WO2007040444A1 - Method, device and computer program product for diagnosing of at least one exhaust emission control unit - Google Patents

Method, device and computer program product for diagnosing of at least one exhaust emission control unit Download PDF

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Publication number
WO2007040444A1
WO2007040444A1 PCT/SE2006/001137 SE2006001137W WO2007040444A1 WO 2007040444 A1 WO2007040444 A1 WO 2007040444A1 SE 2006001137 W SE2006001137 W SE 2006001137W WO 2007040444 A1 WO2007040444 A1 WO 2007040444A1
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Prior art keywords
lactobacillus
kit according
cells
compositions
ixio
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PCT/SE2006/001137
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English (en)
French (fr)
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Jan Alenfall
Anna Berggren
Carola Rask
Agnes Wold
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Probi Ab
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
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    • A—HUMAN NECESSITIES
    • A21—BAKING; EDIBLE DOUGHS
    • A21D—TREATMENT OF FLOUR OR DOUGH FOR BAKING, e.g. BY ADDITION OF MATERIALS; BAKING; BAKERY PRODUCTS
    • A21D13/00—Finished or partly finished bakery products
    • A21D13/40—Products characterised by the type, form or use
    • A—HUMAN NECESSITIES
    • A21—BAKING; EDIBLE DOUGHS
    • A21D—TREATMENT OF FLOUR OR DOUGH FOR BAKING, e.g. BY ADDITION OF MATERIALS; BAKING; BAKERY PRODUCTS
    • A21D13/00—Finished or partly finished bakery products
    • A21D13/02—Products made from whole meal; Products containing bran or rough-ground grain
    • A—HUMAN NECESSITIES
    • A21—BAKING; EDIBLE DOUGHS
    • A21D—TREATMENT OF FLOUR OR DOUGH FOR BAKING, e.g. BY ADDITION OF MATERIALS; BAKING; BAKERY PRODUCTS
    • A21D13/00—Finished or partly finished bakery products
    • A21D13/06—Products with modified nutritive value, e.g. with modified starch content
    • A—HUMAN NECESSITIES
    • A21—BAKING; EDIBLE DOUGHS
    • A21D—TREATMENT OF FLOUR OR DOUGH FOR BAKING, e.g. BY ADDITION OF MATERIALS; BAKING; BAKERY PRODUCTS
    • A21D2/00—Treatment of flour or dough by adding materials thereto before or during baking
    • A21D2/08—Treatment of flour or dough by adding materials thereto before or during baking by adding organic substances
    • A21D2/24—Organic nitrogen compounds
    • A21D2/26—Proteins
    • A21D2/267—Microbial proteins
    • A—HUMAN NECESSITIES
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    • A21D8/00—Methods for preparing or baking dough
    • A21D8/02—Methods for preparing dough; Treating dough prior to baking
    • A21D8/04—Methods for preparing dough; Treating dough prior to baking treating dough with microorganisms or enzymes
    • A21D8/045—Methods for preparing dough; Treating dough prior to baking treating dough with microorganisms or enzymes with a leaven or a composition containing acidifying bacteria
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING OR TREATMENT THEREOF
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    • A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
    • A23C9/123—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
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    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
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    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
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    • C12N1/20—Bacteria; Culture media therefor
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    • A—HUMAN NECESSITIES
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    • A23V2400/00—Lactic or propionic acid bacteria
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    • C—CHEMISTRY; METALLURGY
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    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
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    • Y10S435/853—Lactobacillus
    • Y10S435/857—Lactobacillus plantarum

Definitions

  • the present invention relates to a kit comprising at least two compositions, wherein each of said compositions comprises at least one strain chosen from Lactobacillus.
  • the invention further relates to a method for administration of said kit, wherein each of said compositions of the kit is administered in a sequence or in an alternating sequence or in a combination thereof.
