Curcuma longa extract reduces serum inflammatory markers and postprandial hyperglycemia in healthy but borderline participants with overweight and glycemia in the normal/prediabetes range: a randomized, double-blind, and placebo-controlled trial.

The spice turmeric, which has the Latin name Curcuma longa (C. longa), has various physiological effects. This study evaluated the effects of a hot water mixture with supercritical carbon dioxide C. longa extracts, CLE, and the potential active components of C. longa, turmeronols A and B and bisacurone on inflammation and glucose metabolism. First, we investigated the effect of CLE and the potential active components of C. longa on lipopolysaccharide-induced inflammation in RAW264.7 macrophages. We found a significant decrease in the production of interleukin (IL)-1ß, IL-6, tumor necrosis factor (TNF)-α, and nitric oxide with CLE, turmeronol A, and bisacurone, Significant inhibition of each of these substances was also observed, except for TNF-α with turmeronol B. The second part of our work was a 12-week randomized, double-blind, placebo-controlled study in healthy but borderline adults aged 40 to 69 years with overweight and normal/prediabetes glycemia. We compared blood inflammatory and glycometabolic markers in the CLE (n = 55) and placebo groups (n = 55). We found significantly lower serum high-sensitivity C-reactive protein and hemoglobin A1c levels in the CLE group. This group also showed significant improvements in postprandial hyperglycemia and insulin sensitivity indices. Our findings indicate that CLE may reduce low-grade inflammation and thus improve insulin sensitivity and postprandial hyperglycemia. Clinical trial registration: https://center6.umin.ac.jp/cgi-open-bin/ctr/ctr_view.cgi?recptno=R000051492, UMIN-CTR, UMIN000045106.

Turmeronols (A and B) from Curcuma longa have anti-inflammatory effects in lipopolysaccharide-stimulated BV-2 microglial cells by reducing NF-κB signaling.

Turmeronols (A and B), bisabolane-type sesquiterpenoids found in turmeric, reduce inflammation outside the brain in animals; however, their effects on neuroinflammation, a common pathology of various neurodegenerative diseases, are not understood. Inflammatory mediators produced by microglial cells play a key role in neuroinflammation, so this study evaluated the anti-inflammatory effects of turmeronols in BV-2 microglial cells stimulated with lipopolysaccharide (LPS). Pretreatment with turmeronol A or B significantly inhibited LPS-induced nitric oxide (NO) production; mRNA expression of inducible NO synthase; production of interleukin (IL)-1ß, IL-6, and tumor necrosis factor α and upregulation of their mRNA expression; phosphorylation of nuclear factor-κB (NF-κB) p65 proteins and inhibitor of NF-κB kinase (IKK); and nuclear translocation of NF-κB. These results suggest that these turmeronols may prevent the production of inflammatory mediators by inhibiting the IKK/NF-κB signaling pathway in activated microglial cells and can potentially treat neuroinflammation associated with microglial activation.

Corrigendum: Beneficial effect of heat-killed Lactiplantibacillus plantarum L-137 on skin functions in healthy participants: A randomized, placebo-controlled, double-blind study.

[This corrects the article DOI: 10.3389/fmed.2022.912280.].

A Hot Water Extract of Curcuma longa L. Improves Fasting Serum Glucose Levels in Participants with Low-Grade Inflammation: Reanalysis of Data from Two Randomized, Double-Blind, Placebo-Controlled Trials.

