Molecular mimicry of Aggregatibacter actinomycetemcomitans with beta2 glycoprotein I.

INTRODUCTION: beta2-Glycoprotein I (beta 2GPI) is important in the suppression of coagulation, and antibodies against TLRVYK peptides on the beta 2GPI molecule are related to thrombosis. According to the Swiss-Prot database, Aggregatibacter actinomycetemcomitans leukotoxin c has sequences (SIRVYK) that are homologous to the TLRVYK peptides. The aim of this study was to investigate the effects of A. actinomycetemcomitans infection on the antibody response against SIRVYK peptides in patients with periodontitis. METHODS: Serum immunoglobulin G (IgG) antibody and IgG subclass antibody titers against SIRVYK or TLRVYK peptides were measured by enzyme-linked immunosorbent assay in 46 patients with aggressive periodontitis (eight with localized disease, 38 with generalized disease), 28 patients with chronic periodontitis, and 20 periodontally healthy subjects. The presence of A. actinomycetemcomitans in plaque and saliva samples was determined using polymerase chain reaction. RESULTS: The level of anti-SIRVYK antibodies was significantly higher in patients who were A. actinomycetemcomitans-positive than in A. actinomycetemcomitans-negative patients (P < 0.05) in the chronic periodontitis group. A similar trend was found in the antibody response to TLRVYK peptide; however, no statistically significant difference was seen between A. actinomycetemcomitans-positive and -negative patients. The A. actinomycetemcomitans-positive patients displayed significantly higher levels of anti-SIRVYK IgG2 and IgG3 antibodies than A. actinomycetemcomitans-negative patients (P < 0.05 and P < 0.05, respectively). The level of IgG2 was highest among the four IgG subclasses and it predominantly increased in patients who were A. actinomycetemcomitans-positive. Anti-TLRVYK antibody levels were significantly correlated with anti-SIRVYK IgG antibody levels. CONCLUSION: The results suggest that A. actinomycetemcomitans infection may elicit anti-SIRVYK IgG antibodies and modify the anti-TLRVYK antibody response in patients with periodontitis by molecular mimicry with beta2GPI.

Bovine alpha-lactalbumin stimulates mucus metabolism in gastric mucosa.

Bovine alpha-lactalbumin (alpha-LA), a major milk protein, exerts strong gastroprotective activity against rat experimental gastric ulcers induced by ethanol or stress. To elucidate the mechanisms underlying this activity, the influence of alpha-LA on gastric mucus metabolism was investigated in vitro and in vivo. For the in vitro study, RGM1 cells (a rat gastric epithelial cell line) were selected for observation of the direct activity of alpha-LA on gastric mucosal cells and cultured in the presence of either alpha-LA or ovalbumin (OVA), a reference protein showing no gastroprotective activity. Amounts of synthesized and secreted mucin, a major component of mucus, were determined using [3H]glucosamine as a tracer, and prostaglandin E2 (PGE2) levels in the culture medium were determined by RIA. For the in vivo study, the thickness of the mucus gel layer, a protective barrier for gastric mucosa, was evaluated histochemically in rat gastric mucosa. alpha-Lactalbumin (3 mg/mL) significantly stimulated mucin synthesis and secretion in RGM1 cells and also increased PGE2 levels in the culture medium. In contrast, OVA showed no enhancing effects under identical conditions. Neither indomethacin, a cyclo-oxygenase inhibitor, nor AH23848, a prostaglandin EP4 receptor antagonist, affected alpha-LA-induced enhancement of mucin synthesis and secretion. In vivo, oral administration of alpha-LA (300 mg/kg x 3 times/d x 7 d) increased the thickness of the mucus gel layer in rats. These results indicate that alpha-LA fortifies the mucus gel layer by stimulating mucin production and secretion in gastric mucus-producing cells, and that this enhancing effect is independent of endogenous PGE2. Comparison of the efficacy of alpha-LA with OVA suggests that the activities observed in RGM1 cells are closely related to the gastroprotective effects in rat gastric ulcer models. In conclusion, alpha-LA stimulates mucus metabolism, and this action may be responsible for its gastroprotective activity.

CD31 expression on leukocytes is downregulated in vivo during hemodialysis.

