Characterisation of a sucrose-independent in vitro biofilm model of supragingival plaque.

OBJECTIVES: Sugar consumption has been decreasing in Japan, suggesting higher rates of sucrose-independent supragingival plaque formation. For developing an in vitro biofilm model of sucrose-independent supragingival plaque, this study aimed to investigate the compositions and functions on contributing to cariogenicity in comparison with sucrose-dependent biofilm. MATERIALS AND METHODS: An in vitro multispecies biofilm containing Actinomyces naeslundii, Streptococcus gordonii, S. mutans, Veillonella parvula and Fusobacterium nucleatum was formed on 24-well plates in the absence or presence of 1% sucrose. Compositions were assessed by plate culture, scanning electron microscopy and confocal laser scanning microscopy after fluorescent in situ hybridisation or labelling of extracellular polymeric substances (EPS). Functions were assessed by acidogenicity, adherence strength and sensitivities to anticaries agents. RESULTS: Although both biofilms exhibited a Streptococcus predominant bacterial composition, there were differences in bacterial and EPS compositions; in particular, little glucan EPS was observed in sucrose-independent biofilm. Compared with sucrose-dependent biofilm, acidogenicity, adherence strength and antimicrobial resistance of sucrose-independent biofilm were only slightly lower. However, dextranase degradation was substantially lower in sucrose-independent biofilm. CONCLUSION: Our findings suggest that sucrose-independent biofilm may have cariogenicity as with sucrose-dependent biofilm. These in vitro models can help further elucidate plaque-induced caries aetiology and develop new anticaries agents.

Decreased expression of E-cadherin by Porphyromonas gingivalis-lipopolysaccharide attenuates epithelial barrier function.

BACKGROUND AND OBJECTIVE: The gingival epithelium is a first line of defense against bacterial challenge. E-cadherin (E-cad) plays an important role in cell-cell adhesion as a barrier in the epithelium. Recently, a decrease in the expression of E-cad has been observed in inflamed gingival tissue. The aims of this study were to clarify the changes in E-cad expression and barrier function in human gingival epithelial cells stimulated with Porphyromonas gingivalis-lipopolysaccharide (P. gingivalis-LPS) and to evaluate the influence of these changes on the inflammatory reaction. Furthermore, to clarify the mechanism of the E-cad changes induced by P. gingivalis-LPS, we focused on reactive oxygen species (ROS) that are reported to induce a decrease in E-cad expression. MATERIAL AND METHODS: Human gingival epithelial cells were incubated in Humedia-KG2 in the presence or absence of P. gingivalis-LPS and antioxidants to analyze ROS involvement in P. gingivalis-LPS-induced E-cad changes. E-cad protein expression was analyzed by immunofluorescence staining. To investigate barrier function and inflammatory changes, we performed transport and cytokine assays using gingival epithelial cells and macrophages co-culture model in transwell plates. Medium containing 10 µg/mL P. gingivalis-LPS (transport substance) was added to the upper compartment, which harvested gingival epithelial cells, and medium without P. gingivalis-LPS was added to the lower compartment, which harvested macrophages. In the transport assay, P. gingivalis-LPS penetration was analyzed using the Limulus amebocyte lysate test. In the cytokine assay, we examined the change in tumor necrosis factor-α (TNF-α) production from the macrophages in the lower compartment using enzyme-linked immunosorbent assay. RESULTS: Expression of E-cad in human gingival epithelial cells was decreased by P. gingivalis-LPS, and the decrease in E-cad accelerated the penetration of P. gingivalis-LPS through the monolayer. In addition, the concentration of TNF-α was higher under the E-cad reduced monolayer. Antioxidants, particularly vitamin E and l-ascorbic acid 2-phosphate magnesium salt, inhibited the decrease in E-cad expression, penetration of P. gingivalis-LPS and increase in TNF-α. CONCLUSION: These results suggest that the decrease in E-cad caused by P. gingivalis-LPS leads to destruction of the epithelial barrier function in human gingival epithelial cells, and finally accelerates the inflammatory reaction under the barrier. Antioxidants, particularly vitamin E and l-ascorbic acid 2-phosphate magnesium salt, may restore the impaired function by scavenging ROS, which are related to the decrease in E-cad expression by P. gingivalis-LPS.

