CN114350547A - Bifidobacterium lactis strain B-622 and application thereof in preparation of medicines for treating diabetes - Google Patents
Bifidobacterium lactis strain B-622 and application thereof in preparation of medicines for treating diabetes Download PDFInfo
- Publication number
- CN114350547A CN114350547A CN202111550440.0A CN202111550440A CN114350547A CN 114350547 A CN114350547 A CN 114350547A CN 202111550440 A CN202111550440 A CN 202111550440A CN 114350547 A CN114350547 A CN 114350547A
- Authority
- CN
- China
- Prior art keywords
- strain
- diabetes
- bifidobacterium lactis
- preparation
- type
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
- 241000901050 Bifidobacterium animalis subsp. lactis Species 0.000 title claims abstract description 45
- 229940009289 bifidobacterium lactis Drugs 0.000 title claims abstract description 43
- 206010012601 diabetes mellitus Diseases 0.000 title claims abstract description 23
- 239000003814 drug Substances 0.000 title claims abstract description 17
- 238000002360 preparation method Methods 0.000 title claims description 17
- 229940079593 drug Drugs 0.000 title description 7
- 210000004369 blood Anatomy 0.000 claims abstract description 29
- 239000008280 blood Substances 0.000 claims abstract description 29
- 208000001072 type 2 diabetes mellitus Diseases 0.000 claims abstract description 28
- 210000000227 basophil cell of anterior lobe of hypophysis Anatomy 0.000 claims abstract description 20
- 230000035755 proliferation Effects 0.000 claims abstract description 9
- 238000004321 preservation Methods 0.000 claims abstract description 3
- 239000006041 probiotic Substances 0.000 claims description 16
- 235000018291 probiotics Nutrition 0.000 claims description 16
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims description 15
- 239000008103 glucose Substances 0.000 claims description 15
- 230000000529 probiotic effect Effects 0.000 claims description 12
- 230000006907 apoptotic process Effects 0.000 claims description 9
- 230000002218 hypoglycaemic effect Effects 0.000 claims description 9
- 239000000203 mixture Substances 0.000 claims description 6
- 239000000843 powder Substances 0.000 claims description 4
- 239000004480 active ingredient Substances 0.000 claims description 2
- 239000002775 capsule Substances 0.000 claims description 2
- 239000003826 tablet Substances 0.000 claims description 2
- 238000009472 formulation Methods 0.000 claims 4
- 230000000069 prophylactic effect Effects 0.000 claims 1
- 230000001603 reducing effect Effects 0.000 abstract description 17
- 230000000694 effects Effects 0.000 abstract description 14
- 230000002401 inhibitory effect Effects 0.000 abstract description 9
- 108010067722 Dipeptidyl Peptidase 4 Proteins 0.000 abstract description 8
- 230000028327 secretion Effects 0.000 abstract description 8
- 102100025012 Dipeptidyl peptidase 4 Human genes 0.000 abstract description 7
- 230000001737 promoting effect Effects 0.000 abstract description 7
- 230000003914 insulin secretion Effects 0.000 abstract description 6
- 230000001684 chronic effect Effects 0.000 abstract description 5
- GCYXWQUSHADNBF-AAEALURTSA-N preproglucagon 78-108 Chemical compound C([C@@H](C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCCCN)C(=O)NCC(=O)N[C@@H](CCCNC(N)=N)C(=O)NCC(O)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CCCCN)NC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)[C@H](CCC(N)=O)NC(=O)CNC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)[C@H](CO)NC(=O)[C@H](CO)NC(=O)[C@@H](NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CO)NC(=O)[C@@H](NC(=O)[C@H](CC=1C=CC=CC=1)NC(=O)[C@@H](NC(=O)CNC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](C)NC(=O)[C@@H](N)CC=1N=CNC=1)[C@@H](C)O)[C@@H](C)O)C(C)C)C1=CC=CC=C1 GCYXWQUSHADNBF-AAEALURTSA-N 0.000 abstract description 4
- 206010061218 Inflammation Diseases 0.000 abstract description 3
- 230000004054 inflammatory process Effects 0.000 abstract description 3
- 230000001105 regulatory effect Effects 0.000 abstract description 3
- 238000012216 screening Methods 0.000 abstract description 3
- 230000004060 metabolic process Effects 0.000 abstract description 2
- 241000699670 Mus sp. Species 0.000 description 21
- 230000001580 bacterial effect Effects 0.000 description 20
- NOESYZHRGYRDHS-UHFFFAOYSA-N insulin Chemical compound N1C(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(NC(=O)CN)C(C)CC)CSSCC(C(NC(CO)C(=O)NC(CC(C)C)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CCC(N)=O)C(=O)NC(CC(C)C)C(=O)NC(CCC(O)=O)C(=O)NC(CC(N)=O)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CSSCC(NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2C=CC(O)=CC=2)NC(=O)C(CC(C)C)NC(=O)C(C)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2NC=NC=2)NC(=O)C(CO)NC(=O)CNC2=O)C(=O)NCC(=O)NC(CCC(O)=O)C(=O)NC(CCCNC(N)=N)C(=O)NCC(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC(O)=CC=3)C(=O)NC(C(C)O)C(=O)N3C(CCC3)C(=O)NC(CCCCN)C(=O)NC(C)C(O)=O)C(=O)NC(CC(N)=O)C(O)=O)=O)NC(=O)C(C(C)CC)NC(=O)C(CO)NC(=O)C(C(C)O)NC(=O)C1CSSCC2NC(=O)C(CC(C)C)NC(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(N)CC=1C=CC=CC=1)C(C)C)CC1=CN=CN1 NOESYZHRGYRDHS-UHFFFAOYSA-N 0.000 description 14
- 239000007788 liquid Substances 0.000 description 11
- 238000012360 testing method Methods 0.000 description 11
- 239000001963 growth medium Substances 0.000 description 10
- 101800000224 Glucagon-like peptide 1 Proteins 0.000 description 9
- DTHNMHAUYICORS-KTKZVXAJSA-N Glucagon-like peptide 1 Chemical compound C([C@@H](C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCCCN)C(=O)NCC(=O)N[C@@H](CCCNC(N)=N)C(N)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CCCCN)NC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)[C@H](CCC(N)=O)NC(=O)CNC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)[C@H](CO)NC(=O)[C@H](CO)NC(=O)[C@@H](NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CO)NC(=O)[C@@H](NC(=O)[C@H](CC=1C=CC=CC=1)NC(=O)[C@@H](NC(=O)CNC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](C)NC(=O)[C@@H](N)CC=1N=CNC=1)[C@@H](C)O)[C@@H](C)O)C(C)C)C1=CC=CC=C1 DTHNMHAUYICORS-KTKZVXAJSA-N 0.000 description 9
- 102100040918 Pro-glucagon Human genes 0.000 description 9
- 238000010172 mouse model Methods 0.000 description 9
- 210000002966 serum Anatomy 0.000 description 9
- 229940125396 insulin Drugs 0.000 description 8
- 239000000725 suspension Substances 0.000 description 8
- 241000894006 Bacteria Species 0.000 description 7
- 102000004877 Insulin Human genes 0.000 description 6
- 108090001061 Insulin Proteins 0.000 description 6
- 241000699666 Mus <mouse, genus> Species 0.000 description 6
- 238000002474 experimental method Methods 0.000 description 6
- 230000000968 intestinal effect Effects 0.000 description 6
- 239000000047 product Substances 0.000 description 6
- ZSJLQEPLLKMAKR-UHFFFAOYSA-N Streptozotocin Natural products O=NN(C)C(=O)NC1C(O)OC(CO)C(O)C1O ZSJLQEPLLKMAKR-UHFFFAOYSA-N 0.000 description 5
- 239000003242 anti bacterial agent Substances 0.000 description 5
- 235000011389 fruit/vegetable juice Nutrition 0.000 description 5
- 210000001035 gastrointestinal tract Anatomy 0.000 description 5
- 239000002504 physiological saline solution Substances 0.000 description 5
- 239000013641 positive control Substances 0.000 description 5
- 239000007787 solid Substances 0.000 description 5
- ZSJLQEPLLKMAKR-GKHCUFPYSA-N streptozocin Chemical compound O=NN(C)C(=O)N[C@H]1[C@@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O ZSJLQEPLLKMAKR-GKHCUFPYSA-N 0.000 description 5
