熱の予防によりラットに悪影響をストレス<em>枯草菌</em>ひずみ
Summary
We described a protocol for prevention of heat stress effects in rats by oral pre-treatment with beneficial bacteria. This protocol can be modified and used for various routes of administration and for analysis of different compounds.
Abstract
この研究は、熱ストレスに関連する合併症に対するバチルス・ズブチリス株の保護効果を評価するために設計しました。 32個のSprague-Dawleyラットをこの研究で使用しました。動物は、経口Bとの 2日間、1日2回処理しました。 サブチリス BSB3株またはPBS。最後の処置後の翌日、各グループを分割し、2つの実験群（PBS及び枯草菌で処理したものと処理したもの）25分間、45°Cのに置きました。 2つの対照群は、室温で25分間滞在しました。全てのラットを安楽死させ、別のパラメータは、すべての群で分析しました。熱ストレスの悪影響が絨毛の高さと腸上皮における総粘膜の厚さの減少により登録されています。内腔からの細菌の転。赤血球および血液中のリポ多糖（LPS）レベルの上昇の小胞形成を増加させました。治療の予防効果は、事前によって評価されますこれらの副作用の発明。プロトコルは、熱ストレスの合併症の予防のための細菌とラットの経口治療のために設立されたが、このプロトコルは、他の投与経路のために、異なる化合物の分析のために修正して使用することができます。
Introduction
Different stressors affect human and animal health. Temperature is one of the most stressful factors, causing chronic, acute and even lethal illnesses1. Changes in intestinal morphology and a loss of gut barrier integrity after heat stress were documented in many cases2,3. This protective barrier is responsible for defense against translocation of gut bacteria and their toxins, in particular lipopolysaccharides (LPS), from the lumen to the internal circulation4,5. Stability of the gut microbiota significantly influences intestinal barrier function, the immune response of the host, and tolerance to stress conditions6. Thus, modulation of the intestinal microbiota can provide a novel approach for prevention of stress-related adverse effects.
Beneficial bacteria have been used as a promising strategy to modify the gut microbiota for successful management of various clinical conditions, such as diarrhea, inflammatory bowel disease, atopic dermatitis, metabolic disorders7-10. Probiotic effects of beneficial bacteria include production of essential metabolites to support intestinal health, stimulation of the immune system, promotion of lactose tolerance, restoration of epithelial dysfunction. Probiotics were effective in prevention of the stress-related complications in vitro and in animal models11,12.
Researchers have given more attention to Bacillus bacteria as probiotics because these bacteria support intestinal homeostasis13 and have beneficial effects on the host14. Our previous data demonstrated high efficacy of Bacillus probiotics against pathogens in vitro15,16 and in clinical trials17,18. Here, we aim to evaluate the preventive effect of B. subtilis BSB3 strain against complications after heat stress.
Protocol
Representative Results
Discussion
深刻な健康状態29内の高温の結果への暴露。予防と熱ストレスに関連する合併症の早期発見は極めて重要30です。 提示されたプロトコルは、熱ストレスの有害作用の防止のための様々なアプローチの有効性を評価するために使用することができます。
このプロトコル内の重要なステップは次のとおり熱処理手順（45°Cの 、25分）。エ?…
Divulgazioni
The authors have nothing to disclose.
Acknowledgements
This work was supported by the Auburn University fund FOP 101002-139294-2050.
Materials
| Phosphate buffered saline (PBS) | Sigma-Aldrich, St. Louis, MO | P4417 | |
| Ethyl Alcohol | Pharmco products Inc. Brookfield, CT, USA | 64-17-5 | |
| Agar | VWR | 97064-334 | |
| MacConkey agar plates | VWR | 470180-742 | |
| 5% blood agar plates | VWR | 89405-024 | |
| Brucella blood with Hemin, and Vitamin K, plates |
VWR | 89405-032 | |
| Bouin’s Fixative | Electron Microscopy Sciences, Hatfield, PA, USA | 15990 | |
| IL-10 Rat ELISA kit | Invitrogen, Camarillo, CA, USA | KRC0101 | |
| IL-1beta Rat ELISA Kit | Invitrogen, Camarillo, CA, USA | KRC0011 | |
| IL-6 Rat ELISA Kit | Invitrogen, Camarillo, CA, USA | KRC0061 | |
| INF-gamma Rat ELISA Kit | Invitrogen, Camarillo, CA, USA | KRC4021 | |
| TNF-alpha Rat ELISA Kit | Invitrogen, Camarillo, CA, USA | KRC3011 | |
| Rat LPS ELISA Kit | NeoBioLab, MA, USA | RL0275 | |
| Environmental Chamber 6020-1 | Caron, Marietta, OH, USA | 6020-1 | |
| Centrifuge | Beckman Coulter, Indianapolis, IN, USA | Optima L-90K | |
| Ultra Centrifuge | |||
| Light microscope optical system | CitoViva Technology Inc., Auburn, AL | ||
| Colony counter | Fisher Scientific , Pittsburgh, PA, USA | RE-3325 | |
| Freezer | Haier, Brooklyn, NY, USA | HCMO50LA | |
| Ultra microplate reader | Bio-Tek Instrument, Winooski, VT, USA | ELx 808 | |
| Auto Strip Washer | Bio-Tek Instrument, Winooski, VT, USA | ELx50 | |
| Pipettes | Gilson, Pipetman, France | P100, P200, P1000 | |
| C24 Incubator Shaker | New Brunswick Scientific, Enfield, CT, USA | Classic C24 | |
| Rocking Shaker | Reliable Scientific, Inc., Nesbit, MS, USA | 55 | |
| Petri dishes | Fisher Scientific, Pittsburgh, PA, USA | 875713 | 100mmX15mm |
| SterilGard III Advance | The Baker Company, Sanford, ME, USA | SG403 | |
| Culture Growing Flasks | Corning Incorporated, Corning, NY, USA | 4995 | PYREX 250mL Erlenmeyer flasks |
| Optical Spectrometer Genesys 20 | Thermo Scientific, Waltham, MA, USA. | 4001 | |
| Sony DXC-33 Video camera | Sony | ||
| Richardson test slide | Electron Microscopy Science, Hatfield, PA, USA | 80303 | |
| Millipore water purification system | Millipore | Direct-Q | |
| Image-Pro Plus software | Media Cybernetics, MD, USA | ||
| Triple Beam Balance | OHAUS Corporation, Parsippany, NJ, USA | ||
| Tissue Homogenizer, Dounce | VWR | 71000-518 | |
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