A-type K+ current mediated by the Kv4 channel regulates the generation of action potential in developing cerebellar granule cells.

During neuronal differentiation and maturation, electrical excitability is essential for proper gene expression and the formation of synapses. The expression of ion channels is crucial for this process; in particular, voltage-gated K(+) channels function as the key determinants of membrane excitability. Previously, we reported that the A-type K(+) current (I(A)) and Kv4.2 K(+) channel subunit expression increased in cultured cerebellar granule cells with time. To examine the correlation between ion currents and the action potential, in the present study, we measured developmental changes of action potentials in cultured granule cells using the whole-cell patch-clamp method. In addition to an observed increment of I(A), we found that the Na(+) current also increased during development. The increase in both currents was accompanied by a change in the membrane excitability from the nonspiking type to the repetitive firing type. Next, to elucidate whether Kv4.2 is responsible for the I(A) and to assess the effect of Kv4 subunits on action potential waveform, we transfected a cDNA encoding a dominant-negative mutant Kv4.2 (Kv4.2dn) into cultured cells. Expression of Kv4.2dn resulted in the elimination of I(A) in the granule cells. This result demonstrates that members of the Kv4 subfamily are responsible for the I(A) in developing granule cells. Moreover, elimination of I(A) resulted in shortening of latency before the first spike generation. In contrast, expression of wild-type Kv4.2 resulted in a delay in latency. This indicates that appearance of I(A) is critically required for suppression of the excitability of granule cells during their maturation.

Heterogeneity of beta-conglycinin.

Beta-conglycinin, a major 7 S soybean globulin, purified by ion-exchange and gel chromatography was fractionated into six distinct components on columns of DEAE-Sephadex. The six components (designated B1 to B6-conglycinins) were characterized by disc electrophoresis. Gel electrophoresis and gel electrofocusing in dissociating buffers indicate that the six conglycinins are isomers containing varying proportions of three kinds of subunits (alpha, alpha' and beta). The subunit structures of these isomers are alpha' beta (B1-), alpha beta (B2-), alpha alpha' beta (B3-), alpha beta (B4-), alpha alpha' (B5-), and alpha (B6-conglycinin). Beta subunit is a major constitutent of B1- and B2-conglycinins, whereas B3- to B6-conglycinins are composed predominantly of alpha subunit. The six beta-conglycinins are all glycoproteins containing mannose and glucosamine. They differ in the N-terminal amino acid composition. The isolated B1- to B4-conglycinins are immunologically identical with one another and with the total beta-conglycinin. B5- and Bl-conglycinins which comprise no beta subunit are partially identical with the total protein. Some antigenic determinants that are lacking in the B5- and B6-conglycinins are expected to be located on the beta subunit.

Beta-conglycinin from soybean proteins. Isolation and immunological and physicochemical properties of the monomeric forms.