- 229960001052 streptozocin Drugs 0.000 description 5
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 4
- 206010022489 Insulin Resistance Diseases 0.000 description 4
- 241000186660 Lactobacillus Species 0.000 description 4
- 210000000683 abdominal cavity Anatomy 0.000 description 4
- 229940088710 antibiotic agent Drugs 0.000 description 4
- 244000052616 bacterial pathogen Species 0.000 description 4
- 230000037396 body weight Effects 0.000 description 4
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 description 4
- 230000006870 function Effects 0.000 description 4
- 210000004051 gastric juice Anatomy 0.000 description 4
- 201000001421 hyperglycemia Diseases 0.000 description 4
- 230000001965 increasing effect Effects 0.000 description 4
- 238000002955 isolation Methods 0.000 description 4
- 229940039696 lactobacillus Drugs 0.000 description 4
- 238000010186 staining Methods 0.000 description 4
- 229920001817 Agar Polymers 0.000 description 3
- 241000186000 Bifidobacterium Species 0.000 description 3
- 241000283707 Capra Species 0.000 description 3
- 241000282326 Felis catus Species 0.000 description 3
- 102100023915 Insulin Human genes 0.000 description 3
- 102000004889 Interleukin-6 Human genes 0.000 description 3
- 108090001005 Interleukin-6 Proteins 0.000 description 3
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 241001052560 Thallis Species 0.000 description 3
- 239000008272 agar Substances 0.000 description 3
- 230000003115 biocidal effect Effects 0.000 description 3
- 238000006243 chemical reaction Methods 0.000 description 3
- 238000012258 culturing Methods 0.000 description 3
- 238000011156 evaluation Methods 0.000 description 3
- 210000004153 islets of langerhan Anatomy 0.000 description 3
- 150000002632 lipids Chemical class 0.000 description 3
- 238000009630 liquid culture Methods 0.000 description 3
- 238000002156 mixing Methods 0.000 description 3
- 230000000291 postprandial effect Effects 0.000 description 3
- 230000002829 reductive effect Effects 0.000 description 3
- 238000011160 research Methods 0.000 description 3
- 230000001954 sterilising effect Effects 0.000 description 3
- 238000004659 sterilization and disinfection Methods 0.000 description 3
- 230000004083 survival effect Effects 0.000 description 3
- 108020004465 16S ribosomal RNA Proteins 0.000 description 2
- 241000700199 Cavia porcellus Species 0.000 description 2
- 108050006400 Cyclin Proteins 0.000 description 2
- 241000588724 Escherichia coli Species 0.000 description 2
- WSFSSNUMVMOOMR-UHFFFAOYSA-N Formaldehyde Chemical compound O=C WSFSSNUMVMOOMR-UHFFFAOYSA-N 0.000 description 2
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 2
- 102000009339 Proliferating Cell Nuclear Antigen Human genes 0.000 description 2
- 241000293871 Salmonella enterica subsp. enterica serovar Typhi Species 0.000 description 2
- 241000191967 Staphylococcus aureus Species 0.000 description 2
- 230000003213 activating effect Effects 0.000 description 2
- 230000004913 activation Effects 0.000 description 2
- 210000000481 breast Anatomy 0.000 description 2
- 229940041514 candida albicans extract Drugs 0.000 description 2
- 210000004027 cell Anatomy 0.000 description 2
- 235000012000 cholesterol Nutrition 0.000 description 2
- 230000007547 defect Effects 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 230000002708 enhancing effect Effects 0.000 description 2
- 238000001914 filtration Methods 0.000 description 2
- 235000013305 food Nutrition 0.000 description 2
- 230000002496 gastric effect Effects 0.000 description 2
- 239000011521 glass Substances 0.000 description 2
- 238000001727 in vivo Methods 0.000 description 2
- 239000000859 incretin Substances 0.000 description 2
- 230000002473 insulinotropic effect Effects 0.000 description 2
- 238000002372 labelling Methods 0.000 description 2
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 2
- 238000005259 measurement Methods 0.000 description 2
- 239000002609 medium Substances 0.000 description 2
- 244000005700 microbiome Species 0.000 description 2
- 239000012982 microporous membrane Substances 0.000 description 2
- 229910052757 nitrogen Inorganic materials 0.000 description 2
- 230000003647 oxidation Effects 0.000 description 2
- 238000007254 oxidation reaction Methods 0.000 description 2
- 230000002062 proliferating effect Effects 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 230000035945 sensitivity Effects 0.000 description 2
- 238000012163 sequencing technique Methods 0.000 description 2
- 239000011780 sodium chloride Substances 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- 230000004936 stimulating effect Effects 0.000 description 2
- 208000024891 symptom Diseases 0.000 description 2
- MPLHNVLQVRSVEE-UHFFFAOYSA-N texas red Chemical compound [O-]S(=O)(=O)C1=CC(S(Cl)(=O)=O)=CC=C1C(C1=CC=2CCCN3CCCC(C=23)=C1O1)=C2C1=C(CCC1)C3=[N+]1CCCC3=C2 MPLHNVLQVRSVEE-UHFFFAOYSA-N 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 239000012138 yeast extract Substances 0.000 description 2
- MZOFCQQQCNRIBI-VMXHOPILSA-N (3s)-4-[[(2s)-1-[[(2s)-1-[[(1s)-1-carboxy-2-hydroxyethyl]amino]-4-methyl-1-oxopentan-2-yl]amino]-5-(diaminomethylideneamino)-1-oxopentan-2-yl]amino]-3-[[2-[[(2s)-2,6-diaminohexanoyl]amino]acetyl]amino]-4-oxobutanoic acid Chemical compound OC[C@@H](C(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CCCN=C(N)N)NC(=O)[C@H](CC(O)=O)NC(=O)CNC(=O)[C@@H](N)CCCCN MZOFCQQQCNRIBI-VMXHOPILSA-N 0.000 description 1
- GOZMBJCYMQQACI-UHFFFAOYSA-N 6,7-dimethyl-3-[[methyl-[2-[methyl-[[1-[3-(trifluoromethyl)phenyl]indol-3-yl]methyl]amino]ethyl]amino]methyl]chromen-4-one;dihydrochloride Chemical compound Cl.Cl.C=1OC2=CC(C)=C(C)C=C2C(=O)C=1CN(C)CCN(C)CC(C1=CC=CC=C11)=CN1C1=CC=CC(C(F)(F)F)=C1 GOZMBJCYMQQACI-UHFFFAOYSA-N 0.000 description 1
- 206010067484 Adverse reaction Diseases 0.000 description 1
- 208000037157 Azotemia Diseases 0.000 description 1
- 201000004569 Blindness Diseases 0.000 description 1
- 208000014644 Brain disease Diseases 0.000 description 1
- 229920000742 Cotton Polymers 0.000 description 1
- 102000004127 Cytokines Human genes 0.000 description 1
- 108090000695 Cytokines Proteins 0.000 description 1
- 238000007400 DNA extraction Methods 0.000 description 1
- 208000032274 Encephalopathy Diseases 0.000 description 1
- 208000003790 Foot Ulcer Diseases 0.000 description 1
- 208000014770 Foot disease Diseases 0.000 description 1
- 206010017711 Gangrene Diseases 0.000 description 1
- 102000051325 Glucagon Human genes 0.000 description 1
- 108060003199 Glucagon Proteins 0.000 description 1
- 102000007446 Glucagon-Like Peptide-1 Receptor Human genes 0.000 description 1
- 108010086246 Glucagon-Like Peptide-1 Receptor Proteins 0.000 description 1
- 229920002527 Glycogen Polymers 0.000 description 1
- 108010023302 HDL Cholesterol Proteins 0.000 description 1
- 101000976075 Homo sapiens Insulin Proteins 0.000 description 1
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 1
- 206010020751 Hypersensitivity Diseases 0.000 description 1
- 208000013016 Hypoglycemia Diseases 0.000 description 1
- 108010028554 LDL Cholesterol Proteins 0.000 description 1
- 206010024774 Localised infection Diseases 0.000 description 1
- 206010028813 Nausea Diseases 0.000 description 1
- 102000057297 Pepsin A Human genes 0.000 description 1
- 108090000284 Pepsin A Proteins 0.000 description 1