BACKGROUND/AIM: CD31 on leukocytes is the adhesion molecule involved in the leukocyte extravasation in inflammatory conditions. During hemodialysis with cellulosic membranes, it is considered that activated leukocytes adhere to endothelium, but do not show extravasation. However, it is not elucidated why activated leukocytes do not show endothelial transmigration during hemodialysis with cellulosic membranes. METHODS: In the present study, changes in the expressions of Mac-1 and CD31 on granulocytes and monocytes were analyzed by flow cytometry during hemodialysis in 7 patients treated with regenerated-cellulose (RC) membranes and next with polysulfone (PS) membranes. RESULTS: During dialysis with RC, Mac-1 expressions on granulocytes and monocytes both significantly increased as compared with predialysis values and across the dialyzer. During dialysis with RC, the CD31 expression on granulocytes and monocytes significantly decreased as compared with predialysis values. During dialysis with PS, changes in Mac-1 and CD31 expressions on granulocytes and monocytes were smaller than those during dialysis with RC. CONCLUSIONS: Decreased CD31 expression on leukocytes may affect leukocyte function more in patients chronically hemodialyzed with RC than in those hemodialyzed with PS, since CD31 is important in leukocyte transendothelial migration in inflammatory conditions.

Functional fragments of ingested lactoferrin are resistant to proteolytic degradation in the gastrointestinal tract of adult rats.

Pharmaceutical and food-related applications of lactoferrin, an 80-kDa iron-binding glycoprotein found predominantly in milk, have attracted interest lately, but the process of digestion of lactoferrin has been poorly characterized. The digestive fate of bovine lactoferrin in adult rats after oral administration of a single dose and after dietary supplementation was studied by (125)I-labeling and by surface-enhanced laser desorption/ionization (SELDI) affinity mass spectrometry. The latter method was designed to detect multiple forms of degraded lactoferrin as simple molecular ion peaks corresponding to one of the core regions of lactoferrin, namely, the lactoferricin region (Phe17-Ala42). Radioactive fragments with molecular masses of 42, 36, 33 and 29 kDa were observed at 20, 60 and 180 min postingestion in the contents of the lower small intestine. Rats were given free access to milk enriched with lactoferrin at 482 micromol/L (40 mg/mL). The concentrations of lactoferrin fragments in the contents of the stomach, small intestine and lower small intestine as determined by SELDI affinity mass spectrometry were approximately 200, 20 and 1 micromol/L, respectively. These data indicate that functional fragments of LF such as fragments containing glycosaminoglycan-binding site(s), as well as large fragments with a mass >20 kDa, indeed survive proteolytic degradation in the small intestine of adult rats.

New biological function of bovine alpha-lactalbumin: protective effect against ethanol- and stress-induced gastric mucosal injury in rats.

Although several studies have shown that milk protein components have a wide range of biological activities, the potential role of these proteins in the gastrointestinal mucosal defense system is less well elucidated. In this study, we investigated the effect of the major proteins in cow's milk on gastric mucosal injury by using two acute ulcer models in Wistar rats. Gastric mucosal injury was induced by either intragastric 60% ethanol-HCl or water-immersion restraint stress (23 degrees C, 7 h). Each test milk protein was orally administered 30 min before the induction of gastric injury. Among the major milk proteins, alpha-lactalbumin (alpha-LA) is demonstrated to have a marked protective effect against ethanol-induced gastric injury, with the same potency as that of the typical antiulcer agent, Selbex. Whey protein isolate (WPI), which contained 25% alpha-LA, also protected against gastric injury, while casein showed no effect. Comparative studies on the protective effect of the four major components of WPI, beta-lactoglobulin, alpha-LA, bovine serum albumin and gamma-globulins (immunoglobulins), on the basis of their contents in WPI revealed that alpha-LA was responsible for the protective effect of WPI, being about 4-fold more effective than WPI itself. Alpha-LA showed dose-dependent protection against gastric injury induced by stress as well as ethanol. Pretreatment with indomethacin (10 mg/kg body weight, s.c.), which is a potent inhibitor of endogenous prostaglandin synthesis, resulted in a significant reduction in the protective effect of alpha-LA. These results indicate that alpha-LA has marked antiulcer activity as an active component of cow's milk protein, and suggest that alpha-LA intake may serve to protect against gastric mucosal injury, in part through endogenous prostaglandin synthesis.

