Bovine lactoferrin enhances osteogenesis through Smad2/3 and p38 MAPK activation.

OBJECTIVES: Lactoferrin (LF) possesses diverse biological functions. We previously reported that bovine LF (bLF) attenuates lipopolysaccharide-induced bone resorption in osteoblasts. In addition to its ability to inhibit osteoclastogenesis, bLF has been implicated in stimulating bone formation. However, the molecular mechanisms of bLF in bone cell anabolism remain unclear. Here, we tried to analyse the molecular mechanisms involved in osteogenesis in the presence of bLF. METHODS: Alkaline phosphatase activity, Runx2 activity, gene expression, and Alizarin red staining were analyzed to evaluate the osteogenic differentiation status. The expression of the Smads and mitogen-activated protein kinase (MAPK) signaling molecules was analyzed via western blotting. Ex vivo organ cultures of mouse calvariae were performed to evaluate the effect of bLF on bone regeneration. RESULTS: bLF enhanced the osteoblastic differentiation of mesenchymal stem cells through activation of Smad2/3 and p38 MAPK, which increased the transcriptional activity of Runx2. bLF treatment also enhanced osteoblastic differentiation and mineralized nodule formation of osteoblast-lineage cells, and repaired bone defects ex vivo. Moreover, inhibition of Smad2/3 or p38 MAPK signaling reduced the anabolic effects of bLF. Together, these results suggested that bLF is a potent osteogenic factor, which mediates its function via activation of the Smad2/3 and p38 MAPK signaling pathways. CONCLUSIONS: Here, we described a novel function of bLF and its signal transduction mechanisms in osseous tissue. Along with inhibiting osteoclastogenesis, bLF may limit further osteoclast formation and contribute to bone mass enlargement. Thus, bLF represents a potentially valuable therapeutic agent for bone regeneration and destructive bone diseases.

Fiber-rich diet with brown rice improves endothelial function in type 2 diabetes mellitus: A randomized controlled trial.

BACKGROUND & AIMS: A fiber-rich diet has a cardioprotective effect, but the mechanism for this remains unclear. We hypothesized that a fiber-rich diet with brown rice improves endothelial function in patients with type 2 diabetes mellitus. METHODS: Twenty-eight patients with type 2 diabetes mellitus at a single general hospital in Japan were randomly assigned to a brown rice (n = 14) or white rice (n = 14) diet and were followed for 8 weeks. The primary outcome was changes in endothelial function determined from flow debt repayment by reactive hyperemia using strain-gauge plethysmography in the fasting state. Secondary outcomes were changes in HbA1c, postprandial glucose excursions, and markers of oxidative stress and inflammation. The area under the curve for glucose after ingesting 250 kcal of assigned rice was compared between baseline (T0) and at the end of the intervention (T1) to estimate glucose excursions in each group. RESULTS: Improvement in endothelial function, assessed by fasting flow debt repayment (20.4% vs. -5.8%, p = 0.004), was significantly greater in the brown rice diet group than the white rice diet group, although the between-group difference in change of fiber intake was small (5.6 g/day vs. -1.2 g/day, p<0.0001). Changes in total, HDL-, and LDL-cholesterol, and urine 8-isoprostane levels did not differ between the two groups. The high-sensitivity C-reactive protein level tended to improve in the brown rice diet group compared with the white rice diet group (0.01 µg/L vs. -0.04 µg/L, p = 0.063). The area under the curve for glucose was subtly but consistently lower in the brown rice diet group (T0: 21.4 mmol/L*h vs. 24.0 mmol/L*h, p = 0.043, T1: 20.4 mmol/L*h vs. 23.3 mmol/L*h, p = 0.046) without changes in HbA1c. CONCLUSIONS: Intervention with a fiber-rich diet with brown rice effectively improved endothelial function, without changes in HbA1c levels, possibly through reducing glucose excursions.

A high-fiber, low-fat diet improves periodontal disease markers in high-risk subjects: a pilot study.