- 239000001888 Peptone Substances 0.000 description 1
- 108010080698 Peptones Proteins 0.000 description 1
- 208000001647 Renal Insufficiency Diseases 0.000 description 1
- 208000017442 Retinal disease Diseases 0.000 description 1
- 206010038923 Retinopathy Diseases 0.000 description 1
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 1
- 102000004142 Trypsin Human genes 0.000 description 1
- 108090000631 Trypsin Proteins 0.000 description 1
- 108060008682 Tumor Necrosis Factor Proteins 0.000 description 1
- 102000000852 Tumor Necrosis Factor-alpha Human genes 0.000 description 1
- 206010054094 Tumour necrosis Diseases 0.000 description 1
- 206010067584 Type 1 diabetes mellitus Diseases 0.000 description 1
- 206010047700 Vomiting Diseases 0.000 description 1
- 230000002159 abnormal effect Effects 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 230000006838 adverse reaction Effects 0.000 description 1
- 208000026935 allergic disease Diseases 0.000 description 1
- 230000007815 allergy Effects 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 238000010171 animal model Methods 0.000 description 1
- 230000003042 antagnostic effect Effects 0.000 description 1
- 230000000259 anti-tumor effect Effects 0.000 description 1
- 239000003472 antidiabetic agent Substances 0.000 description 1
- 230000001640 apoptogenic effect Effects 0.000 description 1
- 230000036528 appetite Effects 0.000 description 1
- 235000019789 appetite Nutrition 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 230000003385 bacteriostatic effect Effects 0.000 description 1
- 230000004888 barrier function Effects 0.000 description 1
- 235000015278 beef Nutrition 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 230000036772 blood pressure Effects 0.000 description 1
- 238000010241 blood sampling Methods 0.000 description 1
- 210000005252 bulbus oculi Anatomy 0.000 description 1
- VTYYLEPIZMXCLO-UHFFFAOYSA-L calcium carbonate Substances [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 description 1
- 229910000019 calcium carbonate Inorganic materials 0.000 description 1
- KLOIYEQEVSIOOO-UHFFFAOYSA-N carbocromen Chemical compound CC1=C(CCN(CC)CC)C(=O)OC2=CC(OCC(=O)OCC)=CC=C21 KLOIYEQEVSIOOO-UHFFFAOYSA-N 0.000 description 1
- 150000001720 carbohydrates Chemical class 0.000 description 1
- 235000014633 carbohydrates Nutrition 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
- 230000003915 cell function Effects 0.000 description 1
- 208000026106 cerebrovascular disease Diseases 0.000 description 1
- 238000012512 characterization method Methods 0.000 description 1
- 208000037976 chronic inflammation Diseases 0.000 description 1
- 230000006020 chronic inflammation Effects 0.000 description 1
- 239000000701 coagulant Substances 0.000 description 1
- 239000011248 coating agent Substances 0.000 description 1
- 238000000576 coating method Methods 0.000 description 1
- 230000001276 controlling effect Effects 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 238000006731 degradation reaction Methods 0.000 description 1
- 230000018044 dehydration Effects 0.000 description 1
- 238000006297 dehydration reaction Methods 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000029087 digestion Effects 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 229940090124 dipeptidyl peptidase 4 (dpp-4) inhibitors for blood glucose lowering Drugs 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000016097 disease of metabolism Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 239000012154 double-distilled water Substances 0.000 description 1
- 238000012137 double-staining Methods 0.000 description 1
- 230000002124 endocrine Effects 0.000 description 1
- 239000002158 endotoxin Substances 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 239000000284 extract Substances 0.000 description 1
- 208000030533 eye disease Diseases 0.000 description 1
- 201000005577 familial hyperlipidemia Diseases 0.000 description 1
- 230000002550 fecal effect Effects 0.000 description 1
- MHMNJMPURVTYEJ-UHFFFAOYSA-N fluorescein-5-isothiocyanate Chemical compound O1C(=O)C2=CC(N=C=S)=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 MHMNJMPURVTYEJ-UHFFFAOYSA-N 0.000 description 1
- 235000010855 food raising agent Nutrition 0.000 description 1
- 230000030136 gastric emptying Effects 0.000 description 1
- 238000001502 gel electrophoresis Methods 0.000 description 1
- 235000021472 generally recognized as safe Nutrition 0.000 description 1
- MASNOZXLGMXCHN-ZLPAWPGGSA-N glucagon Chemical compound C([C@@H](C(=O)N[C@H](C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H]([C@@H](C)O)C(O)=O)C(C)C)NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](C)NC(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H](CO)NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)[C@H](CCCCN)NC(=O)[C@H](CO)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CO)NC(=O)[C@@H](NC(=O)[C@H](CC=1C=CC=CC=1)NC(=O)[C@@H](NC(=O)CNC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CO)NC(=O)[C@@H](N)CC=1NC=NC=1)[C@@H](C)O)[C@@H](C)O)C1=CC=CC=C1 MASNOZXLGMXCHN-ZLPAWPGGSA-N 0.000 description 1
- 229960004666 glucagon Drugs 0.000 description 1
- 238000007446 glucose tolerance test Methods 0.000 description 1
- 229940096919 glycogen Drugs 0.000 description 1
- 208000019622 heart disease Diseases 0.000 description 1
- 229940088597 hormone Drugs 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 230000004957 immunoregulator effect Effects 0.000 description 1
- 230000002757 inflammatory effect Effects 0.000 description 1
- 230000005764 inhibitory process Effects 0.000 description 1
- 230000006362 insulin response pathway Effects 0.000 description 1
- 229940100601 interleukin-6 Drugs 0.000 description 1
- 210000004347 intestinal mucosa Anatomy 0.000 description 1
- 239000007928 intraperitoneal injection Substances 0.000 description 1
- 208000017169 kidney disease Diseases 0.000 description 1
- 201000006370 kidney failure Diseases 0.000 description 1
- 239000004310 lactic acid Substances 0.000 description 1
- 235000014655 lactic acid Nutrition 0.000 description 1
- 230000037356 lipid metabolism Effects 0.000 description 1
- XFIKMPRBSORKQP-JATHGWPISA-M lithium;9-[(e)-4-[(2s,3r,4r,5s)-3,4-dihydroxy-5-[[(2s,3s)-3-[(2s,3s)-3-hydroxybutan-2-yl]oxiran-2-yl]methyl]oxan-2-yl]-3-methylbut-2-enoyl]oxynonanoate Chemical class [Li+].C[C@H](O)[C@H](C)[C@@H]1O[C@H]1C[C@@H]1[C@@H](O)[C@@H](O)[C@H](C\C(C)=C\C(=O)OCCCCCCCCC([O-])=O)OC1 XFIKMPRBSORKQP-JATHGWPISA-M 0.000 description 1
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 1
- 238000013227 male C57BL/6J mice Methods 0.000 description 1
- SQQMAOCOWKFBNP-UHFFFAOYSA-L manganese(II) sulfate Chemical compound [Mn+2].[O-]S([O-])(=O)=O SQQMAOCOWKFBNP-UHFFFAOYSA-L 0.000 description 1
- 229910000357 manganese(II) sulfate Inorganic materials 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 230000010534 mechanism of action Effects 0.000 description 1
- 239000000155 melt Substances 0.000 description 1
- 208000030159 metabolic disease Diseases 0.000 description 1
- XZWYZXLIPXDOLR-UHFFFAOYSA-N metformin Chemical compound CN(C)C(=N)NC(N)=N XZWYZXLIPXDOLR-UHFFFAOYSA-N 0.000 description 1
- 229960003105 metformin Drugs 0.000 description 1
- 238000000034 method Methods 0.000 description 1
- 229940126619 mouse monoclonal antibody Drugs 0.000 description 1
- 230000008693 nausea Effects 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 230000008621 organismal health Effects 0.000 description 1
