In Vivo Pharmacology

Inflammation


We offer a range of in vitro, ex vivo & in vivo models for identification & confirmation of anti – inflammatory potential of NCEs & formulations

  • Acute Inflammation Models
    • Carageeneen Induced
      Model Carrageenan induced paw edema
      Test system Rat or Mice
      Method
      • Administration of test item at appropriate dose and route
      • Inject carrageenan at sub-plantar region
      • Measure the paw volume change at defined time points
      End points
      • Paw volume change at predetermined time points
      • % Inhibition or ED50 Determination
      X
    • Xylene Induced
    • LPS Induced
      Model Cytokine profiling of potential anti-inflammatory compounds in LPS-induced murine endotoxic shock model
      Test system Balb/C mice, Female, 20-30 g
      Method
      • Preadministration of mice with test compounds
      • Septic shock induction with LPS
      • Collection of blood after 90 minutes
      • Estimation of cytokines in serum samples using ELISA
      End points
      • Quantitation of TNF-a, IL-1-b & IL-6
      X
  • Chronic Inflammation Models
    • MIA Induced OA
    • Collagen Induced RA
      Model Collagen Induced Arthritis (CIA)
      Test system Mouse or Rat
      Method
      • Sub-plantar immunization of collagen with appropriate adjuvant
      • Monitor the development of inflammatory response
      • Dosing for defined dose and schedule
      • Observe the parameters during and post compound administration
      End points
      • Paw volume change
      • Clinical Scoring
      • Body Weight
      • Hyperalgesia
      • Photomicrograph
      • Cytokine levels (Serum & Homogenate)
      • Histopathology
      X
    • DSS / TNBS Induced Colitis
      Model Inflammatory Bowel Disease (IBD)
      Method Dextran Sodium Sulfate (DSS) induced Colitis in Mice Model
      Test system C57BL/6J mice, Male, 20-30 g
      Method
      • Randomize the animal based on body weight
      • 3% DSS solution will be given from day 1 as drinking solution
      • Administration of test item for 7 days
      • Daily body weight and Disease Activity Index (DAI) will be recorded.
      • On day 7, all animals will be Humane sacrifice to measure colon length and weight
      End points
      • Body weight change
      • Colon weight and length
      • Disease Activity Index (DAI) based on weight loss, Stool consistency and Fecal Occult Blood
      • Histopathological slides and scoring
      • Colon homogenate – Colon MPO activity (Optional)
      Model Inflammatory Bowel Disease (IBD)
      Method TNBS induced in Mice Model
      Test system BALB/c mice, Male, 20-30 g
      Method
      • Randomize the animal based on body weight
      • Intrarectal administration of TNBS for all the animals
      • Administration of test item for 7 days
      • Daily body weight and Clinical observation will be done.
      • On day 7, all animals will be Humane sacrifice to measure colon length and weight
      • Colon histopathology.
      End points
      • Body weight
      • Colon weight and length
      • Histopathological of colon and its scoring
      • Clinical scoring for IBD
      • Colon homogenate – Colon MPO activity (Optional)
      X
    • Oxazolone / DNFB Induced DTH Inflammation
      Model Delayed Type Hypersentivity (DTH) Reaction
      Method
      • Oxazolone induced DTH inflammation Model
      • 2,4-DNFB induced DTH inflammation Model
      Test system Balb/C mice, Female, 20-30 g
      Method
      • On day 1 apply 150µL of 3% Oxazolone solution /30µL of 0.5% DNFB solution (Acetone : olive oil) to the shaved abdomen area
      • On day 7 animals will be challenged with 20µL of 3% Oxazolone/20µL of 0.2% DNFB on the right ear
      • Test item will be applied topically to the ear 6h and 23h after the challenge
      • Ear thickness will be measured before treatment (0h), 6h and 23h after the challenge
      • Animals will be sacrificed & biopsies of both the ears will be taken
      End points
      • Ear thickness/Ear edema
      • Ear punch weight
      • %change in ear punch weight
      • Myeloperoxidase activity
      • Cytokine estimation-Optional (IL-1beta, IL-4. MIP-1alpha, MCP-1, KC, RANTES, IP-10 and G-CSF)
      X