Periodontal disease is related to aging, smoking habits, diabetes mellitus, and systemic inflammation. However, there remains limited evidence about causality from intervention studies. An effective diet for prevention of periodontal disease has not been well established. The current study was an intervention study examining the effects of a high-fiber, low-fat diet on periodontal disease markers in high-risk subjects. Forty-seven volunteers were interviewed for recruitment into the study. Twenty-one volunteers with a body mass index of at least 25.0 kg/m(2) or with impaired glucose tolerance were enrolled in the study. After a 2- to 3-week run-in period, subjects were provided with a test meal consisting of high fiber and low fat (30 kcal/kg of ideal body weight) 3 times a day for 8 weeks and followed by a regular diet for 24 weeks. Four hundred twenty-five teeth from 17 subjects were analyzed. Periodontal disease markers assessed as probing depth (2.28 vs 2.21 vs 2.13 mm; P < .0001), clinical attachment loss (6.11 vs 6.06 vs 5.98 mm; P < .0001), and bleeding on probing (16.2 vs 13.2 vs 14.6 %; P = .005) showed significant reductions after the test-meal period, and these improvements persisted until the follow-up period. Body weight (P < .0001), HbA1c (P < .0001), and high-sensitivity C-reactive protein (P = .038) levels showed improvement after the test-meal period; they returned to baseline levels after the follow-up period. In conclusion, treatment with a high-fiber, low-fat diet for 8 weeks effectively improved periodontal disease markers as well as metabolic profiles, at least in part, by effects other than the reduction of total energy intake.

4-Hydroxy hexenal derived from docosahexaenoic acid protects endothelial cells via Nrf2 activation.

Recent studies have proposed that n-3 polyunsaturated fatty acids (n-3 PUFAs) have direct antioxidant and anti-inflammatory effects in vascular tissue, explaining their cardioprotective effects. However, the molecular mechanisms are not yet fully understood. We tested whether n-3 PUFAs showed antioxidant activity through the activation of nuclear factor erythroid 2-related factor 2 (Nrf2), a master transcriptional factor for antioxidant genes. C57BL/6 or Nrf2(-/-) mice were fed a fish-oil diet for 3 weeks. Fish-oil diet significantly increased the expression of heme oxygenase-1 (HO-1), and endothelium-dependent vasodilation in the aorta of C57BL/6 mice, but not in the Nrf2(-/-) mice. Furthermore, we observed that 4-hydroxy hexenal (4-HHE), an end-product of n-3 PUFA peroxidation, was significantly increased in the aorta of C57BL/6 mice, accompanied by intra-aortic predominant increase in docosahexaenoic acid (DHA) rather than that in eicosapentaenoic acid (EPA). Human umbilical vein endothelial cells were incubated with DHA or EPA. We found that DHA, but not EPA, markedly increased intracellular 4-HHE, and nuclear expression and DNA binding of Nrf2. Both DHA and 4-HHE also increased the expressions of Nrf2 target genes including HO-1, and the siRNA of Nrf2 abolished these effects. Furthermore, DHA prevented oxidant-induced cellular damage or reactive oxygen species production, and these effects were disappeared by an HO-1 inhibitor or the siRNA of Nrf2. Thus, we found protective effects of DHA through Nrf2 activation in vascular tissue, accompanied by intra-vascular increases in 4-HHE, which may explain the mechanism of the cardioprotective effects of DHA.

Willow bark extract increases antioxidant enzymes and reduces oxidative stress through activation of Nrf2 in vascular endothelial cells and Caenorhabditis elegans.