- 230000004792 oxidative damage Effects 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 210000000496 pancreas Anatomy 0.000 description 1
- 230000001717 pathogenic effect Effects 0.000 description 1
- 229940111202 pepsin Drugs 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
- 230000002093 peripheral effect Effects 0.000 description 1
- 230000035790 physiological processes and functions Effects 0.000 description 1
- 229920000136 polysorbate Polymers 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 108090000765 processed proteins & peptides Proteins 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 208000037821 progressive disease Diseases 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 210000002955 secretory cell Anatomy 0.000 description 1
- 238000004088 simulation Methods 0.000 description 1
- 208000017520 skin disease Diseases 0.000 description 1
- 239000001632 sodium acetate Substances 0.000 description 1
- 235000017281 sodium acetate Nutrition 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- UFTFJSFQGQCHQW-UHFFFAOYSA-N triformin Chemical compound O=COCC(OC=O)COC=O UFTFJSFQGQCHQW-UHFFFAOYSA-N 0.000 description 1
- 239000012588 trypsin Substances 0.000 description 1
- 239000012137 tryptone Substances 0.000 description 1
- 238000011144 upstream manufacturing Methods 0.000 description 1
- 208000009852 uremia Diseases 0.000 description 1
- 210000003462 vein Anatomy 0.000 description 1
- 238000012795 verification Methods 0.000 description 1
- 230000008673 vomiting Effects 0.000 description 1
Images
Classifications
-
- 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
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Landscapes
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses a bifidobacterium lactis strain B-622 and application thereof in preparing a medicament for treating diabetes. The preservation number of the strain B-622 is CCTCC NO.M: 20211355, deposited at the China center for type culture Collection on 11/2/2021. The strain obtained by screening plays a role in reducing blood sugar by promoting the secretion of glucagon-like peptide-1 (GLP-1), repairing the insulin secretion function, regulating blood fat metabolism, reducing chronic low-grade inflammation and inhibiting the activity of dipeptidyl peptidase-4, and can also inhibit a type 2 diabetes model mouse and stimulate the proliferation of islet beta cells of the type 2 diabetes model mouse.
Description
Technical Field
The invention belongs to the technical field of microorganisms, and particularly relates to a bifidobacterium lactis strain B-622 and application thereof in preparation of a medicament for treating diabetes.
Background
Diabetes (diabetes) is an endocrine metabolic disease associated with abnormal production and action of insulin, characterized primarily by hyperglycemia, and primarily includes type I and type II diabetes. The diabetes mellitus people in China mainly take II type diabetes mellitus, and account for more than 90 percent of the total number of diabetes mellitus patients. Type II diabetes, which is dominated by relative insulin deficiency and insulin resistance, eventually leads to hyperglycemia, and chronic hyperglycemia symptoms can cause a series of chronic complications, such as foot disease (foot ulcers, infection and gangrene), nephropathy (renal failure, uremia), eye disease (retinopathy, blurring, blindness), encephalopathy (cerebrovascular disease), heart disease, skin disease, venereal disease, and the like. Type II diabetes is a slowly progressive disease whose central link in onset is insulin resistance and a defect in islet beta cell function. At present, the number of people with type II diabetes mellitus in China is the first in the world, so that the development of prevention and treatment measures for type II diabetes mellitus is not slow. Many studies found that probiotics have a good biological effect on improving type II diabetes, but the mechanism of action is not clearly elucidated, and many strains of functional probiotics, leavening agents, and core technologies are monopolized abroad.
Incretins are a hormone secreted by small intestinal secretory cells that helps the body to produce a postprandial insulin response after eating carbohydrates, and their insulinotropic effect accounts for about 60% of the total amount of postprandial insulin secretion. To date, 2 incretins, glucose-dependent insulin release peptide (GIP) and Glucagon-like peptide-1 (GLP-1), have been isolated. Clinical researches show that GLP-1 and analogues thereof have important application values in the treatment process of type 2 diabetes, and GLP-1 has physiological functions of promoting insulin secretion, proliferating and inhibiting islet beta cells, inhibiting apoptosis, slowing down gastric emptying, inhibiting secretion of postprandial glucagon, reducing synthesis of glycogen, improving insulin sensitivity, controlling appetite and the like. Thus, GLP-1 is receiving increasing attention as a safe and effective insulinotropic secretion. However, GLP-1 is degraded by Dipeptidyl peptidase-4 (DPP-4) in vivo very rapidly, the half-sadness period is only 1-2min, and when the endogenous GLP-1 secretion amount is insufficient, the dose for activating the GLP-1 receptor cannot be accumulated, so that the application in clinic is difficult. The DPP-4 inhibitor inhibits the degradation of GLP-1 by DPP-4, so that endogenous GLP-1 is accumulated in vivo to achieve the effect of reducing blood sugar. The two medicines have the functions of protecting islet cells, reducing weight and the like, rarely cause hypoglycemia response, and are increasingly widely applied clinically. The most common adverse reactions of the two medicines are nausea, vomiting and other gastrointestinal reactions.
Probiotics (probiotic) is a micro-ecological preparation which has a promoting effect on the health of organisms, mainly comprises lactobacillus and bifidobacterium, and is a microorganism which is generally recognized as safe to eat. The probiotic has the effects of improving intestinal tract, enhancing immunity, resisting allergy, resisting oxidation, reducing cholesterol, lowering blood pressure, reducing weight, etc. In recent years, some lactic acid bacteria have been found to have hypoglycemic effects. The mechanism for reducing blood sugar mainly comprises: improving the barrier function of the intestinal mucosa; enhancing immunoregulatory activity; repairing the oxidative damage of the organism and improving the oxidation resistance; promoting insulin secretion and improving insulin sensitivity; inhibiting or delaying the absorption of glucose by intestinal tract, and promoting the utilization of glucose by peripheral tissues and target organs.
Disclosure of Invention
Aiming at the defects in the prior art, the invention provides a bifidobacterium lactis strain B-622 and application thereof in preparing a medicament for treating diabetes, wherein the strain plays a role in reducing blood sugar by promoting secretion of glucagon-like peptide-1 (GLP-1), repairing insulin secretion function, regulating blood lipid metabolism, reducing chronic low-grade inflammation and inhibiting activity of dipeptidyl peptidase-4, and can also inhibit a type 2 diabetes model mouse and stimulate proliferation of islet beta cells of the type 2 diabetes model mouse.
