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. 2020 Jul 17;29(1):19–27. doi: 10.5978/islsm.20-OR-02

Helicobacter pylori Eradication Using Laser Endoscope and Methylene Blue

Kouji Ogasawara 1,2,✉, Susumu Nakajima 3, Hiroshi Sato 4, Tadashi Sasaki 4
PMCID: PMC7447828  PMID: 32904076

Abstract

Background and Aims

Helicobacter pylori (H. pylori) eradication has become increasingly unsuccessful due to the prevalence of antibiotic resistance. To address this global issue, a novel strategy for eradication without antibiotics must be developed. The purpose of this study was to examine the effect of methylene blue (MB) with sodium bicarbonate (NaHCO3) on H. pylori using photodynamic antimicrobial chemotherapy.

Materials and Methods

MB was basified using NaHCO3. The basic effect of MB with NaHCO3 was examined using an endoscope equipped with a laser light source. H. pylori was smeared on the culture media with basic MB, followed by illumination at approximately 1,100 lux for 10 and 20 seconds.

After 4 days of culture, the basic effects were determined according to the bacterial growth.

Results

The basic effects of MB appeared at a pH from 8.6 to 9.0 and at NaHCO3 concentrations between 2% and 6.5%. MB concentrations of > 0.05% exhibited the basic effects. The duration of irradiation had no remarkable effects.

Conclusions

Our results showed that the laser endoscope and basic MB were effective for H. pylori eradication.

Keywords: Helicobacter pylori, methylene blue, sodium bicarbonate, basic effect, laser endoscope

Introduction

The bacteria Helicobacter pylori (H. pylori) are gram-negative, flagellated, microaerophilic, and spiral or curved bacilli present in almost all patients with active chronic gastritis, duodenal ulcers and gastric ulcers 1) and H. pylori infects half of the worldwide population and plays a causal role in ulcer diseases and gastric cancer 2).

For eradication, a combination therapy of proton pump inhibitors and antibiotics has been used. However, because of the development of widespread antibiotic resistance, complete eradication is difficult to achieve.

Although the development of antibiotic-independent methods has garnered much attention, it yet has to be established.

One study on the motility of H. pylori in gastric mucin with an acidic or neutral pH and in the absence of urea showed that the bacteria swam freely at a high pH but were strongly constrained at a low pH 3).

To infect the stomach and establish colonies on the mucosal epithelial surface, the bacterium has to move across the gel-like gastric mucus lining under acidic conditions 4).

An acid–base disturbance regulated by an external pH appears to destroy the survival mechanisms of H. pylori.

Photodynamic antimicrobial chemotherapy (PACT), which combines light and a photosensitizing drug, is a potential therapy that promotes a phototoxic effect on the targeted cells by oxidative damage 5).

We developed a basic pH condition by adding sodium bicarbonate (NaHCO3) to methylene blue (MB) and, consequently, devised a new photochemical method using basic MB. We performed PACT and aimed to verify its bactericidal effect on H. pylori using laser endoscope and basic MB.

Materials and Methods

Materials and preparation

We used H. pylori strain (Japan Collection of Microorganisms: No.12093) obtained from the National Research and Development Agency, Institute of Physical and Chemical Research (RIKEN, Japan). H. pylori was identified using the API Campy kit (bioMerieux Japan Ltd., Tokyo, Japan).

The H. pylori colonies were placed in a sterile physiological solution for an adjustment of the count to 2.0 × 108 cells/mL and were stored in a sterile test tube.

We used a 500 mL solution containing 25 g of MB diluted in ethanol (FUJIFILM Wako Pure Chemical Corporation, Japan) as the stock mixture and NaHCO3 powder (Nichi-Iko Pharmaceutical Co., Ltd., Japan). A series of solutions containing MB concentrations of 0.01%, 0.05%, 0.1%, 0.2%, 0.5%, and 1% in sterile water for injection were prepared in sterile test tubes. NaHCO3 solutions at concentrations of 2%, 3%, 4%, 5%, 6%, and 6.5% were made in a similar manner.