The dietary spice Curcuma longa L. (C. longa), also known as turmeric, has various biological effects. A hot water extract of C. longa was shown to have anti-inflammatory activities in preclinical and clinical studies. Chronic low-grade inflammation is associated with the disruption of glucose homeostasis, but the effect of C. longa extract on glucose metabolism in humans is poorly understood. Therefore, we investigated the effect of C. longa extracts on serum glucose levels in the presence of low-grade inflammation. We reanalyzed our published data from two randomized, double-blind, placebo-controlled trials in overweight participants aged 50 to 69 years and performed a stratified analysis using the inflammatory marker high-sensitivity C-reactive protein (hsCRP). In both studies, participants took a test food with a hot water extract of C. longa (C. longa extract group, n = 45 per study) or without C. longa extract (placebo group, n = 45 per study) daily for 12 weeks, and we measured the levels of serum hsCRP and fasting serum glucose. The mean baseline hsCRP value was used to stratify participants into two subgroups: a low-hsCRP subgroup (baseline mean hsCRP < 0.098 mg/dL) and a high-hsCRP subgroup (baseline mean hsCRP ≥ 0.098 mg/dL). In the low-hsCRP subgroup, we found no significant difference in fasting serum glucose levels between the two groups in either study, but in the high-hsCRP subgroup, the C. longa extract group had significantly lower levels of serum hsCRP (p < 0.05) and fasting serum glucose (p < 0.05) than the placebo group in both studies. In conclusion, a hot water extract of C. longa may help to improve systemic glucose metabolism in people with chronic low-grade inflammation.

Beneficial Effect of Heat-Killed Lactiplantibacillus plantarum L-137 on Skin Functions in Healthy Participants: A Randomized, Placebo-Controlled, Double-Blind Study.

To determine whether consuming heat-killed Lactiplantibacillus plantarum L-137 (HK L-137) influences skin functions, we performed a randomized, placebo-controlled, double-blind study in healthy participants who were conscious of dry skin. A total of 80 healthy participants (20 men, 60 women; mean age, 47.3 years) were assigned to receive a tablet containing HK L-137 or a placebo tablet daily for 12 weeks. Every 4 weeks, the skin water content and transepidermal water loss (TEWL) were measured at the forearm and face, and participants completed two skin-related questionnaires, the Dermatology Life Quality Index and a self-evaluation. The HK L-137 group tended to show greater increases from baseline of water content at the forearm and larger decreases of TEWL at the face. The total scores of both questionnaires improved significantly more in the HK L-137 group. Water content and TEWL improved significantly in participants in the HK L-137 group who were above the median age of study participants or had relatively dry skin. These findings suggest that daily HK L-137 intake can improve dry skin, thereby contributing to skin satisfaction.

Curcuma longa extract improves serum inflammatory markers and mental health in healthy participants who are overweight: a randomized, double-blind, placebo-controlled trial.

BACKGROUND: The dietary spice Curcuma longa, also known as turmeric, has various biological effects. Both a water extract and a supercritical carbon dioxide extract of C. longa showed anti-inflammatory activities in animal studies. However, the anti-inflammatory effect in humans of a mixture of these two C. longa extracts (CLE) is poorly understood. Therefore, we investigated the effect of CLE containing anti-inflammatory turmeronols on chronic inflammation and general health. METHODS: We performed a randomized, double-blind, placebo-controlled study in healthy subjects aged 50 to 69 years with overweight. Participants took two capsules containing CLE (CLE group, n = 45) or two placebo capsules (placebo group, n = 45) daily for 12 weeks, and serum inflammatory markers were measured. Participants also completed two questionnaires: the Medical Outcomes Study (MOS) 36-Item Short-Form Health Survey (SF-36) and the Profile of Mood States (POMS) scale. Treatment effects were analyzed by two way analysis of variance followed by a t test (significance level, p <  0.05). RESULTS: After the intervention, the CLE group had a significantly lower body weight (p <  0.05) and body mass index (p < 0.05) than the placebo group and significantly lower serum levels of C-reactive protein (p < 0.05) and complement component 3 (p < 0.05). In addition, the CLE group showed significant improvement of the MOS SF-36 mental health score (p < 0.05) and POMS anger-hostility score (p < 0.05). CONCLUSION: CLE may ameliorate chronic low-grade inflammation and thus help to improve mental health and mood disturbance. TRIAL REGISTRATION: UMIN-CTR, UMIN000037370. Registered 14 July 2019, https://upload.umin.ac.jp/cgi-open-bin/ctr/ctr_view.cgi?recptno=R000042607.