  • Probiotoic bacteria are defined as live microorganisms which when administered in adequate amounts beneficially affect the host. Lactobacilli and bifidobacteria are the most frequently used bacteria in probiotic products. These bacteria are generally safe, as are probiotics based on these organisms. The lack of pathogenicity extends across all age groups and to immunocompromised individuals.
  • a method to expand and/or increase the functional activity of regulatory T cells has been studied, thereby reducing the risk of developing immunoregulatory diseases such as allergy, autoimmune diseases (type 1-diabetes and multiple sclerosis) and inflammatory bowel disease.
  • immunoregulatory diseases such as allergy, autoimmune diseases (type 1-diabetes and multiple sclerosis) and inflammatory bowel disease.
  • the immune system is activated in a manner that leads to formation of functionally active regulatory T cells. These cells can then down-regulate potentially harmful immune mediated diseases, such as allergy.
  • An object of the present invention is a kit comprising at least two compositions, wherein each of said compositions comprises at least one strain chosen from Lactobacillus.
  • Another object of the present invention is a method for administration of above mentioned kit, wherein each of said compositions of the kit is administered in a sequence or in an alternating sequence or in a combination thereof.
  • Figure 1 shows the numbers of volunteers reporting any minor adverse gastrointestinal effects during the trial.
  • Figure 2 shows base line numbers (day 0) of different lymphocytes per ml blood (mean ⁇ (SEM) )
  • Figure 3 shows base line (day 0) percentages or GMFI (mean + (SEM) ) of lymphocytes positive for different cell activation and memory markers.
  • FIG. 4 Subjects were randomly assigned to nine different study groups. The trial started with a wash out period of two weeks. Thereafter, the active study period followed. During this period, the subjects consumed one dose of study product per day for 14 (L. Heal 19, L. fermentum, L. paracasel, L. gasseri, L. rhamnosus, P. lundensis groups) or 35 days (L. plantarum and placebo group) . Each dose contained 10 10 coloni forming units (CFU) ⁇ lactobacilli groups) or 10 9 CFU bacteria (P. lundensis group) . Subjects were supplied with a list of probiotic products, which should not be consumed during the trial. A diary, in which each subject stated adverse effects, health conditions and confirmed intake of study product, was kept during the trial.
  • CFU coloni forming units
  • FIG. 1 Percentages of lymphocytes expressing the activation phenotypes CD3CD8CD25, CD3CD8HLA-DR, CD3CD4CD25 'and CD3CD4HLA-DR was analysed by flowcytometry . Group means ( ⁇ SEM) based on individual ratios, day 14/day 0 and day 35/day 0 (for L. plantarum and placebo group only) is shown.
  • FIG. 6 Percentages of lymphocytes expressing the memory phenotypes CD3CD8CD45RO and CD3CD4CD45RO was analysed by flowcytometry .
  • FIG. 7 Percentages of lymphocytes positive for the NKT cell markers (CD56CD16CD3) was analysed by flowcytometry . Group calculations are based on individual ratios (day 14/day 0) .
  • FIG. 8 The phagocytic activity of neutrophils was analysed by incubating whole blood cells with FITC- labelled E. coli or S. aureus. The ratio between mean fluorescence values obtained at day 14 and day 0 was determined individually and group calculations are shown in this figure.
  • Figure 9 shows the ratio of lymphocytes expressing the activation phenotypes CD4CD25 from experiment 2.
  • Figure 10 shows the ratio of lymphocytes expressing the activation phenotypes CD4 + CD25 ++ from experiment 2.
  • Figure 11 shows the ratio of lymphocytes expressing the activation phenotypes CD8 + HLA-DR + from experiment 2.
  • Figure 12 shows the ratio of lymphocytes expressing the activation phenotypes CD8+CD25+ from experiment 2.
  • Figure 13 shows the ratio of lymphocytes expressing the activation phenotypes CD4CD45RO from experiment 2.