Irradiation experiments using a laser endoscope

Using a sterile pipette tip,100 µL of MB, 20 µL of H. pylori, and 10 µL of NaHCO3 were taken out of test tubes and were smeared on a Helicobacter agar medium (Nissui Pharmaceutical Co., Ltd., Japan) using a sterile bacteria spreader. The bacterial smear area on the medium was adjusted to a circle with a diameter of approximately 4 cm. NaHCO3 was diluted 13-fold before being added to the media, to obtain final concentrations of 0.15%, 0.23%, 0,30%, 0.38%, 0.46%, and 0.50%.

After being left in the dark for 5 minutes, the bacterial cultures were irradiated vertically from a distance of 4 cm using an endoscope system (EG-L600ZW7 scope, Processor VP-7000 system, and Light source device LL-7000; FUJIFILM Co., Tokyo, Japan) with lasers (LASEREO®). The light intensity was adjustable from a darker level (−4) to a lighter level (+5).

For this experiment, the light intensity applied was adjusted to the standard level of automatic control, which was in the middle range of the light intensities. Endoscopic irradiation was performed at approximately 1,100 lux using white light and linked color imaging (LCI) for 10 and 20 seconds.

LCI is an image-processing method that can emphasize minute color differences in the gastric mucosa by simultaneously irradiating a narrowband of short wavelength light and white light 6). Compared with white light alone, LCI can make reddish colors appear redder and whitish colors appear whiter, thus enabling an easy detection of the subtle changes in colors 6).

After irradiation, which was defined as both white laser light and LCI, the culture media were placed in jars and were incubated under microaerophilic conditions at 37ºC for 4 days.

To maintain microaerophilic and moist conditions in the jars, an oxygen absorber-carbon dioxide generator (DIA-Microaerophilic Pack; LSI Medience Corporation, Tokyo, Japan) and 20 ml of sterile water for injection were used.

Determination of effectiveness

The bactericidal effect was evaluated according to the H. pylori growth area, which was categorized into six grades. The response was considered ineffective when 1) there was a bactericidal effect in the absence of irradiation and MB; 2) there was a bactericidal effect from irradiation in the absence of MB; or 3) there was a bactericidal effect from NaHCO3 alone, regardless of application of irradiation.

The blank test included the bacteria and each MB concentration with no irradiation.

(−) indicated ineffectiveness.

Compared with the blank test that used an ineffective concentration of 0.01% MB, scores of (1+), (2+), (3+), and (4+) indicated decrease in the growth area by 20%, 40%, 60%, and 80%, respectively.

A score of (5+) indicated no growth area.

Measurements of pH

The pH of MB, NaHCO3, and basic MB solutions were measured. The stock mixture of 0.01% to 1% MB and 2% to 6.5% NaHCO3 were diluted in sterile water in sterile test tubes. The pH values of the 2000 µL MB solution, 400 µL of sterile physiological saline, and 200 µL of NaHCO3 solution that were prepared in sterile test tubes were measured using the Seven Compact instrument (METTLER TOLEDO International Inc.). The concentration of MB was increased from 0.01% to 1% and that of NaHCO3 was increased from 2% to 6.5%; pH was measured for each sample in the series.

Lux measurement of the laser endoscope

Light intensity at the standard level was measured vertically from a distance of 4 cm, using a luminometer (T10-A, KONICA MINOLTA JAPAN, INC.) in the endoscopic room. Lux measurement was performed after adjusting for a room light intensity of 0 lux.

Results

pH of the materials

Table 1 shows the pH values of the MB, NaHCO3, and basic MB solutions.

Table 1: pH of MB According to the NaHCO3 Solutions.

NaHCO3 concentration% MB concentration%
0 0.01 0.05 0.1 0.2 0.5 1
0 7.435 6.997 6.856 6.962 6.505 5.890
2 8.141 8.550 8.574 8.589 8.620 8.708 8.869
3 8.109 8.598 8.612 8.617 8.658 8.748 8.939
4 8.082 8.605 8.619 8.672 8.653 8.782 9.000
5 8.037 8.600 8.617 8.632 8.674 8.785 8.929
6 8.030 8.617 8.624 8.628 8.671 8.777 8.972
6.5 8.045 8.668 8.667 8.701 8.739 8.849 9.000
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MB, methylene blue; NaHCO3, sodium bicarbonate

As the MB concentration increased from 0.01% to 1%, the pH decreased from 7.435 to 5.890. Moreover, as the concentration of NaHCO3 increased from 2% to 6.5%, the pH decreased from 8.141 to 8.045.