Beta-conglycinin consisting of six major isomers (designated B1- to B6-conglycinin) was dissociated and fractionated on columns of DEAE- and CM-Sephadex in buffers containing 6 M urea. Three major (alpha, alpha' and beta) and one minor (gamma) subunits were isolated and further characterized by gel electrophoresis and gel electrofocusing. Gel electrophoresis in urea and in sodium dodecyl sulfate, and gel filtration in 6 M guanidine hydrochloride gave a molecular weight of 57 000 for alpha, alpha' subunits; and 42 000 for beta and gamma subunits. The isoelectric points of the isolated subunits, measured by disc gel electrofocusing, were as follows: alpha, 4.90; alpha', 5.18; beta, 5.66-6.00. On gel electrofocusing, beta subunit showed four microheterogeneous components; three of them comprised 95% of the total beta subunit. Leucine and valine were the N-terminal amino acids of beta and alpha alpha' subunits, respectively. The isolated subunits contained mannose and glucosamine in varying quantities. Two carbohydrate moieties were calculated for one mole of alpha, alpha' subunits; and one carbohydrate moiety for the beta subunit. Considerable similarity in the amino acid composition of alpha and alpha' subunits was observed. The beta subunit was devoid of cysteine and methionine; and in comparison with alpha, alpha' subunits, had a higher content of hydrophobic amino acids. The isolated subunits exhibited antigen-antibody reaction with antisera to the native beta-conglycinin. Each of them was partglycinins. The alpha and alpha' subunits were in addition identical with each other and with B5-, B6-conglycinins. They were immunologically unrelated with beta subunit. The recovery of immuno-properties from the individual subunits may be attributed to the reconstruction of the three-dimensional structure upon removal of denaturing reagents.

Colonic carcinoma resembling submucosal tumor: report of a case and review of the literature.

Submucosal tumor-like colorectal carcinoma, most of whose surface is covered with normal mucosa, is very rare. We report a case of colonic carcinoma resembling submucosal tumor. A 54-yr-old man visited our institution for an evaluation of a positive fecal occult blood test. Colonoscopic examination revealed a small, mainly red polypoid lesion with a central deep ulceration and many white spots in the sigmoid colon. Indigocarmine staining demonstrated that the white spots were faint shallow depressions. Magnifying colonoscopic examination showed that the lesion surface, except for the ulceration and the depressions, was covered with normal mucosa. Although the tumor was small, we strongly suspected its malignancy due to a deep ulceration. As we could not excise it endoscopically, we performed sigmoidectomy. The lesion was 12 mm in size. Histologic examination revealed that the lesion was a moderately differentiated adenocarcinoma that was mainly covered with normal mucosa, that carcinoma was exposed only at the ulceration and the depressions on the surface, and that it had expanded to the muscularis propria. Together with considerations from the literature, this type of colorectal carcinoma is supposed to be invasive and surgical resection should be considered, no matter how small it may be.

Effect of interferon on incidence of hepatocellular carcinoma in patients with chronic hepatitis C.

BACKGROUND/AIMS: The aim of this study was to evaluate whether IFN prevents the development of HCC in patients with chronic hepatitis C. METHODOLOGY: 103 patients with chronic hepatitis C received IFN and 30 control patients were enrolled in this study. RESULTS: In 33 patients (32.0%) who received IFN, alanine aminotransferase (ALT) decreased to normal range and HCV RNA became negative (complete response: CR). In 7 patients (6.7%), ALT decreased to less than 50 IU/L or stayed within the normal range, but HCV RNA remained positive (biochemical response: BR). In 63 patients (61.1%) and 30 control patients, ALT did not change and HCV RNA remained positive (no response: NR). HCC developed in 5 (4.9%) of the 103 patients who received IFN and 7 (23.3%) of the control patients (p<0.01). In 5 patients who developed HCC, the response to IFN was NR and no HCC developed in patients with CR or BR. In addition, 5-year cumulative rate of development of HCC in 63 IFN NR patients and in control was 7.9% and 23.3% (p<0.05). CONCLUSIONS: IFN decreased the development of HCC in not only patients with CR or BR but also patients with NR.

Pharmacological properties of YM-26365, a low molecular weight, orally active renin inhibitor.