Safety and efficacy of using heat-killed Lactobacillus plantarum L-137: High-dose and long-term use effects on immune-related safety and intestinal bacterial flora.

Heat-killed Lactobacillus plantarum L-137 (HK L-137) promotes immune function in animals. In healthy people, T-cell proliferation was shown to be enhanced by taking 10 mg HK L-137 daily for 12 weeks. However, the safety and efficacy of higher doses or longer treatments have not yet been investigated in humans. To investigate the high-dose and long-term use effects of HK L-137 on immune-related safety and on host intestinal bacterial flora, 15 healthy volunteers took a daily HK L-137 (50 mg) preparation for 4 weeks. An additional 29 participants who regularly visited a clinic for health care took HK L-137 (10 mg) daily for 12 months. Measures for anthropometrics, hematology, biochemistry, and urinalysis were taken at scheduled timepoints for all participants. Stool and blood samples were also collected and evaluated for microbes and short-chain fatty acids (SCFA); isolated T-cells were assessed for levels of proliferation induced by phytohemagglutinin in the long-term study. Adverse events or shifts in clinical measures from normal ranges due to the dietary intervention were not observed in the high-dose or long-term studies. Long-term intake also did not result in immune exhaustion due to any chronic immunostimulation; ex vivo T-cell proliferation was significantly greater at 12 months than at baseline (p < 0.01). In addition, the Firmicutes/Bacteroidetes ratio in stool samples was significantly lower at 12 months than at baseline (p < 0.05) due to the long-term intake of the HK L-137. Lastly, fecal SCFA concentrations were significantly greater (p < 0.05) at 6 months than at baseline. From these data, it can be concluded that the efficacy of HK L-137 is maintained with no overt adverse effects as a result of high-dose and/or long-term consumption.

Heat-killed Lactobacillus plantarum L-137 attenuates obesity and associated metabolic abnormalities in C57BL/6†J mice on a high-fat diet.

Heat-killed Lactobacillus plantarum L-137 (HK L-137) has anti-allergic, antitumor, and antiviral effects in mice, as well as an anti-inflammatory effect in rats with metabolic syndrome through regulation of immunity. To evaluate the influence of HK L-137 on chronic inflammation in mice with diet-induced obesity, C57BL/6 J mice were fed a normal diet (16% of energy as fat) or a high-fat diet (62% of energy as fat) with or without 0.002% HK L-137 for 4 to 20 weeks. It was found that HK L-137 supplementation alleviated weight gain and elevation of plasma glucose, cholesterol, alanine aminotransferase, and aspartate transaminase levels in mice with diet-induced obesity. Expression of several inflammation-related genes, including F4/80, CD11c, and IL-1ß, in the epididymal adipose tissue of these mice was significantly downregulated by HK L-137. In addition, plasma levels of lipopolysaccharide-binding protein, a marker of endotoxemia, tended to be decreased by administration of HK L-137. These findings suggest that HK L-137 supplementation ameliorates obesity-induced metabolic abnormalities and adipose tissue inflammation, possibly through improvement of intestinal permeability.

Daily intake of heat-killed Lactobacillus plantarum L-137 improves inflammation and lipid metabolism in overweight healthy adults: a randomized-controlled trial.