  • compositions comprised in the kit according to the invention and/or the compositions according to the invention may be independently selected from food compositions, dietary supplements, functional foods, medical foods and nutritional products.
  • the above mentioned food compositions may e.g. be beverages, yoghurts, juices, ice creams, breads, biscuits, cereals, health bars, spreads, or nutritional products .
  • the kit according to the invention comprises at least 10 of the above mentioned compositions. More preferably the kit according to the invention comprises at least 7 of the above mentioned compositions. Most preferably the kit according to the invention comprises at least 5 of the above mentioned compositions .
  • the Lactobacillus used according to the invention may be selected from, but not limited to, the group consisting of Lactobacillus plantarum, Lactobacillus rhamnsosus, Lactobacillus fermentum, Lactobacillus paracasei and Lactobacillus gasseri.
  • the Lactobacillus plantarum used according to the invention may be selected from, but not limited to, the group consisting of Lactobacillus plantarum 299, DSM 6595, Lactobacillus plantarum 299v, DSM 9843, Lactobacillus plantarum HEAL 9, DSM 15312, Lactobacillus plantarum HEAL 19, DSM 15313, and Lactobacillus plantarum HEAL 99, DSM 15316.
  • the Lactobacillus paracasei used according to the invention may be selected from, but not limited to, the group consisting of Lactobacillus paracasei 8700:2, DSM 13434, and Lactobacillus paracasei 02A, DSM13432.
  • the Lactobacillus gasseri used according to the invention is selected from, but not limited to, Lactobacillus gasseri VPG44, DSM 16737.
  • probiotic bacterial strains may naturally be used in the present invention and are comprised of the invention.
  • compositions of the kit according to the invention and/or the composition according to the invention further comprise a carrier material.
  • Said carrier material may independently be selected from the group consisting of oat meal gruel, lactic acid fermented foods, resistant starch, dietary fibres, carbohydrates, proteins, and glycosylated proteins.
  • said at least one strain in the compositions of the kit according to the invention is present in an amount from about IxIO 6 to about IxIO 14 CFU. More preferred is that said at least one strain in the compositions of the kit according to the invention is present in an amount from IxIO 8 to IxIO 12 . Even more preferred is that that said at least one strain in the compositions of the kit according to the invention is present in an amount from IxIO 9 to IxIO 11 .
  • said at least two strains in the composition according to the invention are each present in an amount from about IxIO 5 to about IxIO 14 CFU. More preferred is that said at least two strains in the composition according to the invention are each present in an amount from IxIO 8 to IxIO 12 .
  • said at least two strains in the composition according to the invention are each present in an amount from IxIO 9 to IxIO 11 . It is also possible that the at least two strains are present in a total amount from about IxIO 6 to about IxIO 14 CFU, preferably IxIO 8 to IxIO 12 , and more preferably IxIO 9 to IxIO 11 .
  • the kit according to the invention may be administered in a sequence or in an alternating sequence or in a combination thereof. This means that the compositions comprising the respective strain or strains could be administered alternating, e.g. strain A is administered first, then strain B is administered and then strain C is administered. Thereafter the sequence is repeated. It is also possible that first an alternating strain is administered (i.e.
  • compositions of the kit may also be administered simultaneously .
  • the kit is administered in a way such that each composition of the kit is administered at time intervals of between approximately 12 h and approximately 14 days, preferably approximately 24 h and approximately 7 days, more preferably between 24 h and 48 h.
  • one composition comprising at least one probiotic bacterial strain is administered per day.
  • Treatable virus infections are i.e. those caused by a virus selected from the group consisting of common cold virus, rhinovirus, adenovirus, parainfluenza virus, respiratory syncytial virus, enterovirus and corona virus.
  • kit according to the invention may be very beneficial in the sense of being usable prophylactically, i.e. before the virus infection has developed. It is for instance very convenient for a normal healthy individual to take to composition of the invention prophylactically to stay healthy. Examples Example 1
  • the control group took skim milk powder (1 g) .
