The Effect of Intralesian Injection of Umbilical Cord Mesenchymal Stem Cells, Its Conditioned Medium, and Triamcinolone Acetonide on Type 1:3 Collagen Ratio and Interleukin-10 Levels in Keloid: A Randomised Controlled Trial
Condition: Keloid; Stem Cell
MeSH terms: Keloid
Study type: Interventional
Phase: Phase 4

Summary: 
      The objective of this study is to observe the potency of umbilical cord-derived mesenchymal
      stem cells (UC-MSC) and umbilical cord-derived conditioned medium (UC-CM), or triamcinolone
      acetonide (TA) in keloid therapy, measured by the decrease in the type 1:3 collagen ratio and
      the increase of IL-10 levels carried out using CONSORT statement.
    

Detailed description: 
      Screening of patients were done by measuring keloid dimensions using a ruler. Patients
      meeting the inclusion criteria were then randomly divided into 3 groups. The same injection
      volume (1 ml) in every cm3 keloid volume was given using a 1 mL syringe and 27G needle to the
      subjects. Ultrasound guidance was used to administer the injection using an in-plane
      technique into the center of the lesion, with inclinations of 30-45 degrees, ensuring uniform
      pressure in every subject. Group 1 was given UC-MSC 2 million cells/mL/cm3, group 2 receives
      UC-CM 1mL/cm3 while group 3 was administered TA 40 mg/mL/cm3. In order to obtain UC-MSC and
      UC-CM, 10 cm of umbilical cord tissue was collected in 50 mL transport medium containing the
      following substances: amphotericin B (Final concentration 7500 ng/ml [JR Scientific 50701]),
      alpha minimal essential medium (MEM [GIBCO 12000-022 1]), penicillin/streptomycin (final
      concentration 300 U/ml [Gibco 15140-122]). Samples were then processed within 8 hours of
      collection. The umbilical cord was then dissected, briefly washed in 0.5% povidone-iodine
      (betadine ©) with phosphate buffered saline of pH 7.4 (PBS [Sigma P3813]), and then washed in
      PBS to remove betadine and blood. The umbilical vessels were excised before the umbilical
      cord was minced into a complete medium. UC-CM was created using Alpha-MEM and Dulbecco's
      modified Eagles medium (DMEM [GIBCO 31600-034]). The finalized complete medium contains 1%
      L-glutamine (Lonza 17-605C), 10% TC (Indonesian Red Cross), amphotericin B (final
      concentration 2500 ng/ml), and penicillin/streptomycin (final concentration 100 U/ml).
      Supplementation of culture with 10% autologous or allogeneic cord blood serum and 10% human
      AB serum (Gibco 34005-100) was done to create MEM. Each well of a 12-well plate (growth are
      3.8 cm2 [Biolite]) was filled with three explants (diameter 2-5 mm) and Wharton's Jelly,
      followed by addition of several drops of complete medium. Triplication of culture was done to
      each medium. The plate was incubated at 37°C and 5% CO2 and observed daily for contamination
      or cellular growth, in which the contaminated wells were removed. After the explants was
      attached, addition of 200-500 µL of appropriate medium was done. Medium changes, involving
      removal of half of the medium and addition of half of each medium, were performed every 2-3
      days. Growth of the cells beyond the explants with 90% confluence marks viability for
      harvesting using TrypLE Select (GIBCO 12563-011). Dye exclusion method was used to count the
      viable/non-viable cell yield. After each harvest, with the explant still attached to the
      plate, new appropriate medium was added with the plate being re-incubated at 37°C and 5% CO2.
      Similar treatment was repeated for the second and subsequent cultures, enabling multiple
      harvests from a single explant. Biopsies of the keloid tissues were then conducted twice, in
      the first meeting and 17 weeks after. Parameters to be evaluated in anatomic pathology
      examination are Sirius red staining to evaluate collagen structure under a polarizing lens,
      as well as In Vitro quantitative examination using the ELISA method to examine IL-10.
      Calculation of changes in the ratio of type 1 to type 3 collagen levels was carried out by
      dividing the ratio of collagen before treatment from the ratio of collagen after each
      treatment. When visualized under a polarizing lens, type-3 collagen will appear
      green-birefringence and type-1 collagen will appear yellow-birefringence. The collagen ratio
      was obtained by dividing the composition of collagen type 1 to collagen type 3 in Sirius red
      staining under polarizing lenses.
    

Intervention name(s): umbilical cord-derived mesenchymal stem cells (UC-MSC); umbilical cord-derived mesenchymal stem cells conditioned medium (UC-CM); Triamcinolone Acetonide (TA)
Intervention type(s): Biological; Biological; Drug
Intervention description(s): Group 1 was injected UC-MSC 2 million cells/mL/cm3, given the same injection volume (1 mL) in every cm3 keloid volume using a 1 mL syringe and 27G needle. The injections were ultrasound-guided into the center of the lesion with a 30-45-degree angle using an in-plane technique, therefore the same pressure.; Group 2 was injected UC-CM 1 mL/cm3, given the same injection volume (1 mL) in every cm3 keloid volume using a 1 mL syringe and 27G needle. The injections were ultrasound-guided into the center of the lesion with a 30-45-degree angle using an in-plane technique, therefore the same pressure.; Group 3 was injected TA 40 mg/mL/cm3, given the same injection volume (1 mL) in every cm3 keloid volume using a 1 mL syringe and 27G needle. The injections were ultrasound-guided into the center of the lesion with a 30-45-degree angle using an in-plane technique, therefore the same pressure.
Intervention MeSH terms: Triamcinolone; Triamcinolone Acetonide; Triamcinolone hexacetonide; Triamcinolone diacetate
Allocation: Randomized
Intervention model: Parallel Assignment
Intervention model description: Patients were screened initially by measuring the length and thickness of the keloids using a ruler. Patients who met the study inclusion criteria were randomly divided into 3 groups. Each patient in the groups was given the same injection volume (1 mL) in every cm3 keloid volume using a 1 mL syringe and 27G needle. The injections were ultrasound-guided into the center of the lesion with a 30-45-degree angle using an in-plane technique, therefore the same pressure. Group 1 was given UC-MSC 2 million cells/mL/cm3, group 2 was given UC-CM 1 mL/cm3, and group 3 was given TA 40 mg/mL/cm3.
Masking: Quadruple (Participant, Care Provider, Investigator, Outcomes Assessor)
Primary purpose: Treatment
Keywords: Keloid therapy; mesenchymal stem cells; Interleukin-10; collagen ratio