In order to achieve the purpose, the technical scheme adopted by the invention for solving the technical problems is as follows:
a Bifidobacterium lactis (Bifidobacterium animalis subsp.Lactis) strain B-622, wherein the preservation number of the strain B-622 is CCTCC NO.M: 20211355, deposited at the China center for type culture Collection on 11/2/2021, address: wuhan university in Wuhan, China.
Further, the strain B-622, disrupted product or culture of the strain B-622 according to claim 1 is contained as an active ingredient.
Further, the probiotic preparation for reducing blood sugar is powder, tablets or capsules.
The hypoglycemic probiotic preparation is applied to the preparation of medicaments for preventing or assisting in reducing blood sugar.
The application of the strain B-622 in preparing hypoglycemic drugs.
The application of the strain B-622 in preparing a medicament for treating diabetes.
The application of the probiotic preparation for reducing blood sugar in the preparation of medicaments for treating diabetes is provided.
Further, the strain B-622 or the hypoglycemic probiotic preparation can inhibit apoptosis of islet beta cells and promote proliferation of the islet beta cells.
Further, diabetes is type ii diabetes.
The invention has the beneficial effects that:
the invention takes lactobacillus which is separated and identified from a 2-year-old healthy excrement sample which is breast-fed in the west-Tibet region and is not contacted with antibiotics as a research object, and obtains a new lactobacillus strain named as bifidobacterium lactis B-622 through a large number of experimental screens. The invention adopts high-fat feed and streptozotocin to establish a diabetes model, and the bifidobacterium lactis B-622 is continuously administered to a mouse by intragastric administration for 28 days to discuss the treatment effect of the bifidobacterium lactis B-622 on the diabetes model mouse. Research results show that the bifidobacterium lactis B-622 plays a role in reducing blood sugar by promoting the secretion of glucagon-like peptide-1 (GLP-1), repairing the secretion function of insulin, regulating blood fat metabolism, reducing chronic low-grade inflammation and inhibiting the activity of dipeptidyl peptidase-4.
The bifidobacterium lactis B-622 strain has the effects of preventing and treating diabetes, so that the bifidobacterium lactis B-622 strain can be used for preparing foods, health-care products or medicines and the like with the function of reducing blood sugar, and meanwhile, the bifidobacterium lactis B-622 strain provided by the invention can also be used as functional probiotics to be applied to preparing foods, health-care products or medicines, so that the bifidobacterium lactis B-622 strain is safe, free of toxic and side effects and wide in application prospect. The screening of the lactobacillus with the blood sugar reducing effect is developed, so that the audience population is wide, and the market space is huge.
Drawings
FIG. 1 shows the effect of Bifidobacterium lactis B-622 on blood glucose in type 2 diabetic mice;
FIG. 2 shows that Bifidobacterium lactis B-622 inhibits apoptosis of islet beta cells in type 2 diabetes model mice;
FIG. 3 shows that Bifidobacterium lactis B-622 stimulates the proliferation of islet beta cells in type 2 diabetes model mice.
Detailed Description
The following description of the embodiments of the present invention is provided to facilitate the understanding of the present invention by those skilled in the art, but it should be understood that the present invention is not limited to the scope of the embodiments, and it will be apparent to those skilled in the art that various changes may be made without departing from the spirit and scope of the invention as defined and defined in the appended claims, and all matters produced by the invention using the inventive concept are protected.
Example 1 isolation screening and characterization of Bifidobacterium lactis B-622
1. Collection of samples
A 2 year old healthy stool sample from the tibetan region that was breast-fed and not exposed to antibiotics was collected.
2. Strain isolation
Placing 1g of sample into 9mL of physiological saline, shaking and uniformly mixing, carrying out gradient dilution on the bacterial suspension, sucking 100 mu L of appropriate gradient bacterial liquid on a bifidobacterium isolation medium agar plate added with mupirocin lithium salt, uniformly coating, replacing air with anaerobic air, and then standing at 37 ℃ for constant-temperature culture for 48 hours. And (3) selecting a single bacterial colony which is opaque, milky white, round, glossy, neat in edge, convex in surface and wet when the bacterial colony grows out, performing repeated streak purification culture on an MRS solid culture medium to obtain 1 bacterial strain of the infant fecal bacteria, and storing the bacterial strain at the temperature of 4 ℃ for later use. Bifidobacterium isolation medium (g/L): 10 parts of beef extract, 10 parts of peptone, 5 parts of yeast extract, 5 parts of glucose, 5 parts of sodium acetate, 2 parts of diammonium hydrogen citrate, 801 parts of Tween and K parts of2HPO4 2、MgSO4·7H2 O 0.58、 MnSO4·H2O 0.25、CaCO3Sterilizing at 20 and 121 deg.C for 20min, adding 1.5% agar powder into MRS solid culture medium, the agar has stable chemical structure, and only acts as coagulant in the culture medium, and melts at 100 deg.CThe mixture is formed into liquid and cooled to below 45 ℃ to be solidified again. The bacterial colony of the strain is opaque and milky white, round and glossy, neat in edge, convex and moist in surface. The strain is sensitive to oxygen and is strictly anaerobic, and the air is replaced by anaerobic air during culture. The optimal growth temperature of the strain is 37 ℃, and the optimal initial pH value is 6.8.
3. Molecular biological identification of bifidobacterium lactis B-622
And (3) purifying and culturing the primary screened thalli, collecting the primary screened thalli, extracting the strain genome DNA by adopting a genome DNA extraction kit (TIANGEN company), and performing the steps of: AGTTTGATCMTGGCTCAG (SEQ ID NO.1) and a downstream primer 1492R: GGTTACCTTGTTACGACTT (SEQ ID NO.2), and the 16S rDNA sequence is amplified to obtain a PCR product. And sequencing the PCR product. Wherein the PCR reaction system comprises: 2 mu L of each of the upstream and downstream primers, 2 mu L of the DNA template, 2 mu L of ddH2O 21 mu L, of Super PfX MasterMix 25 mu L (CW2965M, kang century, China). And (3) PCR reaction conditions: 10min at 95 ℃; 30s at 94 ℃, 30s at 55 ℃, 2min at 72 ℃ and 35 cycles; 10min at 72 ℃. The PCR product was detected by gel electrophoresis and sent to Beijing optimak company for sequencing. The determined gene sequences were submitted to NCBI database (www.ncbi.nlm.nih.gov) for BLAST analysis and alignment, using the 16S rDNA gene sequence homology of 99% or more as the identification standard.
Example 2 evaluation of tolerance of Bifidobacterium lactis B-622 to simulated artificial gastric and intestinal juices
Inoculating 100 μ L of test bacterial liquid into nitrogen-filled anaerobic glass tube filled with 7mL of MRS liquid culture medium, standing at 37 deg.C for culture and activation for 2 times. And centrifuging 200 mu L of activated bifidobacterium lactis B-622 bacterial liquid, collecting thalli, respectively adding 1mL of simulated artificial gastric juice with the pH value of 2.5 and simulated artificial intestinal juice with the pH value of 8.0, fully mixing, digesting for 3.0h at 37 ℃, sampling, detecting the number of viable bacteria, and calculating the survival rate, wherein the results are shown in table 1. Wherein the artificial gastric juice and the artificial intestinal juice are prepared immediately and stored at 4 ℃ for standby in a short time. The survival rate is the viable count of the activated bacteria liquid/the viable count after digestion is multiplied by 100 percent.