After adding NaHCO3 to MB, the pH increased in the range from 8.550 to 9.0.

Effects of basic MB using white light

Table 2 shows the basic effect of MB with NaHCO3 using white light. MB concentrations > 0.5% showed Grade 5+ bactericidal activity in the absence of light. MB concentrations > 0.1% exhibited Grade 5+ bactericidal effect using white light for 10 and 20 seconds. After adding the 2% to 6.5% NaHCO3 concentrations to MB, Grade 5+ bactericidal effect was shown at MB concentrations > 0.05%. The effectiveness did not differ according to the exposure time.

Table 2: Basic Effect of MB with NaHCO3 Irradiated at Approximately 1,100 Lux for 10 and 20 Seconds Using White Light.

NaHCO3 concentration MB conceniration %
% irradiation (sec) 0 0.01 0.05 0.1 0.2 0.5 1
0 10 — (−) (4+) (5+) (5+) (5+) (5+)
20 — (−) (3+) (4+) (5+) (5+) (5+)
2 10 — (4+) (5+) (5+) (5+) (5+) (5+)
20 — (4+) (5+) (5+) (5+) (5+) (5+)
3 10 — (4+) (5+) (5+) (5+) (5+) (5+)
20 — (4+) (5+) (5+) (5+) (5+) (5+)
4 10 — (−) (2+) (5+) (5+) (5+) (5+)
20 — (−) (2+) (5+) (5+) (5+) (5+)
5 10 — (−) (5+) (5+) (5+) (5+) (5+)
20 — (−) (4+) (5+) (5+) (5+) (5+)
6 10 — (4+) (5+) (5+) (5+) (5+) (5+)
20 — (3+) (5+) (5+) (5+) (5+) (5+)
6.5 10 — (4+) (5+) (5+) (5+) (5+) (5+)
20 — (4+) (5+) (5+) (5+) (5+) (5+)
Blank test (no irradiation) — (−) (−) (3+) (4+) (5+) (5+)
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MB, methylene blue; NaHCO3, sodium bicarbonate

These effects on the media are shown in Figures 1 to 3. The basic effect of MB with NaHCO3 was more efficacious, compared with that of MB alone.

Figure 1:

Figure 1:

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No Irradiation.

Figure 3:

Figure 3:

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Basic Effect of Methylene Blue with 5% Sodium Bicarbonate Irradiated at Approximately 1,100 Lux for 10 Seconds Using White Light.

Figure 2:

Figure 2:

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Effect of Methylene Blue Irradiated at Approximately 1,100 Lux for 10 Seconds Using White Light.

Effects of basic MB using LCI

The effect of MB with NaHCO3 using LCI is shown in Table 3. MB concentrations > 0.2% had Grade 5+ bactericidal effect after LCI for 10 and 20 seconds. After adding the 2% to 6.5% NaHCO3 concentrations to MB, Grade 5+ bactericidal effect was achieved at MB concentrations > 0.05%. The duration of irradiation had no effect on the basic effect of Grade 5+.

Table 3: Basic Effect of MB with NaHCO3 Irradiated at Approximately 1,100 Lux for 10 and 20 Seconds Using LCI.

