secretion of the rats in CAG treated by He-Ne laser irradiation was increased, the best effect was on the animals treated by small dosage He-Ne laser. Some changes in gastric mucus such as thickness of gastric mucosa, alleviated ingluvitis, and parietal cell hyperplasia were found in this experiment. We think the mechanism of He-Ne laser treated CAG in rats as follows: (1) partial microcirculation was ameliorated; (2) body immunity was enhanced; (3) the organized growth of gastric mucosa was promoted; (4) inflammation was eliminated. Moreover, in our experiment we found that three different dosages of He-Ne laser preceding the incitement to the rat in CAG, resulted in different effects. The best effect was the small dosage of He-Ne laser, the inside dose and the large dose were not as well as the small dose of He-Ne laser, which showed the dependence on dosage[20,30,31]. Perhaps it is because the biology effect of He-Ne laser closely related to the action time, power density that organized He-Ne laser. Under the same power density (8 mW/cm²), different action time (small dose for 7 min, inside dose for 10 min, large dose for 13 min), produced different effects. These results showed that small dose He-Ne laser was a positive action, but large dose was an inhibitory action to incite lombricine organization[32]. Besides, some damaged cells, individual swelling, afterbirth syrup degeneration and ingluvitis edema in the inside dose He-Ne laser group and the big dose He-Ne laser group were found in gastric mucus of rats under microscope. This may indicate the light dynamic damage of He-Ne laser. So we draw a further conclusion that big doses of He-Ne laser were deformative. In a word, the experiment showed that 3.36 J/cm² dose of He-Ne laser is a valid dose of external irradiation to incite life, it could promote the secretion of gastric acid and recover the function of gastric mucus to the CAG in rats.

In summary, 3.36 J/cm² /d dose of He-Ne laser irradiation is a well-tolerated, safe, and effective treatment in rats with CAG. The technique is easy, inexpensive, and of short duration[25]. It is necessary to translate the outcome of this study into clinically relevant interventions by further studies which would develop a new way for the treatment of CAG.