Willow bark extract (WBE) is listed in the European Pharmacopoeia and has been traditionally used for treating fever, pain, and inflammation. Recent studies have demonstrated its clinical usefulness. This study investigated the antioxidative effects of WBE in human umbilical vein endothelial cells (HUVECs) and Caenorhabditis elegans. WBE prevented oxidative-stress-induced cytotoxicity of HUVECs and death of C. elegans. WBE dose-dependently increased mRNA and protein expression levels of the nuclear factor erythroid 2-related factor 2 (Nrf2) target genes heme oxygenase-1, γ-glutamylcysteine ligase modifier and catalytic subunits, and p62 and intracellular glutathione (GSH) in HUVECs. In the nematode C. elegans, WBE increased the expression of the gcs-1::green fluorescent protein reporter, a well-characterized target of the Nrf2 ortholog SKN-1, in a manner that was SKN-1-dependent. WBE increased intranuclear expression and DNA binding of Nrf2 and the activity of an antioxidant response element (ARE) reporter plasmid in HUVECs. WBE-induced expression of Nrf2-regulated genes and increased GSH levels in HUVECs were reduced by Nrf2 and p38 small interfering (si) RNAs and by the p38-specific inhibitor SB203580. Nrf2 siRNA reduced the cytoprotective effect of WBE against oxidative stress in HUVECs. Salicin, a major anti-inflammatory ingredient of WBE, failed to activate ARE-luciferase activity, whereas a salicin-free WBE fraction showed intensive activity. WBE induced antioxidant enzymes and prevented oxidative stress through activation of Nrf2 independent of salicin, providing a new potential explanation for the clinical usefulness of WBE.

Molecular mechanisms of the inhibitory effects of bovine lactoferrin on lipopolysaccharide-mediated osteoclastogenesis.

Lactoferrin (LF) is an important modulator of the immune response and inflammation. It has also been implicated in the regulation of bone tissue. In our previous study we demonstrated that bovine LF (bLF) reduces LPS-induced bone resorption through a reduction of TNF-α production in vivo. However, it was not known how bLF inhibits LPS-mediated TNF-α and RANKL (receptor activator of nuclear factor κB ligand) production in osteoblasts. In this study we show that bLF impairs LPS-mediated TNF-α and RANKL production. bLF inhibited LPS-mediated osteoclastogenesis via osteoblasts in a co-culture system. Furthermore, bLF pretreatment inhibited LPS-induced NFκB DNA binding activity as well as IκBα and IKKß (IκB kinase ß) phosphorylation. MAP kinase activation was also inhibited by bLF pretreatment. However, bLF pretreatment failed to block the degradation of IRAK1 (interleukin-1 receptor-associated kinase 1), which is an essential event after its activation. Remarkably, we found that bLF pretreatment inhibited LPS-mediated Lys-63-linked polyubiquitination of TNF receptor-associated factor 6 (TRAF6). We also found that bLF is mainly endocytosed through LRP1 (lipoprotein receptor-related protein-1) and intracellular distributed bLF binds to endogenous TRAF6. In addition, bLF inhibited IL-1ß- and flagellin-induced TRAF6-dependent activation of the NFκB signaling pathway. Collectively, our findings demonstrate that bLF inhibits NFκB and MAP kinase activation, which play critical roles in chronic inflammatory disease by interfering with the TRAF6 polyubiquitination process. Thus, bLF could be a potent therapeutic agent for inflammatory diseases associated with bone destruction, such as periodontitis and rheumatoid arthritis.

Human trial of liposomal lactoferrin supplementation for periodontal disease.