In Vivo Models for anti – inflammatories

  • Carrageen induced paw edema
    Model Carrageenan induced paw edema
    Test system Rat or Mice
    Method
    • Administration of test item at appropriate dose and route
    • Inject carrageenan at sub-plantar region
    • Measure the paw volume change at defined time points
    End points
    • Paw volume change at predetermined time points
    • % Inhibition or ED50 Determination
    X
  • Collagen induced arthritis
    Model Collagen Induced Arthritis (CIA)
    Test system Mouse or Rat
    Method
    • Sub-plantar immunization of collagen with appropriate adjuvant
    • Monitor the development of inflammatory response
    • Dosing for defined dose and schedule
    • Observe the parameters during and post compound administration
    End points
    • Paw volume change
    • Clinical Scoring
    • Body Weight
    • Hyperalgesia
    • Photomicrograph
    • Cytokine levels (Serum & Homogenate)
    • Histopathology
    X
  • LPS induced inflammation
    Model Cytokine profiling of potential anti-inflammatory compounds in LPS-induced murine endotoxic shock model
    Test system Balb/C mice, Female, 20-30 g
    Method
    • Preadministration of mice with test compounds
    • Septic shock induction with LPS
    • Collection of blood after 90 minutes
    • Estimation of cytokines in serum samples using ELISA
    End points
    • Quantitation of TNF-a, IL-1-b & IL-6
    X
  • InVivo models for chronic inflammatory pain
    Model MIA (Mono sodiumiodoacetate) Induced Osteoarthritis (OA)
    Test system Wistar Rat
    Method
    Injection 50µL of MIA ( Contains 3mg ) to patellar ligament of right knee Inflammation of synovial joints (~5days) Selection, randomization and grouping Administration of test item (TI) at required dose Scoring of Nocturnal mobility & Microscopic examination of isolated arthritic knee Estimation of S-GAG ( Sulphated Glycosaminoglycan) and PGE2 in synovial lavage.
    End points
    Mobility Scoring, Bone weight of patella, Histopathology of knee joint and estimation of S-GAG & PGE2
    Model Adjuvant-Induced Arthritis (RA)
    Test system Wistar Rat / Lewis Rat
    Method
    Injection 0.1ml of Freund’s Complete Adjuvant (DIFCO) to left hind paw 12 day after FCA injection, Administration of test item (TI) at required dose and route Dosing should be continued till day 28. Animals will be observed for Inflammatory parameters like paw volume, joint diameter, Clinical scoring, Pain response etc… At terminal stage, Blood will be collected for biochemical marker estimation Paw will be subjected for radiography and histopathological analysis
    End points
    Arthritic Scoring, Paw Volume, Joint diameter, Von Frey filament test, Joint pain by PAM, Biochemical markers ( TNF alpha, IL1beta, IL6, MPO activity, Hematology & RA Factor, ESR etc…) Histopathology of ankle joint and X-Ray analysis
    Model Collagen Induced Arthritis (CIA)
    Test system DBA/1 mice / Lewis rat
    Method
    Immunization with 100µL emulsion of CFA ( Contains 100µg of C-II & 250µg of M.tuberclosis) On day 21 booster injection of CFA Selection, randomization and grouping Administration of test item (TI) at required dose Measure paw volume using plethysmometer Clinical scoring for arthritic index Collect serum for cytokine estimation, X-ray analysis, spleen weight and Hb level
    End points
    inhibition, arthritic index, Histopathology etc.
    Model Evaluation of Test Items on Lipopolysaccharide (LPS) Induced TNF-a Release in Mice
    Test system Mice/Balb/c / Male
    Method
    • Mice will be randomized and grouped based on the body weight and treated with test item for 6 days (-6, -5, -4, -3, -2 ,-1). On day “0” an hour post dosing, the animals will be challenged with LPS 50 - 100µg/animal (as per sponsor’s suggestion).
    • Ninety minutes post challenge (LPS), blood will be collected; serum separated and samples subjected for TNFa analysis. At the end of the experiment the animals will be humanely sacrificed.
    End points
    • TNF-a levels in serum
    X
  • InVivo models for chronic inflammatory pain
    Model MIA (Mono sodiumiodoacetate) Induced Osteoarthritis (OA)
    Test system Wistar Rat
    Method
    Injection 50µL of MIA ( Contains 3mg ) to patellar ligament of right knee Inflammation of synovial joints (~5days) Selection, randomization and grouping Administration of test item (TI) at required dose Scoring of Nocturnal mobility & Microscopic examination of isolated arthritic knee Estimation of S-GAG ( Sulphated Glycosaminoglycan) and PGE2 in synovial lavage.
    End points
    Mobility Scoring, Bone weight of patella, Histopathology of knee joint and estimation of S-GAG & PGE2
    Model Adjuvant-Induced Arthritis (RA)
    Test system Wistar Rat / Lewis Rat
    Method
    Injection 0.1ml of Freund’s Complete Adjuvant (DIFCO) to left hind paw 12 day after FCA injection, Administration of test item (TI) at required dose and route Dosing should be continued till day 28. Animals will be observed for Inflammatory parameters like paw volume, joint diameter, Clinical scoring, Pain response etc… At terminal stage, Blood will be collected for biochemical marker estimation Paw will be subjected for radiography and histopathological analysis
    End points
    Arthritic Scoring, Paw Volume, Joint diameter, Von Frey filament test, Joint pain by PAM, Biochemical markers ( TNF alpha, IL1beta, IL6, MPO activity, Hematology & RA Factor, ESR etc…) Histopathology of ankle joint and X-Ray analysis
    Model Collagen Induced Arthritis (CIA)
    Test system DBA/1 mice / Lewis rat
    Method
    Immunization with 100µL emulsion of CFA ( Contains 100µg of C-II & 250µg of M.tuberclosis) On day 21 booster injection of CFA Selection, randomization and grouping Administration of test item (TI) at required dose Measure paw volume using plethysmometer Clinical scoring for arthritic index Collect serum for cytokine estimation, X-ray analysis, spleen weight and Hb level
    End points
    inhibition, arthritic index, Histopathology etc.
    Model Evaluation of Test Items on Lipopolysaccharide (LPS) Induced TNF-a Release in Mice
    Test system Mice/Balb/c / Male
    Method
    • Mice will be randomized and grouped based on the body weight and treated with test item for 6 days (-6, -5, -4, -3, -2 ,-1). On day “0” an hour post dosing, the animals will be challenged with LPS 50 - 100µg/animal (as per sponsor’s suggestion).
    • Ninety minutes post challenge (LPS), blood will be collected; serum separated and samples subjected for TNFa analysis. At the end of the experiment the animals will be humanely sacrificed.
    End points
    • TNF-a levels in serum
    X
  • Target Based Screens fo Interstitial cystitis
    Model in vivo Model for screening Interstitial cystitis
    Test system Wistar /SD
    Method
    Day 0 (Intravesical instillation of Acetic acid/hydrochloric acid (irritant) Administration of test item and reference item at predetermined time Interim sacrifice of the animals to confirm the disease induction At the end of the experiment collection of bladder for further evaluation.
    End points
    • TNFa (bladder tissue)
    • Nitric oxide (bladder tissue)
    • Histopathology evaluation (bladder tissue)
    X