[Factors influencing noncompliance with medication regimens in the elderly].

Poor medication compliance is a major issue in the health care of older patients. To identify risk factors for medication noncompliance in the elderly, inpatients aged 65 years and older at Nagoya University Hospital and at Chubu National Hospital underwent a comprehensive geriatric assessment and tests for the assessment of medication compliance. The dependency of medication assistance by a caregiver is associated with low physical function activity, cognitive impairment, depression and communication inability. Medication noncompliance was not associated with the score of any component of comprehensive geriatric assessment. There was a good relationship between patient's knowledge of medications and the frequency of dosage interval, both of which were associated with the score of instrumental ADL, cognitive function and communication ability. The knowledge was also associated with the medication compliance at Nagoya University Hospital but not at Chubu National Hospital. These results may suggest that the elderly patient's understanding of a medication regimen is important but that other factors are also required to maintain their treatment regimen.

Milk and dairy products in cancer prevention: focus on bovine lactoferrin.

Milk and dairy products constitute an important part of the western style diet. A large number of epidemiological studies have been conducted to determine effects of consumption on cancer development but the data are largely equivocal, presumably reflecting the different included components. It has been proposed that whereas fats in general could promote tumor development, individual milk fats like conjugated linoleic acid could exert inhibitory effects. There is also considerable evidence that calcium in milk products protects against colon cancer, while promoting in the prostate through suppression of circulating levels of 1,25-dihydroxyvitamin D3. Whey protein may also be beneficial, as shown by both animal and human studies, and experimental data have demonstrated that the major component bovine lactoferrin (bLF), inhibits colon carcinogenesis in the post-initiation stage in male F344 rats treated with azoxymethane (AOM) without any overt toxicity. The incidence of adenocarcinomas in the groups receiving 2% and 0.2% bLF were thus 15% and 25%, respectively, in contrast to the 57.5% control value (P<0.01 and P<0.05, respectively). Results in other animal models have provided further indications that bLF might find application as a natural ingredient of milk with potential for chemoprevention of colon and other cancers.

Orally administered lactoferrin exerts an antimetastatic effect and enhances production of IL-18 in the intestinal epithelium.

The effects of oral administration of bovine lactoferrin (bLF) and its hydrolysate on the lung colonization by colon 26 carcinoma were investigated. At doses of 100 or 300 mg/kg/day for seven successive days, bLFs demonstrated a significant inhibitory effect on experimental metastasis, which indicated effectiveness before and after tumor implantation. Oral administration of bLFs augmented CD4+, CD8+, and asialoGM1+ cells in the spleen and peripheral blood. Their cytotoxic activities against Yac-1 and colon 26 carcinoma were enhanced by bLF. In the small intestinal epithelium, CD4+ and CD8+ cells were markedly increased, and, simultaneously, enhanced production of interleukin-18 (IL-18) was confirmed in the intestinal epithelial cells. In this model, intravenous injection of murine IL-18 showed significant inhibition of the lung colonization by colon 26 carcinoma. These results suggested that inhibition of experimental metastasis by oral administration of bLF and pepsin hydrolysate of bLF might be due to enhanced cellular immunity, presumably mediated by enhanced IL-18 production in the intestinal epithelium.

Inhibitory effects of bovine lactoferrin on colon carcinoma 26 lung metastasis in mice.

In order to determine the effects of the multifunctional iron-binding glycoprotein, lactoferrin (LF), and related compounds on tumor growth and metastasis, bovine LF (bLF), and bLF hydrolysate and lactoferricin (bLFcin), active products generated by acid-pepsin hydrolysis were administered orally to BALB/c mice bearing subcutaneous (s.c.) implants of the highly metastatic colon carcinoma 26 (Co 26Lu). bLF and the bLF hydrolysate demonstrated significant inhibition of lung metastatic colony formation from s.c. implanted tumors without appreciable effects on tumor growth. bLFcin displayed a tendency for inhibition of lung metastasis. On the other hand, bLF did not exert marked anti-metastatic activity in athymic nude mice bearing Co 26Lu, though bLF had a tendency to inhibit the lung metastatic colony formation associated with anti-asialoGM1 antibody (Ab) treatment. AsialoGM1+ and CD8+ cells in white blood cells were increased after treatment with bLF. In vitro, the viability of Co 26Lu-F55 cells was markedly decreased when co-cultured with white blood cells from mice administrated bLF p.o., but recovered on treatment with anti-asialoGM1 Ab or anti-CD8 mAb and complement. The results suggest bLF and related compounds might find application as tools in the control of metastasis and that asialoGM1+ and CD8+ cells in the blood are important for their inhibitory effects.