PURPOSE: The effects of heat-killed Lactobacillus plantarum L-137 (HK L-137) on inflammation and lipid metabolism were investigated in overweight volunteers. METHODS: One hundred healthy subjects with a body mass index from 23.0 to 29.9 (51 men and 49 women; mean age: 41.4 years) were enrolled in this randomized, double-blind, placebo-controlled, parallel group study. Subjects were randomly assigned to daily administration of a tablet containing HK L-137 (10 mg) or a placebo tablet for 12 weeks. Blood samples were collected every 4 weeks to measure biomarkers of lipid metabolism and inflammatory mediators. RESULTS: The percent change of concanavalin A-induced proliferation of peripheral blood mononuclear cells was significantly larger in the HK L-137 group than in the control group, similar to previous studies. The decreases of aspartate aminotransferase and alanine aminotransferase over time were significantly larger in the HK L-137 group than in the control group, as were the decreases of total cholesterol, low-density lipoprotein cholesterol, and the leukocyte count at one time point. These effects of HK L-137 were stronger in the subjects with higher C-reactive protein levels. CONCLUSIONS: These findings suggest that daily intake of HK L-137 can improve inflammation and lipid metabolism in subjects at risk of inflammation.

High dietary vitamin C intake reduces glucocorticoid-induced immunosuppression and measures of oxidative stress in vitamin C-deficient senescence marker protein 30 knockout mice.

Vitamin C (VC) is a vital micronutrient for humans and some other mammals and also has antioxidant activity. Stress-induced elevation of glucocorticoid production is well known to cause immunosuppression. The present study evaluated the effect of high VC intake on glucocorticoid-induced immune changes in mice. Senescence marker protein 30 knockout mice with genetic VC deficiency were fed a diet containing the recommended VC content (20 mg/kg per d; 0·02 %VC group) or a high VC content (200 mg/kg per d; 0·2 %VC group) for 2 months, then dexamethasone was given by intraperitoneal injection. After administration of dexamethasone, the plasma ascorbic acid concentration decreased significantly in the 0·02 %VC group and was unchanged in wild-type C57BL/6 mice on a VC-deficient diet (wild-type group), while it was significantly higher in the 0·2 %VC group compared with the other two groups. In the 0·02 %VC and wild-type groups, dexamethasone caused a significant decrease in the cluster of differentiation (CD)4+ and CD8+ T cells among splenocytes as well as a significant decrease in IL-2, IL-12p40 and interferon-γ protein production by splenocytes and a significant decrease in T-cell proliferation among splenocytes. In the 0·2 %VC group, these dexamethasone-induced immunosuppression improved when compared with the other two groups. In addition, reduction in the intracellular levels of ascorbic acid, superoxide dismutase and glutathione in splenocytes by dexamethasone as well as elevation in thiobarbituric acid-reactive substances were significantly suppressed in the 0·2 %VC group. These findings suggest that high dietary VC intake reduces glucocorticoid-induced T-cell dysfunction by maintaining intracellular antioxidant activity.

Lactobacillus plantarum L-137 upregulates hyaluronic acid production in epidermal cells and fibroblasts in mice.

Heat-killed Lactobacillus plantarum L-137 (HK L-137), an immunobiotic lactic acid bacterium, has been reported to enhance IFN-γ production through induction of IL-12. In this study, we investigated the effects of HK L-137 on skin moisturizing and production of hyaluronic acid (HA), an extracellular matrix associated with the retention of skin moisture. Oral administration of HK L-137 suppressed the loss of water content in the stratum corneum in hairless mice. Treatment of primary epidermal cells with HK L-137 increased HA production. Supernatant from immune cells stimulated by HK L-137, which contained proinflammatory cytokines such as IL-12, TNF-α, and IFN-γ, upregulated HA production and hyaluronan synthase 2 (HAS2) messenger RNA expression by BALB/3T3 fibroblasts via activation of transcription factor nuclear factor κB (NFκB). Although treatment of the supernatant with anti-TNF-α antibody (Ab) alone did not inhibit the HA production, combination of anti-TNF-α Ab with anti-IFN-γ Ab significantly inhibited the HA production. Thus, HK L-137-induced IFN-γ plays a critical role in upregulated HA production in collaboration with TNF-α. HK L-137 may be useful for improvement of skin functions such as moisture retention by inducing HA production.