  • the study had a duration period of 6 or 9 weeks consisting of two weeks wash out period, 2 or 5 weeks active study period and 2 weeks follow up period (Fig. 4) .
  • Each subject was supplied with a list of products containing probiotic products, which should not be consumed during the whole study period.
  • Peripheral blood samples were withdrawn from subjects by venipuncture at two or three time points, day 0, day 14 and day 35.
  • a diary in which each subject stated adverse effects, health conditions and confirmed intake of study product, was kept during the trial.
  • Phenotypic analysis of lymphocytes in whole blood was performed by flow cytometry.
  • the following anti-human monoclonal antibodies were used as surface markers for different cell populations: CD3 FITC (SK7), CD4 APC (SK3), CD8 PerCP (SKl), CD19 PerCP (SJ25C1) , CD56 PE (MY31), CDl ⁇ PE (B73.1), and CD5 FITC (L17F12).
  • CD3 FITC SK7
  • CD4 APC SK3
  • CD8 PerCP SKl
  • CD19 PerCP SJ25C1
  • CD56 PE MY31
  • CDl ⁇ PE B73.1
  • CD5 FITC L17F12
  • HB7 human HB7
  • CD27 PE L1248
  • CDlIb PE D12
  • All antibodies were purchased from Becton-Dickinson (Erembodegum, Belgium) .
  • Whole blood 100 ⁇ l was incubated with antibodies (10 ⁇ l/antibody) for 30 min at 4° C in the dark. Thereafter, 2 ml of FACS lysing solution (Becton- Dickinson) was added and incubated for 15 min at 20° C in the dark. Cells were washed by adding 3 ml FACSFlow and centrifuged at 300 x g for 5 min.
  • Washed cells were resuspended in 200 ⁇ l FACSFlow and analysed on a FacsCalibur (Becton-Dickinson) with CellQuest software. Phagocytosis assay The phagocytic activity of granulocytes and monocytes were quantified with PHAGOTEST ® (Orpegen Pharma, Heidelberg, Germany) according to manufacturers instruction with some modifications. Briefly, 20 xlO 6 FITC labelled E. coli or FITC labelled S. aureus was added to pre-cooled whole blood (100 ⁇ l) . Blood cells and bacteria were incubated on 37° C for 10 FacsCalibur with CellQuest software. Calculations
  • NKT cells NKT cells
  • Relative increase/decrease compared to day 0 could not be detected regarding other cell populations, such as CD4+ T cells, CD8+ T cells, B cells, B-I cells (CD19+CD5+) , NK cells, granulocytes and monocytes. Phagocytic activity
  • the primary task of the immune system is to react rapidly and violently to micro-organisms thereby preventing and curing infections.
  • the killing of microorganisms employs powerful mechanisms that also cause harm to our own tissues. Therefore, it is necessary that it neither reacts to our own tissues, nor to innocuous substances present in the environment. Therefore, the immune system develops and maintains tolerance both to the components of our own body, and to food and inhaled proteins. If this fails, a number of diseases may arise.
  • Means to develop specific immune tolerance are an essential task of the immune system.
  • T helper cell A central role in all immune reactions is played by the T helper cell.
  • a T helper cell becomes activated by its specific antigen, it becomes activated, divides, matures and produces a range of cytokines which direct the action of other types of cells in the immune system, such as cytotoxic T cells and B cells.
  • cytokines which direct the action of other types of cells in the immune system, such as cytotoxic T cells and B cells.
  • Activation of T helper cells is necessary in order to produce most types of immune reactions, including production of antibodies. Conversely, if activation of T helper cells is prevented, most types of immune reactions are paralysed.
  • T helper cells There are several mechanisms by which activation of T helper cells and maintenance of tolerance is ensured.
  • One mechanism is elimination in the thymus of T cells with capacity to recognize and react to own tissue.