NaHCO3 concentration MB conceniration %
% irradiation (sec) 0 0.01 0.05 0.1 0.2 0.5 1
0 10 — (−) (3+) (4+) (5+) (5+) (5+)
20 — (−) (3+) (4+) (5+) (5+) (5+)
2 10 — (4+) (5+) (5+) (5+) (5+) (5+)
20 — (4+) (5+) (5+) (5+) (5+) (5+)
3 10 — (4+) (5+) (5+) (5+) (5+) (5+)
20 — (4+) (5+) (5+) (5+) (5+) (5+)
4 10 — (−) (5+) (5+) (5+) (5+) (5+)
20 — (−) (5+) (5+) (5+) (5+) (5+)
5 10 — (−) (5+) (5+) (5+) (5+) (5+)
20 — (−) (5+) (5+) (5+) (5+) (5+)
6 10 — (4+) (5+) (5+) (5+) (5+) (5+)
20 — (3+) (5+) (5+) (5+) (5+) (5+)
6.5 10 — (4+) (5+) (5+) (5+) (5+) (5+)
20 — (4+) (5+) (5+) (5+) (5+) (5+)
Blank test (no irradiation) — (−) (−) (3+) (4+) (5+) (5+)
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MB, methylene blue; NaHCO3, sodium bicarbonate; LCI, linked color imaging

Figures 4 and 5 show these effects on the media. At MB concentrations of > 0.05%, the basic effect of MB using white light was consistent with that of LCI for 10 and 20 seconds.

Figure 4:

Figure 4:

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Effect of Methylene Blue Irradiated at Approximately 1,100 Lux for 10 Seconds Using Linked Color Imaging.

Figure 5:

Figure 5:

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Basic Effect of Methylene Blue with 5% Sodium Bicarbonate Irradiated at Approximately 1,100 lux for 10 Seconds Using Linked Color Imaging.

Extent of the basic effect

Figures 6 and 7 demonstrate the relationships among the minimal MB concentration for achieving a Grade 5+ bactericidal effect, NaHCO3 concentration, and pH using white light and LCI.

Figure 6:

Figure 6:

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Basic Effect of Methylene Blue with Sodium Bicarbonate Irradiated at Approximately 1,100 Lux for 10 and 20 Seconds Using White Light.

The basic effect is extensively distributed between 2% and 6.5% of sodium bicarbonate (NaHCO3). The black circle (●) and white circle (○) represent the minimal MB concentrations for achieving Grade 5+ bactericidal effects for 10 and 20 seconds, respectively.

Figure 7:

Figure 7:

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Basic Effect of Methylene Blue with Sodium Bicarbonate Irradiated at Approximately 1,100 Lux for 10 and 20 Seconds Using Linked Color Imaging.

The basic effect is widely present in the range of 2.5% to 6.5% of NaHCO3. The black circle (●) and white circle (○) represent the minimal MB concentrations for achieving Grade 5+ bactericidal effects for 10 and 20 seconds, respectively.

The colored part of the figures represents the region with extensive basic effect. The basic effects of MB were estimated to be present at a pH between 8.6 and 9.0 and at NaHCO3 concentrations between 2% and 6.5%. The actual NaHCO3 concentrations were in the range of 0.15%–0.50%.

Discussion

In this study, laser endoscopic eradication of H. pylori using basic MB with NaHCO3 was highly effective. The combination of acid-base disturbance and PACT is a novel approach to antibiotic-independent eradication of H. pylori.

However, this method cannot be applied to patients who do not consent to endoscopy and may not achieve eradication in one session. In such cases, repeated endoscopic eradication is required.

The mechanism of action of the basic effect was considered to involve the properties of PACT as well as the dye used.

General features of dyes

MB, a widely known histological dye 7), was used as a photosensitizer for PACT. Dyes are generally divided into acidic, basic, and amphoteric types 8). In an aqueous solution, acidic dyes are negatively charged, basic dyes are positively charged, and amphoteric dyes have both features and can maintain an equilibrium state 8). MB is a water-soluble and basic dye, which has shown affinity to most bacteria and an enhanced staining ability in the basic state 8, 9).

Moreover, MB is a phenothiazine dye that has a strong absorbance in the range of 550–700 nm 10), which is included in the broad wavelength range of endoscopic light. Red light with a wavelength of 660 nm corresponds to the absorption spectrum of MB 11).

Therefore, a laser endoscope can function as a new therapeutic approach for H. pylori eradication.

