an innovation for in vitro cell exposure Hario Science Co., Ltd. about us Tokyo, Japan Scientific branch of Hario Glass Corporation Kazuo Kanazawa Hario Science Co., Ltd. 2/34 Laboratory glassware Hario Science Co., Ltd. products Animal Testing Equipments Inhalation Toxicity Testing Test substances are exposed through respiratory organs. General Scientific Equipments Animal Testing Equipments nasal cavity nose mouse larynx trachea fharynx esophagus bronchi bronchioli alveoli 3/34 4/34 Animal Testing Equipments Inhalation Toxicity Testing Typical setup for inhalation exposure testing Generator Test substance generation Animals are inhaled inside chamber Animal Testing Equipments Inhalation Toxicity Testing in vivo inhalation exposure Whole body exposure Nose only exposure Cultex exposure system System Control Unit Exposure Chamber Exhaust Air Treatment Inhale Air Aerosol concentration control/monitoring Environment Measuring Devices Exhaust air treatment Exhaled Air 5/34 6/34 1
An alternative method for inhalation exposure testing. Patented in 1999 Many publications Various line up Cultex exposure system Aufderheide et al, Novel approaches for studying pulmonary toxicity in vitro 7/34 8/34 An alternative method for inhalation exposure testing. Conventional method Air Liquid Interface (ALI) exposure cell culture insert cell medium Air Liquid Interface(ALI) Exposure: Cells are alive on micro pores membrane Cells are exposed direct to test substances. ALI environment mimics living body 9/34 10/34 Set Air Liquid Inside into Cultex Interface incubator module Exposure Medium Cell from are bottom grown Medium Exhaust Aerosols Exhaust Cell culture insert Cells Medium Cells (primary / cell line): A549 (Human lung adenocarcinoma epithelial cell line) HFBE 21 (Human lung epithelial cell) Lk004 (Human lung fibroblasts) HaCaT (Human skin keratinocyte) CHO (Chinese hamster ovary cell) etc... Bacteria: S. Typhimurium Escherichia coli etc... 11/34 12/34 2
Evaluations: WST 1 (proliferation assay) LDH (cytoxicity (y yassay) IL 8 (inflammation test) Comet assay (genotoxicity) Glutathione (oxidative stress test) ROS (oxidative stress) neutral red (cytotoxicity) Heme oxygenase (oxidative stress) Ames assay (genotoxicity) 13/34 14/34 Clean bench Cigarette smoke exposure Aerosol control Exhaust air control Aerosol generator CULTEX module Suction pump Water bath 15/34 16/34 17/34 18/34 3
Cell culture unit (Bottom module) Exposure unit (Top module) Three chambers are located radially Electrostatic charger to improve deposition 19/34 20/34 Medium flow warm water circulation Cell culture insert Cell culture insert Water in Water out Medium out Medium in 21/34 22/34 Aerosol flow aerosol Aerosol flow aerosol exhaust 23/34 24/34 4
Aerosol flow Warm water circulation Water in Water out aerosol 25/34 26/34 maintains cell culture and exposure Aerosol supply CULTEX RFS Various kind/size of cell culture insert Warm ater supply Warm water supply Medium supply 27/34 28/34 Various kind/size of cell culture insert Various kind/size of cell culture insert 29/34 30/34 5
Cigarette smoke exposure Cigarette smoke exposure 31/34 32/34 Test substance: Chemicals Pharmaceutical products Consumer products Complex atmospheres Environmental aerosols and more... Applications: Risk Assessment Screening Biomarker and more... Reduction of animal testing Cost & Time effective Endpoints: Cytotoxicity i Mutagenicity Inflammatory & allergic reactions Organ specific toxicity and more... ขอบค ณ มาก คร บ 33/34 34/34 6