  • this elimination is not complete and, furthermore, we also need to develop specific immune tolerance to exogenous antigens. Otherwise we would react violently to all types of inhaled and ingested substances, leading to massive inflammation and wasted immune resources.
  • a cell type that is central for maintenance of tolerance is the regulatory T cell.
  • This cell type can be recognized by certain markers, such as surface expression of CD4 and CD25, possession of intracellular CTLA-4, and transcription of the nuclear protein Foxp3.
  • the regulatory T cells are capable of preventing other T cells to become activated when encountering harmless substances and, hence, prevent all types of unwanted immune reactions .
  • the symbol "+” in connection with a certain marker such as CD4+ and CD25+ means that the marker is expressed on a T cell.
  • CD4+CD25+ T cells are T cells that expresss both the CD4 marker and CD25 marker on its surface. However, nothing is said about the amount of the marker that is expressed, only that it is present.
  • the symbol "++" in connection with a marker such as CD4++ or CD25++ means that there is a lot of marker expressed.
  • the regulatory T cells are those cells with a lot of CD25 on the surface, i.e. CD4+CD25++ cells.
  • CD4+CD25+ T cells are only activated T cells.
  • CD4CD25 is the same as CD4+CD25+.
  • regulatory T cells it is always written as CD4+CD25++ cells.
  • helper T cells after intake of L. plantarum.
  • Expression of activation markers indicates that the T cells have started to proliferate in response to antigen- specific or non-specific stimuli and that these cells more readily exert their effector functions compared to resting T cells.
  • the mechanisms behind L. plantarum induced activation of T cells could be via antigen presenting cells that are activated by toll-like receptors binding to microbial compounds. Activation of antigen presenting cells makes them more efficient in presenting antigen to T cells.
  • helper and cytotoxic T cells have shown to have various expressions of toll-like receptors, which probably make these cells sensible for non-specific activation by microbial components and products.
  • CD45RO+ T cells can secrete a broad spectrum of cytokines.
  • CD45RO+ T cells can proliferate and produce IL-2 when the CD3-TCR complex is stimulated under suboptimal conditions, whereas na ⁇ ve T cells require a strong CD3-TCR stimulus to carry out these functions.
  • the formation of memory T cells is important for induction of an efficient immune response after infection and vaccination.
  • NKT natural killer T
  • NKT cells constitute a lymphocyte subpopulation that coexpress the NK cell marker CD56 and the T cell marker CD3-T cell receptor complex.
  • Studies in both humans and mice have demonstrated that NKT cells play a central role in the regulation of autoimmune diseases, such as multiple sclerosis, type I diabetes, and systemic lupus. NKT cells also exert effector functions against tumour and virus infected cells. Thus, NKT cells are pleotropic in their functions.
  • Other clinical studies evaluating the immunological effects of probiotic bacteria have shown that intake of L.
  • the goal of this example was to investigate the effect on the immune system by giving the same species of lactobacilli for a longer period of time compared to several lactobacilli (different species) administered in a sequence one after the other.
  • gram-positive bacteria the probiotic bacteria Lactobacillus plantarum 299v is used alone or in combination with L. rhamnosus, L. fermentum, L. paracasei, and L. gasseri.
  • gram-negative bacteria Psedomonas lundensis is given. The following groups are studied:
  • helper T cells CD4+
  • CD25 helper T cells
  • results On day 14, there was a borderline significance of CD4+CD25++ T cells being expanded in individuals consuming the sequence of five different lactobacilli strains .
  • T helper cells (CD4+) expressing high density of the CD25 molecule (CD4+CD25++) have been shown to be important in order to protect against autoimmune diseases, allergies and inflammatory bowel diseases. The finding that these cells are expanded after intake of a sequence of different lactobacilli indicate that intake of these bacteria might be beneficial for the individual concerning the risk of developing the above mentioned diseases .

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