Cell wall and membrane of gram-negative bacteria

Being a microaerophilic gram-negative organism, H. pylori has a cell wall, an outer membrane, a periplasmic space, and an inner or cytoplasmic membrane that separates the cytoplasm from the medium 12).

The outer membrane of gram-negative organisms is a phospholipid bilayer that contains a variety of proteins; some exhibit membrane spanning, whereas others are associated with only one face of the bilayer as well as lipopolysaccharides 13).

To exert bactericidal activity, MB has to be extensively distributed within the cell. Notably, MB has no difficulty in crossing bacterial cell walls 14); because of its cationic charge, it can bind to the negative charge of the lipopolysaccharides of gram-negative bacteria 14). Moreover, even without light, MB has been reported to show natural antifungal and antibacterial activities 14).

The results of this study implied that the natural antibacterial activity of MB was enhanced through the basic effect and in the presence of a laser light source.

Mechanisms of pH homeostasis

(1). Extent of pH for survival and growth

H. pylori bacteria reside deep in the gastric mucus layer; this gastric Helicobacter species has been shown to utilize the mucus pH gradient for precise spatial orientation 15).

After colonizing the human stomach, H. pylori can encounter a number of environmental stress inducers; one of the most obvious factors is the drastic fluctuation in stomach pH 16).

Nevertheless, H. pylori has evolved to acquire a desirable homeostasis for survival under acid–base conditions. Urease-dependent elevation of local pH to nontoxic levels has been thought to allow H. pylori survival in vivo 17).

Survival of H. pylori has been demonstrated at a pH range of 4.0 to 8.0, whereas its protein synthesis occurred at a pH between 6.0 and 8.0 12). The major energy source for H. pylori is glucose, which is converted into metabolic acids 17). The pH range for H. pylori metabolism was shown to be 3.5–8.6; outside these limits, there was an irreversible loss of metabolic capacity 17).

These survival mechanisms were considered a wide range of acid–base adaptation.

(2). Proton motive force

The proton motive force (PMF) allows aerobic bacteria to survive by functioning as an electrochemical gradient for protons across the plasma membrane 18) and as a means for conversion of the energy of substrate metabolism to ATP synthesis 19)

The PMF can be maintained at a relatively constant pH of 4.0 to 8.0, mostly by an alteration of membrane potential; beyond these pH limits, the PMF was shown to decay irreversibly, which correlated with the survival of organism 18).

Figure 8 depicts this concept of acid-base disturbance and pH homeostasis of H. pylori. The regulation of pH homeostasis is critically involved in the H. pylori survival and growth.

Figure 8:

Figure 8:

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pH Range for the Survival and Metabolism of H. pylori.

H. pylori responds to extensive pH fluctuations to maintain survival homeostasis.

(3). Effect of pH fluctuation

Generation of pH > 8.6 or < 3.5 exceeds the desirable range for survival, metabolism, protein synthesis, and PMF integrity.

H. pylori encounters basic fluctuation as the pH rises to > 8.6, whereas it falls into acidic fluctuation as the pH declines to < 3.5. Therefore, H. pylori eradication is strongly influenced by the pH fluctuations.

Photochemical pathway underlying the basic effect

In this study, the addition of NaHCO3 to MB further increased the basic effect in the presence of light.

It is particularly important that PACT is conducted under basic pH conditions. It is estimated that the basic effect is exerted when both the adjusted NaHCO3 concentration and the basic pH are optimized.

A photochemical pathway underlying the basic effect seemed to have been present and could have altered the role of basic MB from staining to bactericidal therapy.

PACT Mechanisms

The mechanism of PACT results from the interaction of photons of visible light with photosensitizers 20). Following the absorption of a light photon of specific wavelength, the photosensitizer is promoted to a singlet state; thereafter, it is converted to the triplet state, which has a lower energy and longer lifetime relative to the singlet state 20).The excited triplet state of the photosensitizer can generate the production of reactive species, either by Type I or Type II photochemical mechanisms 20, 21, 22); the former is caused by an electron transfer between the triplet photosensitizer and biomolecules, and it leads to the formation of reactive oxygen species (ROS) 20, 21, 22), whereas the latter is caused by energy transfer to molecular oxygen, which generates singlet oxygen (1O2) 20, 21, 22). These cytotoxic ROS and 1O2 induce lethal cell damage.