This report describes the pharmacological properties of a novel renin inhibitor (YM-26365: (3R)-3-[3-[(1S)-1-cyclohexylmethyl-2-hydroxy-3- [(1-methyl-5-tetrazolyl)thio]propyl]ureido]-1-methyl-5-phenyl- 2,3-dihydro-1H-1,4-benzodiazepin-2-one) with molecular weight 577 and no peptide bonds. YM-26365 inhibited human plasma renin with an IC50 value of 2.9 x 10(-6) M, but did not affect plasma renin from dogs, rabbits, and rats at 10(-4) M. YM-26365 inhibited not only human renin, but also cathepsin D with an IC50 value of 1.7 x 10(-5) M. This compound competitively inhibited the reaction between recombinant human renin and N-acetyl tetradecapeptide with a Ki value of 1.1 x 10(-6) M. In pithed spontaneously hypertensive rats, YM-26365 at 10 mg/kg i.v. significantly antagonized the pressor response to recombinant human renin, but did not affect responses to angiotensin II, angiotensin I, norepinephrine, or arginine vasopressin. Similarly, oral administration of YM-26365 (10 and 30 mg/kg) to pithed spontaneously hypertensive rats caused a shift to the right of the recombinant human renin dose-pressor response curve. Systemic bioavailability as determined on the basis of the ratio of the total area under the plasma concentration-time curve after 3 mg/kg i.v. and 30 mg/kg orally to rats was 9.6%. These results demonstrate that YM-26365 is a weak but orally absorbed, low molecular weight renin inhibitor.

Ibotenic acid-induced lesions of the medial septum increase hippocampal membrane associated protein kinase c activity and reduce acetylcholine synthesis: prevention by a phosphatidylcholine/vitamin B12 diet.

Ibotenic acid infusion into the medial septum (MS) results in biochemical alterations in the hippocampus. The biochemical events involved in this neuronal lesion are poorly understood. We investigated the effect of a purified diet supplemented with egg phosphatidylcholine (PC) and vitamin B(12) on ibotenic acid-medicated biochemical changes in the rat hippocampus and crude synaptosomal membranes. Male Wistar rats with this MS lesion were fed a purified diet (control diet) or a purified diet supplemented with 5.7 g PC and 125 microg vitamin B(12) per 100 g (experimental diet) for 18 days. Sham-operated rats were fed the control diet. Compared with the sham-operated rats, MS-lesioned rats fed the control diet showed increased activity of membrane-bound protein kinase C (PKC), decreased activity of choline acetyltransferase, and decreased concentrations of acetylcholine in the hippocampus. The ratio of cholesterol to phospholipid in the crude synaptic membrane was lower in the lesioned rats than in the sham-operated rats, but this was not accompanied by any alteration in membrane lipid fluidity. MS-lesioned rats fed the experimental diet showed lowered PKC activity and elevated acetylcholine concentrations than did rats fed the control diet, but there were no significant effects on choline acetyltransferase activity and the lipid ratio. The ibotenic acid-mediated elevation of PKC activity was observed as early as 2 days postinjury in the control diet-fed rats but not in the experimental diet-fed rats. We propose that ibotenic acid mediates pathophysiologic actions through the activation of PKC and that PC combined with vitamin B(12) ameliorates the second messenger-mediated injury.

The role of extracellular Ca2+ in carbachol-induced tonic contraction of the pig detrusor smooth muscle.

The role of extracellular Ca2+ in the tonic-contractile response to muscarinic receptor stimulation was investigated in isolated detrusor smooth muscle from the pig urinary bladder. Carbachol (10(-8)-10(-5) M) produced a concentration-dependent contractile response in isolated pig detrusor smooth muscle strips consisting of an initial phasic component followed by a tonic component. During the plateau of the tonic contractions induced by carbachol at the submaximal concentration of 10(-6) M, the inhibiting effects of atropine, EGTA, nifedipine (a voltage-dependent calcium channel antagonist), H-7 [a protein kinase C (PKC) inhibitor] and YM934 (a potassium channel opener) on the contractions were evaluated. Atropine (10(-10)-3 x 10(-8) M) concentration-dependently inhibited the tonic contractions induced by carbachol. In the same experimental conditions, EGTA (4 mM) and nifedipine (10(-9)-3 x 10(-7) M) depressed the tonic contractions in a concentration-dependent manner as did H-7 (10(-5)-3 x 10(-5) M) and YM934 (10(-8)-10(-6) M). However, H-7 (10(-5)-3 x 10(-5) M) and YM934 (10(-6) M) were very weak in inhibiting the contractions induced by KCl (50 mM) in isolated pig detrusor smooth muscle strips. These results suggest that the tonic-contractile response induced by carbachol in pig detrusor smooth muscle strips is dependent mainly on depolarization of the cell membranes and an influx of extracellular Ca2+, and also suggest that this depolarizing response may be due to inactivation of ATP-sensitive potassium channels through muscarinic activation of PKC.