Possible chemopreventive effects of bovine lactoferrin on esophagus and lung carcinogenesis in the rat.

A milk component, bovine lactoferrin (bLF), previously shown by us to be a strong chemopreventive of colon carcinoma development, was examined for its influence on other organs using a rat multi-organ carcinogenesis model. Male F344 rats, aged 6 weeks, were treated sequentially with diethylnitrosamine (DEN, i.p.), dihydroxy-di-N-propylnitrosamine (DHPN, in drinking water) and N-nitrosomethylbenzylamine (NMBA, s.c.) during the first 8 weeks (DDN treatment), and then bLF was administered in the basal diet, at a dose of 2, 0.2, 0.02 or 0.002%. Other groups were given DDN treatment or bLF alone as controls. All surviving animals were killed at week 41, and major organs were examined histopathologically for neoplastic lesions. In the esophagus, a tendency for reduction in development of papillomas was evident in the bLF-treated animals, along with a significant suppression of relatively large-sized papillomas (more than 50 mm3 volume) at the 0.2% dose (P<0.05, 11% of the control). The multiplicity of tumors (adenomas and carcinomas) in the lung was also decreased in animals fed 0.02% bLF (1.98+/-0.41 per cm2 lung tissue section, P<0.05) compared to the control group (3.48+/-0.33). No enhancing or inhibitory effects of bLF on tumor development in other organs were noted. The present results indicate that bLF exerts chemopreventive effects in the esophagus and lung in addition to the colon.

Inhibition of azoxymethane initiated colon tumor and aberrant crypt foci development by bovine lactoferrin administration in F344 rats.

The influence of bovine lactoferrin (bLf) on colon carcinogenesis was investigated in male F344 rats treated with azoxymethane (AOM). In experiment I, 2% and 0.2% bLf, and Bifidobacterium longum (B. longum) as a positive control at 3% were given in the diet for 4 weeks, along with two s.c. 15 mg/kg injections of AOM on days 1 and 8. The numbers of aberrant crypt foci (ACF) were decreased by both treatments. Similar results were obtained in experiment II of 13 weeks duration. In experiment III, animals were given three weekly injections of AOM and then received 2 or 0.2% bLf, 2% bLf-hydrolysate, or 0.1% bovine lactoferricin (bLfcin) for 36 weeks. No effects indicative of toxicity were noted, but significant reduction in both the incidence and number of adenocarcinomas of the large intestine was observed with almost all the treatments. Thus, the incidences of colon adenocarcinomas in the groups receiving 2 or 0.2% bLf, 2% bLf-hydrolysate, or 0.1% bLfcin were 15%, 25%, 26.3% and only 10%, respectively, in contrast to the 57.5% control value (p < 0.01). ACF values also exhibited reduced development. Investigation of beta-glucuronidase revealed decrease in the cecal contents of animals receiving bLf. In addition, demonstration of enhancement of NK activity by bLf indicated that its inhibitory effects could have been related to elevated immune cytotoxicity.

Inhibitory effects of bovine lactoferrin on intestinal polyposis in the Apc(Min) mouse.