Mucus dissolution under basic pH conditions

Dissolving the gastric mucus and destroying the external environment are necessary prior to eradication. H. pylori uses the gastric mucus pH gradient for chemotactic orientation 15). The proteolytic enzyme (PRONASE®MS; Kaken Pharmaceutical Co., Ltd. Tokyo, Japan), which is used as a pretreatment for endoscopic examination, is unstable under an acidic state, and exhibits an enzyme activity at a pH ranging from 7 to 10 23).

This is the reason we cannot eradicate H. pylori in an acidic state of pH < 3.5. Therefore, H. pylori should be eradicated in a basic state.

Moreover, basic pH conditions can enhance the staining ability of MB and simultaneously optimize the pH of the proteolytic enzyme. At a pH > 8.6, H. pylori encounters a lethal pH crisis and is unable to maintain the mechanisms of survival and growth.

Endoscopic eradication using basic MB

As pretreatment during chromoendoscopy, NaHCO3, proteolytic enzyme, and a gastro intestinal antiflatulent agent (GASCON®; Kissei Pharmaceutical Co., Ltd., Tokyo, Japan) 24) are orally administered in the form of aqueous solutions, followed by an endoscopic examination after15 to 30 minutes 25).

When sprayed onto the stomach, MB will not stain normal gastric mucosa, but it will be absorbed by the intestinal epithelium in the presence of intestinal metaplasia 26).

After sufficiently suctioning the dissolved mucus using endoscopic forceps, basic MB is sprayed throughout the gastric mucosa. NaHCO3 is administered twice to achieve a basic pH to optimize the pH of the proteolytic enzyme and to enhance MB staining ability. Exposure to changes in external pH and mucus would disrupt the survival orientation of H. pylori.

Distribution of H. pylori in the stomach

H. pylori bacteria colonize and typically infect the antral mucosa of the human stomach, but acid suppressive therapy was shown to change the predominance of the infection to the gastric body 27, 28).

After 4 weeks of omeprazole treatment, the histological density of H. pylori was reported to decrease in the antrum and corpus, but it increased in the fundus; this migration of H. pylori from the antrum to the fundus was associated with a corresponding decrease in the activity of antral gastritis 29). Moreover, H. pylori infection was reported to be a significant risk factor for the development of atrophic gastritis and intestinal metaplasia 30). Accordingly, H. pylori eradication can also suppress the progression of these diseases.

Cmparison of endoscopic images

Figure 9 shows the mucosal color differences in atrophic gastritis between white light imaging (WLI) and LCI. Compared with WLI, LCI was able to emphasize the minute color changes in the gastric mucosa. The WLI function can be easily converted to an LCI function by a manual push button on the top of the endoscope. The use of LCI permitted endoscopic eradication of H. pylori, while observing the minute mucosal findings from the infection.

Figure 9:

Figure 9:

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Comparison of the Laser Endoscopic Images of the Gastric Antrum in the Antegrade View. Compared with (A) WLI, (B) LCI highlights the mucosal color changes in atrophic gastritis infected with H. pylori.

WLI, white light imaging; LCI, linked color imaging

Figure 10 schematically illustrates the mechanisms of laser endoscopic eradication.

Figure 10:

Figure 10:

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Schema of Laser Endoscopic Eradication of H. pylori in the Stomach.

An acid–base disturbance at a pH > 8.6 is a lethal crisis for survival and growth of H. pylori. The basic effect of methylene blue (MB) with sodium bicarbonate (NaHCO3) is caused by reactive oxygen species (ROS) and singlet oxygen (1O2), which are generated through a photochemical activation by endoscopic light. Both ROS and 1O2 induce cell death.

Considering the spatial distribution of H. pylori in the stomach, laser endoscopic irradiation needs to be extended entirely from the antrum and corpus to the fundus.

A 10-second irradiation on one site of the stomach is conducted multiple times to irradiate the entire stomach.