New approaches to periodontal health have been in strong demand in addition to conventional local plaque control. In this study, liposomal bovine lactoferrin (L-bLF) was orally administered to subjects with periodontal disease to investigate whether it could be a useful treatment. L-bLF composed of soy phosphatidylcholine was given as a supplement for four weeks in tablet form (180 mg bLF/d) to twelve subjects with multiple sites of more than 3 mm probing depth (PD). PD, bleeding on probing (BOP), gingival crevicular fluid (GCF) volume and the levels of tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, IL-6, and monocyte chemoattractant protein-1 (MCP-1) in GCF were evaluated for 51 sites with more than 4 mm PD in five subjects. Blood samples of all subjects were collected 0, 2 and 4 weeks after supplementation. Isolated peripheral blood mononuclear cells (PBMCs) were incubated for 24 h with or without lipopolysaccharide (LPS) (100 ng/ml) from Porphyromonas gingivalis, and TNF-α, IL-1ß, IL-6 and MCP-1 in the culture media were measured. Toll-like receptor 2 (TLR2) and TLR4 mRNA expressions of isolated PBMCs were also quantitatively analyzed using real-time reverse transcription-polymerase chain reaction (RT-PCR). The PD was significantly reduced by L-bLF supplementation, but the BOP and GCF volume were not significantly changed. The MCP-1 level in GCF was significantly reduced, while levels of other cytokines were not changed. Four-week L-bLF supplementation also showed significant decreases of LPS-induced cytokine production from PBMCs. Relative gene expressions of TLR2 and TLR4 did not change. These results suggest that L-bLF supplementation can be effective in the treatment of periodontal disease, although prospective controlled large-scale studies are required.

Low concentration of 4-hydroxy hexenal increases heme oxygenase-1 expression through activation of Nrf2 and antioxidative activity in vascular endothelial cells.

Large-scale clinical studies have shown that n-3 polyunsaturated fatty acids (n-3 PUFAs) such as eicosapentaenoic and docosahexaenoic acids reduce cardiovascular events without improving classical risk factors for atherosclerosis. Recent studies have proposed that direct actions of n-3 PUFAs themselves, or of their enzymatic metabolites, have antioxidative and anti-inflammatory effects on vascular cells. Although a recent study showed that plasma 4-hydroxy hexenal (4-HHE), a peroxidation product of n-3 PUFA, increased after supplementation of docosahexaenoic acid, the antiatherogenic effects of 4-HHE in vascular cells remain unclear. In the present study, we tested the hypothesis that 4-HHE induces the antioxidative enzyme heme oxygenase-1 (HO-1) through activation of nuclear factor erythroid 2-related factor 2 (Nrf2), a master regulatory transcriptional factor, and prevents oxidative stress-induced cytotoxicity in vascular endothelial cells. This mechanism could partly explain the cardioprotective effects of n-3 PUFAs. Human umbilical vein endothelial cells were stimulated with 1-10µM 4-HHE or 4-hydroxy nonenal (4-HNE), a peroxidation product of n-6 PUFAs. Both 4-HHE and 4-HNE dose-dependently increased HO-1 mRNA and protein expression, and intranuclear expression and DNA binding of Nrf2 at 5µM. Small interfering RNA for Nrf2 significantly reduced 4-HHE- or 4-HNE-induced HO-1 mRNA and protein expression. Furthermore, pretreatment with 4-HHE or 4-HNE prevented tert-butyl hydroperoxide-induced cytotoxicity. In conclusion, 4-HHE, a peroxidation product of n-3 PUFAs, stimulated expression of the antioxidant enzyme HO-1 through the activation of Nrf2 in vascular endothelial cells. This resulted in prevention of oxidative stress-induced cytotoxicity, and may represent a possible mechanism to partly explain the cardioprotective effects of n-3 PUFAs.

Inhibitory effects of orally administrated liposomal bovine lactoferrin on the LPS-induced osteoclastogenesis.