Expansive laminoplasty with reattachment of spinous process and extensor musculature for cervical myelopathy.

Since 1986 we have performed expansive laminoplasty with reattachment of the spinous processes and extensor musculature in cases of cervical myelopathy, to avoid the late postoperative complications of extensive laminectomy. The operative procedure and results are given in detail. Forty cases (24 men, 16 women) were followed for a mean of 28 months. Postoperative results were satisfactory, with no major complications according to the evaluation criteria of the Japanese Orthopaedic Association. No instability or malalignment was seen on postoperative radiographs.

Thoracic disc herniation. Surgical treatment in 23 patients.

The authors' surgical procedure for the treatment of symptomatic thoracic disc herniation has consisted of total discectomy of the involved intervertebral disc, followed by intervertebral body fusion using autogenous iliac bone through an anterior approach. The authors prefer the extrapleural approach to the thoracic vertebrae because of simplified postoperative care with minimum surgical interference with respiratory function. During the past 16 years, 23 patients with symptomatic thoracic disc herniation have been treated by this procedure. Preoperatively all 23 patients presented with some measure of paraplegia, with difficulty in walking. Their clinical presentation was analyzed and diagnostic aids to the thoracic disc herniation by radiographic examinations were assessed. The radiographic abnormality of ossification of the yellow ligament (OYL) coinciding with the involved thoracic disc level was noted. The results in all 23 patients with this procedure were favorable. Based on the results of this series, this procedure is recommended for the treatment of symptomatic thoracic disc herniation.

Inhibition of experimental bone resorption and osteoclast formation and survival by 2-aminoethanesulphonic acid.

It is known that bone resorption is mediated by osteoclasts, and lipopolysaccharide (LPS) and inflammatory mediators such as interleukin-1 (IL-1) and prostaglandin E2 (PGE2) induce osteoclast differentiation from haemopoietic cells, 2-aminoethanesulphonic acid, which is known as taurine, is an important nutrient and is added to most synthetic human infant milk formulas. In this study, it was found that 2-aminoethanesulphonic acid inhibits the stimulation of bone resorption mediated by LPS of the periodontopathic microorganism Actinobacillus actinomycetemcomitans Y4 in organ cultures of newborn mouse calvaria. The effect of 2-aminoethanesulphonic acid on the development and survival of osteoclast-like multinucleated cells produced in a mouse bone-marrow culture system was also examined. 2-aminoethanesulphonic acid (100 microg/ml) suppressed the formation of these osteoclast-like cells in the presence of LPS of A. actinomycetemcomitans Y4, IL-1alpha or PGE2 in mouse marrow cultures. On the other hand, 2-aminoethanesulphonic acid did not inhibit 1alpha, 25-dihydroxyvitamin D3-mediated osteoclast differentiation. Although IL-1alpha elongated the survival of the osteoclast-like cells, 2-aminoethanesulphonic acid blocked the supportive effect of IL-1alpha on osteoclast survival. 2-aminoethanesulphonic acid showed no effect on the growth of mouse osteoblasts. Finally, it was found that 2-aminoethanesulphonic acid inhibited alveolar bone resorption in experimental periodontitis in hamsters. These results suggest that 2-aminoethanesulphonic acid is an effective agent in preventing inflammatory bone resorption in periodontal diseases.

Serum des-gamma-carboxyprothrombin level by a modified enzyme immunoassay method in hepatocellular carcinoma: clinical significance in small hepatocellular carcinoma.