Stepwise eradication following the generation and maintenance of a basic pH constitutes this photochemical method. PACT holds the potential to improve a successful H. pylori eradication.

Conclusions

An acid–base disturbance may be a new therapeutic strategy for H. pylori eradication, without producing resistant bacteria. Future human clinical trials including cases of eradication failure are required to verify our results. We believe that laser endoscopic eradication using basic MB can provide a promising technology in clinical practice.

Acknowledgments

This paper was based on a lecture at “The 29th Annual Meeting of The Japan Photodynamic Association” held on September 19, 2019 in Tokyo.

References

  • 1: Marshall BJ, Warren JR: Unidentified curved bacilli in the stomach of patients with gastritis and peptic ulceration. Lancet. 1984. June 16;1(8390):1311-5. [DOI] [PubMed] [Google Scholar]
  • 2: Sachs G, Scott DR, Wen Y: Gastric infection by Helicobacter pylori: Curr Gastroenterol Rep. 2011. December;13(6):540-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 3: Celli JP, Turner BS, Afdhal NH, Keates S, Ghiran I, Kelly CP, Ewoldt RH, McKinley GH, So P, Erramilli S, Bansil R: Helicobacter pylori moves through mucus by reducing mucin viscoelasticity. Proc Nat l Acad Sci USA. 2009. August 25;106(34):14321-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 4: Bansil R, Celli JP, Hardcastle JM, Turner BS: The Influence of Mucus Microstructure and Rheology in Helicobacter pylori infection. Front Immunol. 2013. October 10; 4:310. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 5: Giroldo LM, Felipe MP, de Oliveira MA, Munin E, Alves LP, Costa MS: Photodynamic antimicrobial chemotherapy (PACT) with methylene blue increases membrane permeability in Candida albicans. Lasers Med Sci. 2009. January;24(1):109-12. [DOI] [PubMed] [Google Scholar]
  • 6: Nishida H, Ozawa S, Shimomura K, Abe S, Yoshida K, Ohashi E, Mishima T: Development of a New Endoscopic Platform. FUJIFILM RESEARCH & DEVELOPMENT. 2017; 62:1-7. [Google Scholar]
  • 7: Tardivo JP, Del Giglio A, de Oliveira CS, Gabrielli DS, Junqueira HC, Tada DB, Severino D, de Fatima Turchiello R, Baptista MS: Methylene blue in photodynamic therapy: From basic mechanisms to clinical applications. Photodiagnosis Photodyn Ther. 2005. September;2(3):175-91. [DOI] [PubMed] [Google Scholar]
  • 8: Sano Y: Histological Techniques-Theoretical and Applied-. NANZANDO Co., Ltd. Japan: 1981;223-6. [Google Scholar]
  • 9: Mizuguchi K, Ito K, Shitara M, Higashi K, Hiroi S, Fujita K: The newest complete guide to staining techniques Medical Technology Extra Issue. Ishiyaku Publishers Inc., Japan: 2011;88. [Google Scholar]
  • 10: DeRosa Maria C., Crutchley Robert J.: Photosensitized singlet oxygen and its applications. Coordination Chemistry Reviews. 2002. November;233–234:351-71. [Google Scholar]
  • 11: Sousa João Nilton Lopes de, et al. : Photoinactivation of Candida albicans using methylene blue as photosensitizer. RGO, Rev. Gaúch. Odontol. 2015. December;63(4):411-7. [Google Scholar]
  • 12: Scott DR, Weeks D, Hong C, Postius S, Melchers K, Sachs G: The Role of Internal Urease in Acid Resistance of Helicobacter pylori. Gastroenterology. 1998. January;114(1):58-70. [DOI] [PubMed] [Google Scholar]
  • 13: Scott D, Weeks D, Melchers K, Sachs G: The life and death of Helicobacter pylori. Gut. 1998. July;43(Suppl 1): S56-60. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 14: Peloi LS, Soares RR, Biondo CE, Souza VR, Hioka N, Kimura E: Photodynamic effect of light-emitting diode light on cell growth inhibition induced by methylene blue. J. Biosci. 2008. June;33(2):231-7. [DOI] [PubMed] [Google Scholar]