Bovine lactoferrin (bLF) modulates the production of proinflammatory cytokines including tumor necrosis factor (TNF)-alpha, and may thus control alveolar bone destruction associated with periodontitis. In this study, the effects of bLF on mRNA expression in lipopolysaccharide (LPS)-stimulated osteoblasts (OBs) and on LPS-induced osteoclastogenesis were examined. The inhibitory effects of oral administration of liposomal-bLF (L-bLF), which improved the robustness of bLF to digestive enzymes, on alveolar bone resorption using LPS-induced periodontitis rat model are also reported. Three groups of 7-week-old male Wistar rats were treated with L-bLF (L-bLF group), bLF (bLF group), or the vehicle (control group) in drinking water (n=6 in each group). On day 7, LPS was topically applied into the gingival sulcus. Number of osteoclasts and immunoexpression of TNF-alpha were analyzed. The bLF inhibited the upregulation of TNF-alpha-mRNA- and upregulation of receptor activator of NF kappaB (RANKL)-mRNA expression and eliminated downregulation of osteoprotegerin (OPG)-mRNA expression in LPS-stimulated OBs and reduced LPS-induced osteoclastogenesis in co-culture with primary OBs and bone marrow cells. In the control group, the number of osteoclasts increased after LPS treatment. The number of osteoclasts that appeared along the alveolar bone margin was significantly reduced (P<0.01) in the L-bLF but not in the bLF group. Furthermore, L-bLF suppressed upregulation of TNF-alpha immunoexpression in periodontal tissue and TNF-alpha and interleukin (IL)-1 beta-mRNA level in gingival tissue. The results of this study indicate that oral administration of L-bLF significantly reduces alveolar bone resorption induced by LPS stimulation through inhibition of TNF-alpha production and modulation of RANKL/OPG balance in OBs. It is suggested that L-bLF could be a potent therapeutic and preventive agent for attenuating alveolar bone destruction in periodontitis patients.

Soy phosphatidylcholine inhibited TLR4-mediated MCP-1 expression in vascular cells.

Inflammatory signaling via Toll-like receptor 4 (TLR4) has been shown to facilitate atherogenesis. Recent lines of evidence show that saturated fatty acids (SFAs) induce the inflammatory response via the TLR4 pathway in macrophages and adipocytes. The aims of this study are to confirm the role of SFAs in TLR4-mediated inflammatory signaling in vascular cells and to propose soy phosphatidylcholine (SPC) as an effective inhibitor against TLR4-mediated agonists. SFAs such as palmitate and stearate increased the expression and secretion of MCP-1 in human umbilical vein endothelial cells (HUVECs) and rat vascular smooth muscle cells (VSMCs). SFAs up-regulated the activity of MCP-1 promoter through the activation of NF-kappaB. Knockdown of TLR4 using siRNA diminished the SFA-induced MCP-1 expression in HUVECs and rat VSMCs, while PKC or ceramide signal inhibitor did not inhibit the expression. Furthermore, we found that SPC effectively inhibited the MCP-1 expression induced by palmitate or LPS in a dose-dependent manner. However, SPC did not inhibit the mRNA expression of MCP-1 induced by cytokines such as TNF-alpha and IL-1beta, or by agonists binding to TLRs other than TLR4. In addition, SPC did not affect the activity of LPS assessed by clotting activity of the Limulus amoebocyte lysate. These results clearly show that SPC specifically inhibits the inflammatory responses induced by the TLR4-dependent signal. In conclusion, we have demonstrated a role of SFAs for inflammatory response via TLR4-NF-kappaB signaling in vascular cells. Moreover, we propose that SPC can be useful as a selective inhibitor to suppress the TLR4-mediated inflammatory signaling.

Morphological study on dental caries induced in WBN/KobSlc rats (Rattus norvegicus) fed a standard laboratory diet.

In our previous studies, WBN/KobSlc was characterized as a rat strain in which only males began to develop pancreatitis, and then presented with diabetic symptoms. In the course of studying their pancreatic inflammation, we detected molar caries in prediabetic males feeding on a standard diet (CRF-1) widely used for experimental animals. The purpose of this study is to confirm whether the WBN/KobSlc strain is caries-susceptible to the diet reported to be non-cariogenic, and to examine the effect of a prediabetic condition on their dental caries. For a morphological study, 25 male WBN/KobSlc rats aged 3.2-7.8 months and 24 females of the same strain aged 3.3-6.6 months were used, along with 10 males and 10 females of 8.2-month-old F344 rats. Marked dental caries were detected in the mandibular molars of male and female WBN/KobSlc rats regardless of pancreatitis, although no similar changes were observed in any teeth of the F344 strain fed the same diet. Soft X-ray examination revealed that the caries began in the crown and progressed horizontally and vertically, and that a severe radiolucent lesion extensively expanded to the entire crown, corresponding to a macroscopically deleted molar. The caries had gradually developed mainly in the second mandibular molar from more than 3.5 months of age, while none were seen in any rats before that time. The WBN/KobSlc rats were caries-susceptible even to the standard laboratory diet, and pancreatitis was not directly associated with the onset of dental caries in this strain.