The formula of the artificial gastric juice comprises: preparing 0.85% physiological saline, adjusting pH to 2.5 with dilute hydrochloric acid, adding 0.3% pepsin, dissolving completely, and filtering with 0.22 μm microporous membrane for sterilization.
The formula of the artificial intestinal juice comprises: preparing 0.85% physiological saline, adjusting pH to 8.0 with sodium hydroxide, adding 0.1% trypsin, dissolving completely, and filtering with 0.22 μm microporous membrane for sterilization.
TABLE 1 Experimental data table for Bifidobacterium lactis B-622 simulation digestive tract environment
The results in Table 1 show that the survival rate of the Bifidobacterium lactis B-622 in both simulated artificial gastric juice and artificial intestinal juice is high, which indicates that the Bifidobacterium lactis B-622 can effectively resist the influence of the gastrointestinal tract and still has high activity after passing through the gastrointestinal tract.
Example 3 evaluation of bacteriostatic Activity of Bifidobacterium lactis B-622
And inoculating 100 mu L of test bacterial liquid into a nitrogen-filled anaerobic glass tube filled with 7mL of MRS liquid culture medium, and standing and culturing at the constant temperature of 37 ℃ for activation. Respectively inoculating 100 mu L of pathogenic indicator strains of escherichia coli, staphylococcus aureus and salmonella typhi into 10mL of liquid LB culture medium, and culturing and activating by a constant temperature shaker at 37 ℃ at 100 rpm. Diluting the activated pathogenic bacteria liquid to bacterial suspension with the concentration of 108CFU/mL, respectively adding 300 mu L of each pathogenic bacteria suspension into 30mL of MRS solid culture medium with the appropriate temperature (about 50 ℃), uniformly mixing (indicating the viable count of the bacteria is 106CFU/mL), pouring into a flat plate with 4 sterile oxford cups placed in the flat plate in advance, taking out the oxford cups by using tweezers after the culture medium is solidified, adding 160 mu L of test bacteria liquid (the viable count is 108CFU/mL) into holes left after the oxford cups are pulled out, and performing 3 parallels and 1 control, wherein the control is replaced by the MRS liquid culture medium. The plate was placed upright after being covered with a light lid, and after replacing the air with oxygen-free air, the plate was cultured in a constant temperature incubator at 37 ℃ for an appropriate time to observe, and the diameter of the zone of inhibition was measured with a vernier caliper, and the results are shown in Table 2. LB culture medium: 10g of tryptone, 5g of yeast extract powder, 10g of sodium chloride and 1000ml of distilled water, wherein the pH value is 7.0-7.2, and the sterilization is carried out at 121 ℃ for 15 min.
TABLE 2 antagonistic activity of Bifidobacterium lactis B-622 against pathogenic bacteria data Table
The results in Table 2 show that Bifidobacterium lactis B-622 has strong inhibitory effect on common enteric pathogenic bacteria Escherichia coli, Staphylococcus aureus and Salmonella typhi.
Example 4 evaluation of antibiotic susceptibility of Bifidobacterium lactis B-622
After test bacteria are streaked and activated on an MRS solid plate, bacterial mud is picked and placed in physiological saline to prepare bacterial suspension, the concentration of the bacterial suspension is adjusted to be 108CFU/mL, 100 mu L of the bacterial suspension is added to the MRS solid plate poured in advance, the bacterial suspension is uniformly coated on the plate by using an aseptic cotton swab, an antibiotic drug sensitive sheet is attached, air is replaced by anaerobic air, then the plate is cultured at the positive position of 37 ℃, the diameter of the sensitivity of the bacterial strain to the antibiotic is measured by using a vernier caliper after 24h, and the result is shown in Table 3.
TABLE 3 data table of Bifidobacterium lactis B-622 sensitivity to antibiotics
The results in table 3 show that bifidobacterium lactis B-622 is not resistant to 17 common antibiotics, demonstrating that bifidobacterium lactis is safe in use.
Example 5 establishment and experiment of animal models
One, packet processing
100 male C57BL/6J mice, randomly divided into 4 groups of 20 mice each, were treated as follows:
1. blank control group: feeding the test by adopting common feed from the beginning to the end of the test; at the 9 th week, the mice are injected with 30mg/kg of physiological saline at one time in the abdominal cavity after fasting for 12 hours; administration was performed from week 10 to week 13 at the following doses: 12mL of sterile PBS buffer was administered per kilogram of body weight.
2. Model group: feeding with high fat feed in 1-8 weeks, and feeding with common feed in 9-9 weeks; at 9 weeks, 30mg/kg of streptozotocin is injected into the abdominal cavity once after the mice fasting for 12 hours; administration is performed from week 10 to week 13; the administration dose is as follows: 12mL of sterile PBS buffer was administered per kilogram of body weight.
3. Positive control group: feeding with high fat feed in 1-8 weeks, and feeding with common feed in 9-9 weeks; at 9 weeks, 30mg/kg of streptozotocin is injected into the abdominal cavity once after the mice fasting for 12 hours; administration is performed from week 10 to week 13; the administration dose is as follows: a0.1 mg/ml solution of metformin in PBS was administered at 12ml/kg body weight once a day.
Group B-622: feeding with high fat feed in 1-8 weeks, and feeding with common feed in 9-9 weeks; at 9 weeks, 30mg/kg of streptozotocin is injected into the abdominal cavity once after the mice fasting for 12 hours; administration is performed from week 10 to week 13; the administration dose is as follows: 12mL of the bacterial suspension was administered per kg of body weight. The preparation method of the bacterial liquid comprises the following steps: suspending Bifidobacterium lactis B-622 with PBS buffer to obtain Bifidobacterium lactis B-622 with concentration of 1.0 × 109CFU/mL of bacterial liquid.
Week 1 to week 8, i.e., days 1 to 56 of the test, week 10 to week 13, i.e., days 64 to 91 of the test, and so on. Intraperitoneal injection of streptozotocin or saline was performed on day 57 of the experiment.
Second, judging and establishing diabetes model standard
At week 9 (day 63 of the experiment), the mice were bled for Fasting Blood Glucose (FBG). The mice meeting the following criteria are type 2 diabetes model mice: FBG is measured for two times continuously and is more than or equal to 11.1 mmol/L. The results show that on the 57 th day of the experiment, the mice of the model group, the positive control group and the B-622 group are type 2 diabetes model mice.
Third, blood sugar detection
The fasting blood glucose level of the mice was measured by tail vein blood sampling at week 10, week 11, week 12, week 13 and week 14, respectively (the average value in the week was calculated by daily measurement). Test day 99 a glucose tolerance test was performed.
The fasting blood glucose results are shown in FIG. 1. Compared with the model group, the fasting blood glucose value of the mice in the B-622 group is in a descending trend. The fasting blood glucose values of the mice in the B-622 group are obviously reduced at 12 weeks, 13 weeks and 14 weeks, and the blood glucose value at the 14 week is reduced to 12.35mmol/L, which are very obviously different from those of the mice in the model group (P is less than 0.01). The result shows that the bifidobacterium lactis B-622 has a treatment effect on hyperglycemia of type 2 diabetes and can obviously reduce the fasting blood glucose value.
Detection of blood lipid, insulin and inflammatory cytokine
At the end of the test (day 100), each group of mice was collected, the eyeballs were removed, blood was collected, the mixture was coagulated at room temperature for 60min and centrifuged at 3000r/min for 15min, and the supernatant (serum) was collected. Detecting the content of total cholesterol, triglyceride, high density lipoprotein cholesterol, low density lipoprotein cholesterol, insulin, endotoxin, anti-tumor necrosis factor-alpha and interleukin-6 in serum. The results of the measurement of serum lipid-related factors are shown in Table 1. Compared with the model group, the serum TC and TG contents of the mice in the B-622 group are reduced. The result shows that the bifidobacterium lactis B-622 can obviously improve the hyperlipemia symptom of type 2 diabetes.