BACKGROUND/AIMS: In the diagnosis of hepatocellular carcinoma (HCC), serum des-gamma-carboxyprothrombin (DCP) is a useful tumor marker. The conventional immunoassays for measurement of DCP levels, however, are not sensitive enough to detect small HCC. Therefore, we intended to elevate the minimal detection level of DCP by a modified enzyme linked immunosorbent assay method. METHODOLOGY: This modified assay method is similar to the conventional enzyme linked immunosorbent assay (ELISA) method, but the first reaction occurs overnight. As a result, the minimal detection levels of DCP varied from 0.06 AU/ml, by the conventional method, to 0.008 AU/ml, by the modified method. Two hundred and twenty five serum samples from 100 patients with HCC, 75 with liver cirrhosis and 50 with chronic hepatitis were subjected to the present study. Simultaneous determinations of serum DCP by the modified assay and a-fetoprotein (AFP) levels were performed. RESULTS: Eighty five of 100 patients with HCC had increased DCP levels of more than 0.008 AU/ml. This method yielded a sensitivity of 85%, a specificity of 90% and a total accuracy value of 88%. In 27 patients with small HCC (less than 30 mm in diameter), 12 had elevated DCP levels, resulting in a sensitivity of 44%. When the modified DCP assay together with AFP measurement (more than 20 ng/ml) were introduced, the sensitivity was 67% in the 27 patients with small HCC. CONCLUSIONS: This modified ELISA method increased the sensitivity in patients with small HCC, and the combination assay of serum DCP and AFP levels was more useful for the early diagnosis of HCC.

Primary duodenal MALT lymphoma.

Primary duodenal MALT lymphoma (MALToma) is a very rare neoplasm arising from the mucosa-associated lymphoid tissue of the duodenum. We report a 55-year-old woman with MALToma located in the descending duodenum and accompanying Helicobacter pylori infection of the stomach. We performed operative resection due to involvement of the papilla of Vater and submucosal tumor infiltration. Despite wide mucosal spreading, postoperative examination revealed only a small amount of MALToma cells infiltrating into the submucosa. No invasion into the adjacent structure or metastasis to regional lymph nodes was confirmed, suggesting the disease could have been controlled by eradication of Helicobacter pylori.

Therapeutic effects of tiopronin on chronic hepatitis: a double-blind clinical study.

The efficacy of tiopronin (2-mercaptopropionyl glycine) 600 mg daily in the treatment of chronic active or chronic persistent hepatitis has been assessed in a double-blind controlled clinical trial of 12 weeks involving 165 Japanese patients with histologically proven disease. Treatment with the drug was associated with a significant improvement in abnormalities of serum transaminase and gamma glutamyl transpeptidase, with reversion towards baseline values on stopping the drug. Improvement was independent of the histological classification of the disease, or HBsAg status. The drug was well tolerated with few side-effects. The results of this short-term study indicate that tiopronin may be of value in the treatment of chronic hepatitis.

[Serotypes and drug susceptibility of Pseudomonas aeruginosa isolated from clinical specimens].

Between January, 1982 and December, 1994, 236 Pseudomonas aeruginosa strains were isolated from clinical specimens at our division, and were tested for serotypes and drug-susceptibilities to 15 antibiotics. Serotype G strains were isolated at the highest frequency (32.6%), and followed by strains of serotype B (15.7%), A (11.9%), E (9.3%), I (7.2%), F and M (5.5%), non-typable (5.1%), D (3.4%), H (2.1%), C and K (0.8%). We examined the changes of isolation frequencies of different serotypes annually. Isolation frequencies of serotypes E and F showed tendency to decrease, whereas serotype I has been isolated increasingly year by year. MIC90's of the 15 antibiotics were as follows, tosufloxacin: 0.78 microgram/ml, biapenem (BIPM) and ofloxacin (OFLX): 3.13 micrograms/ml, imipenem (IPM), ceftazidime, cefozopran, cefsulodin and gentamicin: 6.25 micrograms/ml, aztreonam and amikacin: 12.5 micrograms/ml, piperacillin, cefoperazone and minocycline (MINO): 25 micrograms/ml, fosfomycin: > 100 micrograms/ml and chloramphenicol: > 200 micrograms/ml. MIC90'S of IPM, BIPM, MINO and OFLX increased 4-fold from stage I (1982-1987) through stage III (1992-1994) and the isolation frequency of drug-resistant strains increased year by year. In other words, antibiotic resistant strains appeared increasing with time. No relationship between serotypes and drug-resistance were observed.