Chemoprevention of tumor metastasis by liposomal beta-sitosterol intake.

To investigate chemopreventive effect of liposomal beta-sitosterol on tumor metastasis, we prepared liposomal beta-sitosterol composed of egg yolk phosphatidylcholine for oral delivery. Although orally administered beta-sitosterol (4 micromol as beta-sitosterol/mouse) was not absorbed into plasma, the amount of immune response cytokines such as IL-12 and IL-18 was increased in the small intestine after the liposome intake. Moreover, after daily oral administration of the liposome for 7 d, natural killer (NK) cell activity in the mice was increased, suggesting that the immune surveillance activity of mice was enhanced by the liposomal beta-sitosterol intake. Thus, we examined metastatic potential of B16BL6 melanoma cells, which were intravenously injected into mice after sequential administration of liposomal beta-sitosterol for 7 d. The number of metastatic colonies in the lungs was significantly less than that of control group two weeks after the injections of the cells. These results suggest that daily liposomal beta-sitosterol intake prevents tumor metastasis may be due to enhancement of gut immune surveillance systems.

Liposomalization of lactoferrin enhanced it's anti-inflammatory effects via oral administration.

It is known that lactoferrin is one of the functional proteins contained in mammalian milk and that it plays an important role in the immune system. In this study, we prepared multi-lamellar liposomal bovine lactoferrin composed of egg yolk phosphatidylcholine and phytosterol for oral delivery, and examined any resulting anti-inflammatory effects. Oral pretreatment of liposomal lactoferrin exhibited more suppressive effects than did non-liposomal lactoferrin on CCl4-induced hepatic injury in rats as well as on lipopolysaccharide-induced TNF-alpha production from mouse peripheral blood mononuclear leukocytes. Further investigation revealed that the liposomalization did not exert influence on the absorbability of lactoferrin to the venous blood or lymph following an intraduodenal administration in rats. Furthermore, there was no significant difference exhibited between the antigenicity of liposomal and non-liposomal lactoferrin, which was measured using the passive cutaneous anaphylaxis reaction following oral sensitization to them in guinea pigs. These results suggest that liposomal lactoferrin might act more effectively than conventional lactoferrin in the intestinal site, which is regarded as an active site of orally administered lactoferrin, although the biological mechanism is not fully understood yet. Consequently we propose that liposomal lactoferrin could be a novel active constituent useful for preventive and therapeutic treatment of inflammatory diseases.

Suppressed permeation of linoleic acid in a liposomal formulation through reconstructed skin tissue.

Since the liposomal formulation of linoleic acid (LA) exhibited an enhanced skin-whitening effect, the influence of liposomalization on the cutaneous absorption of LA was examined using a three-dimensional (3D) reconstructed skin model. Liposome entrapped [(14)C]-LA was applied on the skin model, and the permeation of LA through the skin was monitored. The permeation rate of LA in the liposomal formulation was found to be lower than that in the conventional formulation without liposomes, suggesting the increased retention time of LA in the skin by the liposomal formulation. Next, to investigate the dependence of the LA permeation on melanocyte conditions and intactness of the reconstructed skin model, the effect of UV irradiation on LA permeation was examined. Low-dose UVB irradiation (0.03 J/cm(2) for 3 times), which activated melanocytes in the skin, did not influence the extent of LA permeation, while high-dose irradiation (0.30 J/cm(2) for 3 times) enhanced the permeation of LA in both the conventional and liposomal formulation. The present results suggest the importance of skin intactness for LA permeation and that the 3D reconstructed skin model would be useful for evaluating the characteristics of skin-oriented cosmetics and drugs.