Chemopreventive effects of bovine lactoferrin (bLF), previously shown to strongly inhibit intestinal carcinogenesis in rats (K. Sekine, E. Watanabe, J. Nakamura, N. Takasuka, D.J. Kim, M. Asamoto, V. Krutovskikh, T.H. Baba, T. Ota, M.A. Moore, M. Masuda, H. Sugimoto, H. Nishino, T. Kakizoe, H. Tsuda, Inhibition of azoxymethane-initiated colon tumor by bovine lactoferrin administration in F344 rats, Jpn. J. Cancer Res. 88 (1997) 523-526; K. Sekine, Y. Ushida, T. Kuhara, M. Iigo, H. Baba-Toriyama, M.A. Moore, M. Murakoshi, Y. Satomi, H. Nishino, T. Kakizoe, H. Tsuda, Inhibition of initiation and early stage development of aberrant crypt foci and enhanced natural killer activity in male rats administered bovine lactoferrin concomitantly with azoxymethane, Cancer Lett. 121 (1997) 211-216), on spontaneous intestinal polyp development were assessed in the ApcMin mouse, a model for both familial adenomatous polyposis and sporadic colon cancers. In the experiment, 54 mice at 6 weeks of age were given 2% bLF (15 mice), 0.2% bLF (15 mice) and AIN-93G (24 mice) as basal diet ad libitum for 8 weeks. An overall tendency for a reduction in the total number of polyps in the small intestine was evident in the bLF-treated animals, along with significant suppression in the jejunum at the 2% dose (P < 0.05, 68% of the control). In addition, body growth suppression, presumed to be due to anemia and/or intussusception as a consequence of numerous polyps in the intestine, was alleviated. No toxic effects were observed in the intestinal epithelium. Although not as obvious as observed for the rat case, the data suggest that bLF may be a chemopreventor of intestinal polyposis.

Structures of the sugar chains of recombinant macrophage colony-stimulating factor produced in Chinese hamster ovary cells.

The structures of the N- and O-linked sugar chains of recombinant human macrophage colony-stimulating factor (rhM-CSF) from Chinese hamster ovary (CHO) cells were studied. rhM-CSF is a homodimeric glycoprotein. Sugar composition analysis revealed that rhM-CSF contained 4.1 mol N-acetylgalactosamine, 10.3 mol N-acetylglucosamine, 5.0 mol mannose, 10.0 mol galactose, 1.4 mol fucose, and 11.8 mol sialic acid per mol of the monomer. The N- and O-linked sugar chains liberated by hydrazinolysis were N-acetylated, and the reducing-end sugar residues were tagged with 2-aminopyridine. The pyridylamino (PA-) sugar chains thus obtained were purified by HPLC. The structures of the PA-sugar chains were analyzed by a combination of reversed-phase and size-fractionation HPLC, and exoglycosidase digestions, from which the structures of the rhM-CSF sugar chains were estimated to be as follows: monosialo biantennary sugar chain (9 mol%), monosialo fucosylbiantennary sugar chain (10 mol%), disialo biantennary sugar chain (30 mol%), disialo fucosylbiantennary sugar chain (28 mol%), disialo triantennary sugar chain (7 mol%), trisialo triantennary sugar chain (11 mol%), and trisialo fucosyltriantennary sugar chain (5 mol%) for the N-linked sugar chains, and asialo (27 mol%), monosialo (51 mol%), and disialo (22 mol%) Galbeta1-3GalNAc for the O-linked sugar chains. Sialic acid residues were linked to the N-linked sugar chains through an alpha2-3 linkage.

Inhibition of initiation and early stage development of aberrant crypt foci and enhanced natural killer activity in male rats administered bovine lactoferrin concomitantly with azoxymethane.

The influence of concomitant administration of bovine lactoferrin (bLF) on induction of aberrant crypt foci (ACF) by azoxymethane was investigated in male F344 rats. Two percent bLF and 3% Bifidobacterium longum (B. longum), as a positive control, significantly decreased the numbers of ACF as well as the total numbers of aberrant crypts reproducibly in three independent studies (2% bLF, P < 0.01; 3% B. longum, P < 0.05). Most importantly large size foci composed of four or more crypts were always significantly decreased by 2% bLF (P < 0.05). Additional investigation of the natural killer activity of spleen cells demonstrated enhancement by bLF (P < 0.01) and B. longum (P < 0.01) in line with the levels of influence on foci induction, indicating a possible role for elevated immune cytotoxicity in the observed inhibition.

Development of plant growth apparatus using blue and red LED as artificial light source.

It is known that chlorophyll has the second distinct absorption peak in the vicinity of 450nm (blue light region) other than the first peak in the vicinity of 660nm (red light region) in its light absorption spectrum The blue light is also indispensable to the morphologically healthy growth plant. On the other hand, the red light contributes to the plant photosynthesis. Noticing this facts, we have developed various kind of plant growth apparatus using many pieces of blue light LED and red light LED with emission wavelength 450nm and 660nm as artificial light source. In this paper, we introduce our LED plant growth apparatus and systems named such as LED PACK, BIOLED, UNIPACK, and COMPACK with respect to their structure, function, electrical design, and characteristics.