Daily Intake of Heat-killed Lactobacillus plantarum L-137 Decreases the Probing Depth in Patients Undergoing Supportive Periodontal Therapy.

PURPOSE: Heat-killed Lactobacillus plantarum L-137 (HK L-137) has been shown to activate innate and acquired immunity in humans. The aim of this randomised, double-blind, placebo-controlled clinical trial was to examine the effects of the oral administration of HK L-137 on the outcome of periodontal therapy. MATERIALS AND METHODS: Thirty-nine patients undergoing supportive periodontal therapy (SPT) were randomly assigned to receive a capsule containing 10 mg of HK L-137 or a placebo capsule daily for 12 weeks. Nineteen patients in the experimental group and 17 patients in the control group were followed-up. Clinical parameters, including plaque index (PI), gingival index (GI), bleeding on probing (BOP), and probing depth (PD) were scored at baseline and weeks 4, 8 and 12 prior to prophylaxis in conjunction with regular SPT visits. RESULTS: BOP and the number of teeth or sites with PD ≥ 4 mm were significantly reduced in both groups by a successive SPT programme, while there was significantly greater PD reduction (p < 0.05) at teeth with site(s) with PD ≥ 4 mm at baseline in the experimental group than in the control group at week 12. CONCLUSION: These clinical findings suggest that daily HK L-137 intake can decrease the depth of periodontal pockets in patients undergoing supportive periodontal therapy.

Scavenger receptor for lipoteichoic acid is involved in the potent ability of Lactobacillus plantarum strain L-137 to stimulate production of interleukin-12p40.

Heat-killed Lactobacillus plantarum strain L-137 (HK L-137) is a more potent inducer of interleukin (IL)-12 than other heat-killed Lactobacillus strains. To elucidate the mechanism involved in this IL-12p40 induction, we compared HK L-137 with heat-killed L. plantarum strain JCM1149 (HK JCM1149) by nuclear magnetic resonance and mass spectrometry. Results showed that HK L-137 contained lipoteichoic acid (LTA) with a chemical structure similar to that of JCM1149, except for a lower degree of glucosyl substitution in the poly(glycerol phosphate) backbone. Lysozyme sensitivity and electrophoretic moiety analysis revealed that HK L-137 exposed more LTA on its cell surface than HK JCM1149. Phagocytosis of HK L-137 by splenic adherent cells was significantly greater than that of HK JCM1149. Anti-LTA antibody and anti-scavenger receptor-A (SR-A) antibody selectively inhibited phagocytosis of HK L-137, as well as IL-12p40 production, by splenic adherent cells. Thus, a higher efficiency of phagocytosis of HK L-137 via SR-A for LTA is responsible for the potent IL-12p40 induction.

High dietary intake of vitamin C suppresses age-related thymic atrophy and contributes to the maintenance of immune cells in vitamin C-deficient senescence marker protein-30 knockout mice.

Vitamin C (VC) is an essential nutrient for humans and certain other animals. It has antioxidant properties and has been reported to ameliorate oxidative damage to lipids, DNA and proteins. However, the effects of VC on immune function are poorly understood, especially the influence of long-term high-dose VC intake on the number and function of immune cells. In the present study, to evaluate the immune effects of VC, VC-deficient senescence marker protein-30 knockout (SMP30KO) mice were fed a diet containing the recommended level of VC (20 mg/kg per d; 0·02 % VC) or a high level of VC (200 mg/kg per d; 0·2 % VC) for 1 year. The plasma VC concentration of the 0·02 % group was the same as that of age-matched C57BL/6 mice after 1 year of feeding; however, plasma VC concentration and thymus weight were significantly higher in the 0·2 % VC group than in the 0·02 % VC group. The total counts of leucocytes, lymphocytes, granulocytes and monocytes in the peripheral blood, as well as the number of splenocytes and thymocytes, were all significantly higher in the 0·2 % VC group than in the 0·02 % VC group. In addition, the number of naive T cells in peripheral blood lymphocytes, the number of memory T-cell populations in splenocytes, and the number of cluster of differentiation (CD)4⁺CD8⁺ or CD4⁺CD8⁻ or CD4⁻CD8⁺ T cells in thymocytes were all markedly higher in the 0·2 % VC group than in the 0·02 % VC group after 1 year of dietary treatment. These results suggest that a long-term high-dose intake of VC is effective in the maintenance of immune cells, partly through the suppression of age-related thymic involution in VC-deficient SMP30KO mice.