Skin whitening effect of linoleic acid is enhanced by liposomal formulations.

Linoleic acid (LA) is known to have a whitening effect on hyperpigmented skin, and is encapsulated in liposomes for topical application because of its low solubility in aqueous solution, although the effect of liposomalization of LA on the whitening activity has not been evaluated. In the present study, we evaluated the effect of liposomalization on the whitening activity of LA by using LA in ethanol, hydrogel containing LA, and hydrogel containing liposomal LA towards the UV-stimulated hyperpigmented dorsal skin of brownish guinea pigs. The whitening effect was far greater for hydrogel containing liposomal LA (0.1% w/w as a final concentration of LA) than for free LA in ethanol or hydrogel containing LA. Next, the whitening effect of LA was examined with UV-stimulated hyperpigmented human upper arm skin by using a hydrogel containing liposomal LA (0.1% LA) and non-liposomal LA (3.0, 10.0% LA). Liposomal LA (0.1%) showed a whitening effect comparable to 10.0% non-liposomal LA and was far more effective than 3.0% non-liposomal LA. These results indicate that liposomal formulations are favorable for the transdermal application of LA.

Liposomal lactoferrin induced significant increase of the interferon-alpha (IFN-alpha) producibility in healthy volunteers.

Interferon-alpha (IFN-alpha) producibility has been widely accepted as one of the important markers to evaluate the immune status. In this study, preliminary clinical tests were carried out to confirm the immunomodulatory activity of liposomal lactoferrin including IFN-alpha producibility and NK activity. In a primary open trial, the liposomal lactoferrin was administered to five healthy males for one week and various immunological indices were evaluated. Furthermore, ten healthy males were administered 319 mg per day of liposomal or non-liposomal lactoferrin for four weeks, and immune status was monitored at 0, 1 and 4 weeks after the intake as well as three weeks after stopping it. In this double-blinded comparative study, the IFN-alpha producibility was significantly increased only in the liposomal lactoferrin group during administration and decreased 3 weeks after stopping it, while the IFN-alpha producibility was unchanged in the non-liposomal lactoferrin group. Although the biological mechanism of IFN-alpha producibility enforced by liposomal lactoferrin has not been wholly understood, it is suggested to be a novel active constituent having preventive and therapeutic effects on inflammatory diseases, cancer and infectious diseases such as chronic hepatitis C.

[LDL-cholesterol-lowering effect of a mixed green vegetable and fruit beverage containing broccoli and cabbage in hypercholesterolemic subjects].

The serum LDL-cholesterol (LDL-C)-lowering effects of two types of canned beverage containing mixed green vegetables and fruits, with or without broccoli and cabbage (B&C), were examined in a randomized double-blind study design. Seventy-seven adults subjects with mild to moderate hypercholesterolemia participated in this study after giving their informed consent. The subjects were randomly divided into two groups. One group(test group) was allocated a test sample, containing B&C as the main materials. Another group(control group) was allocated a placebo sample made from the same materials but without B&C. The subjects were administered 2 cans of the assigned sample (160 g contents/can) per day for 12 weeks. Forty-nine out of 77 subjects, whose LDL-C levels were greater than or equal to 140 mg/dl and less than 180 mg/dl, were analyzed for the effectiveness. Serum LDL-C levels in the test group were significantly(p < 0.05) reduced at 3, 6 and 9 weeks after administration in comparison to the baseline levels (155.7 mg/dl in average). The average LDL-C value at 9 weeks was 142.5 mg/dl and the reduction rate was 8.5%. But serum LDL-C levels in the control group were not significantly reduced. Significant differences(p < 0.05) between the groups were observed in the LDL-C levels at 6 and 9 weeks and also in the total cholesterol levels at 9 week. Thus daily intake of the beverage tested containing B&C are useful for lowering serum LDL-C levels in hypercholesterolemic subjects.