  • 15: Schreiber S, Konradt M, Groll C, Scheid P, Hanauer G, Werling HO, Josenhans C, Suerbaum S: The spatial orientation of Helicobacter pylori in the gastric mucus. Proc Natl Acad Sci. USA. 2004. April 6:101(14): 5024-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 16: Merrell DS, Goodrich ML, Oz tto G, Tompkins LS, Falkow S: pH-Regulated Gene Expression of the Gastric Pathogen Helicobacter pylori. Infect Immun. 2003. June;71(6):3529-39. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 17: Rektorschek M, Weeks D, Sachs G, Melchers K. Influence of pH on Metabolism and Urease Activity of Helicobacter pylori. Gsstroenterology. 1998. September;115(3):628-41. [DOI] [PubMed] [Google Scholar]
  • 18: Meyer-Rosberg K, Scott DR, Rex D, Melchers K, Sachs G: The Effect of Environmental pH on the Proton Motive Force of Helicobacter pylori. Gastroenterology. 1996. October;111(4):886-900. [DOI] [PubMed] [Google Scholar]
  • 19: Sachs G, Meyer-Rosberg K, Scott DR, Melchers K: Acid, protons and Helicobacter pylori. Yale J Biol Med. 1996. May-Jun;69(3):301-16. [PMC free article] [PubMed] [Google Scholar]
  • 20: Donnelly RF, McCarron PA, Tunney MF: Antifungal photodynamic therapy. Microbiological Research. 2008;163(1):1-12. [DOI] [PubMed] [Google Scholar]
  • 21: Bacellar IO, Pavani C, Sales EM, Itri R, Wainwright M, Baptista MS. Membrane Damage Efficiency of Phenothiazinium Photosensitizers. Photochemistry and Photobiology. 2014. Jul-Aug;90(4):801-3. [DOI] [PubMed] [Google Scholar]
  • 22: Gabrielli D, Belisle E, Severino D, Kowaltowski AJ, Baptista MS: Binding, aggregation and photochemical properties of methylene blue in mitochondrial suspensions. Photochemistry and Photobiology. 2004. March;79(3):227-32. [DOI] [PubMed] [Google Scholar]
  • 23: Pharmaceutical Products Interview Form: PRONASE® MS the third edition. Kaken Pharmaceutical Co., Ltd. Japan: 2010:2-6. [Google Scholar]
  • 24: Pharmaceutical Products Interview Form: GASCON® the second edition. Kissei Pharmaceutical Co., Ltd. Japan: 2012;8-14. [Google Scholar]
  • 25: Japan Gastroenterological Endoscopy Society: Gastroenterological Endoscopy Handbook. Japan Medical Center, Japan: 2012:228-9. [Google Scholar]
  • 26: Kanai M, et al. : Kanai's Manual Clinical Laboratory Medicine. 32nd edition: Kanehara & Co., Ltd., Japan: 2005:1320-1. [Google Scholar]
  • 27: Mollenhauer-Rektorschhek M, Hanauer G, Sachs G, Melchers K: Expression of UreI is required for intragastric transit and colonization of gerbil gastric mucosa by Helicobacter pylori. Res Microbiol. 2002. December; 153(10):659-66. [DOI] [PubMed] [Google Scholar]
  • 28: Marcus EA, Sachs G, Scott DR: Eradication of Helicobacter pylori Infection. Curr Gastroenterol Rep. 2016. July;18(7):33. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 29: Logan RP, Walker MM, Misiewicz JJ, Gummett PA, Karim QN, Baron JH: Changes in the intragastric distribution of Helicobacter pylori during treatment with omeprazole. Gut. 1995. January;36(1):12-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 30: Kuipers EJ, Uyterlinde AM, Peña AS, Roosendaal R, Pals G, Nelis GF, Festen HP, Meuwissen SG: Long-term sequelae of Helicobacter pylori gastritis. Lancet. 1995. June 17;345(8964):1525-8. [DOI] [PubMed] [Google Scholar]

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