TABLE 1 Effect of Bifidobacterium lactis B-622 on blood lipids in type 2 diabetic mice
Note: significant differences (P < 0.05) and very significant differences (P < 0.01) were compared to the model group.
TABLE 2 Effect of Bifidobacterium lactis B-622 on ET, TNF- α, IL-6, INS, GLP-1, DPP-4 in serum of type 2 diabetes model mice
Note: significant differences (P < 0.05) and very significant differences (P < 0.01) were compared to the model group.
As shown in Table 2, the INS content in the serum of mice in each administration group was increased to different extents during the treatment period, and the INS content in the serum of mice in the positive control group and B-622 group was significantly increased (P < 0.01) and the ET level in the serum of mice in the B-622 group was significantly decreased (P < 0.01) as compared with the model group. B-622 can obviously reduce the serum of type 2 diabetic mice, reduce the contents of TNF-alpha and IL-6 and effectively relieve chronic inflammation of organisms. GLP-1 secretion of the B-622 group is obviously increased, and DPP-IV enzyme activity is obviously inhibited.
Fifth, the effect of Bifidobacterium lactis B-622 on islet beta cells of type 2 diabetes model mice
In the experiment, all mice are killed by removing cervical vertebrae after blood is taken on day 100, pancreas samples are taken and fixed by 4% formalin fixing solution, then wax blocks are made by dehydration and transparency, the thickness of the slices is 5um, and subsequent staining is carried out. Verification that bifidobacterium lactis B-622 inhibits apoptosis of islet beta cells of type 2 diabetes model mice and stimulates proliferation of islet beta cells of type 2 diabetes model mice
1. Bifidobacterium lactis B-622 for inhibiting apoptosis of islet beta cells of type 2 diabetes model mouse
Apoptosis test TUNEL apoptosis kit (cat 11684817910, Roche) was stained to show green fluorescence according to the kit's instructions, the primary antibody was polyclonal Guinea pig anti-insulin (cat A0564, DAKO), the secondary antibody was goat anti-guinea pig Texas Red stained to show Red fluorescence, the green fluorescence was apoptotic cells (white arrow mark), the Red fluorescence was islet beta cells (dotted line delineation region), group B-622 was clearly seen in FIG. 2, and the staining results of the positive control group showed that the expression of green fluorescence was significantly lower than that of the model group. Thus, B-622 can inhibit the apoptosis of islet beta cells.
2. Bifidobacterium lactis B-622 for stimulating proliferation of islet beta cells of type 2 diabetes model mouse
Experiment of stimulating proliferation of beta cells of pancreatic islets of mice model of type 2 diabetes by bifidobacterium lactis B-622 staining with double staining of insulin and PCNA, and labeling of beta cells of pancreatic islets: the primary antibody was polyclonal Guinea pig anti-insulin (cat # A0564, DAKO) and the secondary antibody was goat anti-guinea pig Texas Red stained with Red fluorescence (solid line delineation). Labeling of proliferating cells: the primary antibody is mouse monoclonal antibody [ PC10] to PCNA (cat: ab29, abcam), and the secondary antibody is goat anti-mouse FITC stained in green fluorescence (gray arrow mark). In FIG. 3, it can be seen that the green fluorescence expression of the staining results of the B-622 group and the positive control group is significantly higher than that of the model group, indicating that B-622 can stimulate the proliferation of islet beta cells.
Claims (9)
1. A Bifidobacterium lactis (Bifidobacterium animalis subsp.Lactis) strain B-622 is characterized in that the preservation number of the strain B-622 is CCTCC NO.M: 20211355, deposited at the China center for type culture Collection on 11/2/2021.
2. A hypoglycemic probiotic preparation, comprising the strain B-622, disrupted product or culture of the strain B-622 according to claim 1 as an active ingredient.
3. The hypoglycemic probiotic formulation according to claim 2, wherein the hypoglycemic probiotic formulation is a powder, tablet or capsule.
4. Use of a probiotic formulation for lowering blood glucose according to claim 2 or 3, for the preparation of a medicament for the prophylactic or adjunctive lowering of blood glucose.
5. The use of the strain B-622 according to claim 1 for the preparation of a hypoglycemic medicament.
6. The use of the strain B-622 according to claim 1 for the preparation of a medicament for the treatment of diabetes.
7. Use of a probiotic formulation for lowering blood glucose according to any of claims 2 or 3 for the preparation of a medicament for the treatment of diabetes.
8. The use according to claim 6 or 7, wherein the strain B-622 or the hypoglycemic probiotic preparation inhibits apoptosis of islet beta cells and promotes proliferation of islet beta cells.
9. The use according to claim 6 or 7, wherein the diabetes is type II diabetes.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111550440.0A CN114350547B (en) | 2021-12-17 | 2021-12-17 | Bifidobacterium lactis strain B-622 and application thereof in preparation of medicines for treating diabetes |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111550440.0A CN114350547B (en) | 2021-12-17 | 2021-12-17 | Bifidobacterium lactis strain B-622 and application thereof in preparation of medicines for treating diabetes |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN114350547A true CN114350547A (en) | 2022-04-15 |
| CN114350547B CN114350547B (en) | 2023-05-16 |
Family
ID=81099195
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202111550440.0A Active CN114350547B (en) | 2021-12-17 | 2021-12-17 | Bifidobacterium lactis strain B-622 and application thereof in preparation of medicines for treating diabetes |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN114350547B (en) |
Citations (13)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102448477A (en) * | 2009-03-25 | 2012-05-09 | 科·汉森有限公司 | Use of a probiotic to regulate body weight |
| CN102670661A (en) * | 2011-03-08 | 2012-09-19 | 惠宏襄 | Drug for stimulating glucagon-like peptide 1 (GLP-1) secretion |
| CN104548096A (en) * | 2013-10-09 | 2015-04-29 | 深圳翰宇药业股份有限公司 | Pharmaceutical composition containing GLP-1 analogue and DPP-4 inhibitor and preparation method thereof |
| CN104884610A (en) * | 2012-12-20 | 2015-09-02 | 热尔韦·达诺尼公司 | Use of bifidobacterium animalis for treating or preventing body weight gain and insulin resistance |
| US20160213702A1 (en) * | 2015-01-26 | 2016-07-28 | Kaleido Biosciences, Inc. | Glycan therapeutics and related methods thereof |
| CN106535908A (en) * | 2014-02-14 | 2017-03-22 | 凡赛尔医药股份有限公司 | Use of compositions comprising bifidobacterium animalis ssp. lactis LMG P-28149 |
| CN108085285A (en) * | 2018-01-25 | 2018-05-29 | 吉林省命之元生物科技有限公司 | One DM-50 plants of lactobacillus plantarum and its application |
| US20190240100A1 (en) * | 2018-02-02 | 2019-08-08 | Augusta University Research Institute, Inc. | Compositions and Methods for Treating Diabetes |
| CN110150669A (en) * | 2019-04-29 | 2019-08-23 | 河北一然生物科技有限公司 | A kind of probiotic composition and its application suitable for patients with diabetes mellitus |