Oral intake of heat-killed Lactobacillus plantarum L-137 decreases the incidence of upper respiratory tract infection in healthy subjects with high levels of psychological stress.

The immunomodulatory effects of live or non-viable lactic acid bacteria have been extensively investigated. We reported that oral intake of heat-killed Lactobacillus plantarum L-137 (HK L-137) augmented innate and acquired immunity in mice and human subjects. To examine the effects of HK L-137 intake on upper respiratory tract infection (URTI) symptoms and immune functions in human subjects, a randomised, double-blind, placebo-controlled, parallel study was conducted in subjects with high psychological stress levels. A total of seventy-eight healthy subjects (thirty-three men and forty-five women; mean age 50·6 years) with scores of >41 on eighteen-item subscales of psychological distress in the Brief Job Stress Questionnaire were randomly assigned to receive a tablet containing HK L-137 (10 mg) or a placebo tablet daily for 12 weeks. The URTI symptoms were rated once daily on the validated twenty-one-item Wisconsin Upper Respiratory Symptom Survey-21. Immune functions, such as concanavalin A-induced proliferation and percentages of interferon (IFN)-γ- and IL-4-producing CD4 T cells of peripheral blood mononuclear cells (PBMC), and serum IFN-ß concentrations were measured every 4 weeks. URTI incidence was significantly lower in the HK L-137 group than in the control group. URTI incidence, duration and severity, and duration of medication showed significant negative correlations with duration of HK L-137 intake. The percentage change from baseline of concanavalin A-induced proliferation of PBMC was significantly greater in the HK L-137 group than in the control group. These findings suggest that daily HK L-137 intake can decrease URTI incidence in healthy subjects, possibly through augmentation of immune functions.

Enhanced immunomodulatory activity and stability in simulated digestive juices of Lactobacillus plantarum L-137 by heat treatment.

This study reports the effect of heat treating Lactobacillus plantarum L-137 on its in vitro cytokine-inducing activity, on the stability of this activity in simulated digestive juices, and on its in vivo immunomodulatory properties. L-137 cells were harvested at the stationary phase with or without the subsequent heat treatment and then lyophilized. Heat-killed L-137 cells stimulated mouse spleen cells to produce more interleukin-12p40 than unheated L-137. The interleukin-12p40-inducing activity of unheated L-137 was significantly lower when incubated with simulated intestinal juice, but the activity of heat-killed L-137 cells was maintained. Furthermore, heat-killed L-137 was more protective than unheated L-137 in a mouse model of dextran sulfate sodium-induced colitis. A heat treatment may therefore be effective for enhancing the immunomodulatory activity of L-137 cells.

Daily intake of heat-killed Lactobacillus plantarum L-137 enhances type I interferon production in healthy humans and pigs.