[Drug susceptibility of clinically isolated Helicobacter pylori].

Between January 1995 and March 1997, 78 Helicobacter pylori strains were isolated from patients with gastritis and gastric ulcer and their drug-susceptibilities to 8 antimicrobial agents and 3 anti-ulcer drugs were determined. Imipenem was the most active agent and its MICs to all the strains tested were lower than 0.013 microgram/ml. Amoxicillin, cefaclor and minocycline were active against H. pylori with MIC90s of 0.05 microgram/ml, 0.78 microgram/ml and 0.39 microgram/ml, respectively, and no resistant strains against these drugs were isolated. However, resistant strains to clarithromycin (isolation frequency: 9%), erythromycin (13%), ofloxacin (8%) and metronidazole (13%) were isolated. Triple, double and single resistant strains to above 4 antimicrobial agents were noted. No quadruple resistant strain was isolated. Frequencies of those resistance patterns were 14.3% (triple), 28.6% (double), and 57.1% (single), respectively. Seven erythromycin-resistant strains were shown to be cross-resistant to clarithromycin but 3 erythromycin-resistant strains were susceptible to clarithromycin. It seems likely that this phenomenon is caused by the fact that clarithromycin is more active to H. pylori than erythromycin. The MIC90 value of lansoprazole was lower than those of omeprazole and famotidine.

pH response of human dental plaque to chewing gum supplemented with low molecular chitosan.

The effects of low molecular chitosan (LMCS) on pH responses of human dental plaque following exposure to fermentable carbohydrates were investigated by an ion-sensitive field-effect transistor electrode system. After the plaque pH values were minimized by direct application of 5% glucose solution or consumption of sugared caramel, the subjects started chewing the test gums containing 0 (control), 1 or 3% (w/w) LMCS for three minutes. The pH response was monitored until it recovered to over pH5.5. In the case of the glucose solution, chewing 3% LMCS gum caused significantly more rapid pH recovery toward the resting level than did the control gum. Initial pH rising rate during gum chewing was faster with either of the two LMCS gums than with the control gum. In the case of caramel, additional effects of LMCS were observed numerically as LMCS content increased. The findings indicated that LMCS had a potential to promote recovery of plaque pH after acidogenic challenge and to maintain the plaque pH around neutrality.

The influences of the buffer capacity of various substances on pH changes in dental plaque.

This study clarified the suitable pKa value for buffering substances against plaque pH fall in vitro and simultaneously estimated the effect of low molecular chitosan (LMCS) on plaque pH lowered by metabolized acids in vitro and in vivo. Five buffering substances with different pKa, aspartame (pKa: 7.8), phosphate buffer (7.1), LMCS (6.4), maleate buffer (6.2), and monofluorophosphate (4.8), were tested in this study. In the method using S. mutans cells, phosphate inhibited the pH fall from an initial pH of over 7.0, but phosphate exhibited no effect when the initial pH was 6.0. By the addition with lactic acid, LMCS and maleate exhibited more effective inhibition of the pH-fall than the others. These observations imply that pKa value is an important indicator of the ability of a buffering substance to reduce pH fall in dental plaque and that the optimum pKa value may be around pH 6.3. In the plaque pH measurement using ISFET electrode, LMCS showed an additional effect in inhibiting plaque pH fall following direct application of the glucose solution. The findings indicate that LMCS may be useful as a food additive to decrease the cariogenicity of foods.