| CN110446532A (en) * | 2016-12-12 | 2019-11-12 | Gi动力公司 | Treatment method relevant to stomach and intestine implantation material |
| CN110604749A (en) * | 2019-08-30 | 2019-12-24 | 北京农学院 | Bifidobacterium animalis and its use in controlling diabetes or hyperlipidemia, especially weight gain or obesity |
| CN114377037A (en) * | 2022-02-22 | 2022-04-22 | 内蒙古蒙牛乳业（集团）股份有限公司 | Composition of lactobacillus paracasei LC-37 and application thereof in reducing blood sugar |
| CN114752529A (en) * | 2022-04-29 | 2022-07-15 | 科郦有限公司 | Lactobacillus plantarum HOM3201 strain, viable bacteria preparation thereof, preparation method and application |
-
2021
- 2021-12-17 CN CN202111550440.0A patent/CN114350547B/en active Active
Patent Citations (13)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102448477A (en) * | 2009-03-25 | 2012-05-09 | 科·汉森有限公司 | Use of a probiotic to regulate body weight |
| CN102670661A (en) * | 2011-03-08 | 2012-09-19 | 惠宏襄 | Drug for stimulating glucagon-like peptide 1 (GLP-1) secretion |
| CN104884610A (en) * | 2012-12-20 | 2015-09-02 | 热尔韦·达诺尼公司 | Use of bifidobacterium animalis for treating or preventing body weight gain and insulin resistance |
| CN104548096A (en) * | 2013-10-09 | 2015-04-29 | 深圳翰宇药业股份有限公司 | Pharmaceutical composition containing GLP-1 analogue and DPP-4 inhibitor and preparation method thereof |
| CN106535908A (en) * | 2014-02-14 | 2017-03-22 | 凡赛尔医药股份有限公司 | Use of compositions comprising bifidobacterium animalis ssp. lactis LMG P-28149 |
| US20160213702A1 (en) * | 2015-01-26 | 2016-07-28 | Kaleido Biosciences, Inc. | Glycan therapeutics and related methods thereof |
| CN110446532A (en) * | 2016-12-12 | 2019-11-12 | Gi动力公司 | Treatment method relevant to stomach and intestine implantation material |
| CN108085285A (en) * | 2018-01-25 | 2018-05-29 | 吉林省命之元生物科技有限公司 | One DM-50 plants of lactobacillus plantarum and its application |
| US20190240100A1 (en) * | 2018-02-02 | 2019-08-08 | Augusta University Research Institute, Inc. | Compositions and Methods for Treating Diabetes |
| CN110150669A (en) * | 2019-04-29 | 2019-08-23 | 河北一然生物科技有限公司 | A kind of probiotic composition and its application suitable for patients with diabetes mellitus |
| CN110604749A (en) * | 2019-08-30 | 2019-12-24 | 北京农学院 | Bifidobacterium animalis and its use in controlling diabetes or hyperlipidemia, especially weight gain or obesity |
| CN114377037A (en) * | 2022-02-22 | 2022-04-22 | 内蒙古蒙牛乳业（集团）股份有限公司 | Composition of lactobacillus paracasei LC-37 and application thereof in reducing blood sugar |
| CN114752529A (en) * | 2022-04-29 | 2022-07-15 | 科郦有限公司 | Lactobacillus plantarum HOM3201 strain, viable bacteria preparation thereof, preparation method and application |
Non-Patent Citations (5)
| Title |
|---|
| AOKI, R.等: "A proliferative probiotic Bifidobacterium strain in the gut ameliorates progression of metabolic disorders via microbiota modulation and acetate elevation", 《SCI REP》 * |
| CHUNG-HSING WANG等: "Adjuvant Probiotics of Lactobacillus salivarius subsp. salicinius AP-32,L.johnsonii MH-68,and Bifidobacterium animalis subsp. lactis CP-9 Attenuate Glycemic Levels and Inflammatory Cytokines in Patient With Type 1 Diabetes Mellitus", 《FRONT ENDOCRINOL(LAUSANNE)》 * |
| STENMAN LK等: "Potential probiotic Bifidobacterium animalis ssp.lactis 420 prebents weight gain and glucose intolerance in diet-induced obese mice", 《BENEF MICROBES》 * |
| TONG LI等: "Bifidobacterium from breastfed infant faeces prevent high-fat-diet-induced glucose tolerance impairment,mediated by the modulation of glucose intake and the incretin hormone secretion axis", 《J SCI FOOD AGRIC》 * |
| 刘又嘉等: "益生菌预防2型糖尿病研究进展", 《中国微生态学杂志》 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN114350547B (en) | 2023-05-16 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| AU2016344770B2 (en) | Faecalibacterium prausnitzii and Desulfovibrio piger for use in the treatment or prevention of diabetes and bowel diseases | |
| CN110150669B (en) | Probiotic composition suitable for diabetic patients and application thereof | |
| CN106535907A (en) | Composition for treating or preventing metabolic disease, containing, as active ingredient, extracellular vesicles derived from akkermansia muciniphila bacteria | |
| CN114317353B (en) | Lactobacillus plantarum ZJFFYJ 7 and application thereof | |
| CN115153026B (en) | Application of lactobacillus rhamnosus strain LRa05 in preparation of helicobacter pylori and/or streptococcus mutans preparation inhibition | |
| EP3815702A1 (en) | Composition for promoting glucolipid metabolism, and preparation and application thereof | |
| CN113249280B (en) | Streptococcus thermophilus STN26, bacterium powder and application in uric acid reducing product | |
| CN113755409B (en) | Bifidobacterium longum for relieving insulin resistance and application thereof | |
| US20210128649A1 (en) | Composition for promoting glucolipid metabolism, and preparation and application thereof | |
| CN116121154B (en) | Leuconostoc lactis and application thereof | |
| CN111647526B (en) | Lactobacillus plantarum SD-H9, probiotic microbial inoculum thereof and application thereof | |
| CN113308421A (en) | Lactobacillus plantarum BUFX and application thereof in metabolic syndrome | |
| CN115786207A (en) | Lactobacillus salivarius and application thereof in preparation of medicines for treating and/or preventing diabetes | |
| CN108295096B (en) | Application of bacterial strain in preparation for preventing and treating mycotic and bacterial vaginosis | |
| CN114381411B (en) | Lactococcus lactis JYLL-60 and application thereof in preparation of product for improving immunity | |
| WO2017020786A1 (en) | Application of bacteroides fragilis in treating and/or preventing obesity or diabetes | |
| CN116254190A (en) | Lactobacillus paracasei subspecies and application thereof | |
| CN116855413B (en) | Bioactive substance for regulating human body microecological balance prepared from lactobacillus rhamnosus YSs069 and application thereof | |
| WO2005077390A1 (en) | Hypoglycemic agent, remedy/preventive for diabetes and proces for producing the same | |
| CN114350547B (en) | Bifidobacterium lactis strain B-622 and application thereof in preparation of medicines for treating diabetes | |
| CN113969253B (en) | Bifidobacterium lactis JYBR-390 with constipation treatment effect and application and product thereof | |
| CN113005066B (en) | Compound bifidobacterium preparation for resisting allergy, increasing immunity, reducing blood sugar and fat and losing weight and preparation method thereof | |
| CN113249264B (en) | Bifidobacterium adolescentis and application thereof in metabolic syndrome | |
| CN114806953A (en) | Lactobacillus gasseri with characteristic of improving type 1 diabetes | |
| CN110839693B (en) | Application of parabacteroides gibsonii in preventing or treating obesity or related diseases |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20231218 Address after: Room 2404 and 2405, 24th Floor, Unit 3, Building 1, No. 318 West Shun Street, Jinniu District, Chengdu City, Sichuan Province, 610036 Patentee after: Sichuan Min'an Zhichuang Technology Co.,Ltd. Address before: No.32, Section 2, West 1st ring road, Chengdu, Sichuan 610072 Patentee before: Sichuan Academy of Medical Sciences. Sichuan people's Hospital |
|
| TR01 | Transfer of patent right |