We have previously reported that oral administration of heat-killed Lactobacillus plantarum L-137 (HK L-137) stimulates innate immunity for production of type I interferon (IFN) which subsequently augments host defense against influenza A virus infection in mice. We here examined the effect of HK L-137 intake on type I IFN in humans. Sixteen subjects were randomly assigned to receive a tablet containing 10 mg of HK L-137 or a matching tablet for 8 weeks and the serum levels of type I IFN were examined before and after the first or second dose of the trivalent inactivated influenza vaccine. There were no differences in the seroresponse rate, the seroprotection rate and the geometric mean Ab titers after either the first or second dose of vaccine between the HK L-137 group and the control group. On the other hand, the levels of IFN-ß were significantly higher in the HK L-137 group than in the control group before vaccination although the vaccination conferred little additional induction of IFN-ß. We further examined IFN-ß gene expression in the whole blood cells of pigs fed on a diet containing HK L-137 and found that the IFN-ß mRNA levels were significantly higher in the HK L-137 group than in the control group. The finding that daily intake of HK L-137 enhances type I IFN production and host defense against influenza A virus infection in mice may be applied to at least two additional species.

Lipoteichoic acids on Lactobacillus plantarum cell surfaces correlate with induction of interleukin-12p40 production.

Heat-killed cells of Lactobacillus plantarum L-137 are potent inducers of IL-12 in vitro as well as in vivo and have been shown to have antiallergic, antitumor, and antiviral effects through this induction, which leads to a Th1 type immune response. To determine why L-137 cells induce much greater IL-12 production than the type strain Lactobacillus plantarum JCM1149, we examined the differences in their CW components. The L-137 CW was found to have a higher alanine content and IL-12p40 induction was significantly greater in comparison with JCM1149 CW, whereas peptidoglycans isolated from both strains did not cause IL-12p40 induction. Because in purified CW preparations from gram-positive bacteria, the presence of LTA, the major proinflammatory structure on these bacteria, has been known to have high alanine content, we investigated the responsiveness of both strains to anti-LTA antibody by flow cytometry. L-137 cells reacted more with anti-LTA antibody than did JCM1149 cells. Furthermore, derivative strains of L-137, cured of a specific plasmid pLTK11 of the 15 endogenous plasmids in wild-type L-137, had poor responsiveness to anti-LTA antibody and showed lower IL-12p40 inducing activity than the wild-type L-137 with pLTK11. Our results suggest that LTA expression on the cell surface causes IL-12p40 induction, and that the above internal plasmid of L-137 influences LTA synthesis and expression on the cell surface.

Oral administration of heat-killed Lactobacillus plantarum L-137 enhances protection against influenza virus infection by stimulation of type I interferon production in mice.

We have previously reported that heat-killed Lactobacillus plantarum L-137 (HK-LP) stimulates macrophage/dendritic cells to produce T helper (Th) 1-related cytokines in vitro and in vivo in mice. We here examined the effect of oral administration of HK-LP on protection against influenza virus infection in mice. C57BL/6 mice were orally given HK-LP from day -7 to 7 and intranasally infected with influenza virus A/FM/1/47 (H1N1, a mouse-adapted strain) at 100 pfu on day 0. The survival time was significantly prolonged in mice treated with HK-LP than that in mice treated with PBS as controls. The viral titers in the lung were significantly lower in mice treated with HK-LP than controls at the early stage after influenza virus infection. An appreciable level of interferon (IFN)-beta was detected in the serum of mice treated with HK-LP, while no IFN-beta was detected in controls after influenza infection. Our results suggest that HK-LP, a potent IFN-beta inducer, is useful for prevention against influenza infection.

A nonradioisotope, enzymatic microplate assay for in vivo evaluation of 2-deoxyglucose uptake in muscle tissue.

To determine 2-deoxy-D-glucose (2DG) and 2-deoxy-D-glucose 6-phosphate (DG6P) in mouse tissue after injection of 2DG, we have developed a novel assay. This assay is a simple procedure involving incubation of samples with four independent, single reaction mixtures followed by measurement of fluorescence. From differences between the values obtained with the four reactions, each of glucose, glucose 6-phosphate, 2DG and DG6P were able to be quantified in a sensitive manner. Using this assay system, glucose and 2DG in blood and DG6P-accumulation in muscle were easily determined. Therefore, this assay may be useful for measuring in vivo glucose uptake without the use of radioisotopes.