Diabetic Foot Ulcer
Conditions
Keywords
ultrasound assisted wound debridement, surgical debridement, cellular proliferation, microbiology, diabetic foot ulcers
Brief summary
The investigators aimed to elucidate the effects of UAW debridement on cellular proliferation and dermal repair in complicated diabetic foot ulcers as compared to diabetic foot ulcers receiving surgical/sharp wound debridement. A randomized controlled trial was performed involving outpatients with complicated diabetic foot ulcers that either received surgical debridement or UAW debridement every week during a six-week treatment period.
Detailed description
2\. Methods 2.1. Trial design A randomized and controlled parallel clinical trial was performed involving outpatients with complicated DFU that were admitted to specialized diabetic foot unit between November 2017 to December 2019. The study protocol received full approval from the Ethics Committee of the Hospital Clínico San Carlos, Madrid, Spain (C.P. - C.I. 16/484-P). Each patient provided written informed consent before inclusion. 2.1. Intervention Participants were randomized and assigned to receive either surgical debridement or UAW debridement every week during a six-week treatment period. Soft tissue punch biopsies (3mm) were taken after wound debridement sessions at week zero and week six. 2.2. Follow-up Patients were followed-up for 6 months after inclusion. During the follow-up period, the investigators recorded ulcer healing. Ulcer healing was defined as complete epithelialization without any sustained drainage up to 24 weeks after the end of the study follow-up. 2.3. Sample size The sample size was calculated using the Granmo v.12 program (Municipal Institute of Medical Research, Barcelona, Spain) (https://www.imim.cat/ofertadeserveis/software-public/granmo/ ). Therefore, we analyzed 51 patients (24 in surgical group and 27 en UAW group) with an alpha of 0.05 and a statistical power of 0.80. 2.4. Randomization A computer-generated random number table was used to carry out the randomization of the patients into the two groups by an investigator who was blinded to the identity of the participants. 2.5. Blinding None of the participants, care providers, and outcome adjudicators was blinded to the interventions after assignment. 2.6. Statistical Analysis Statistical analysis was performed using SPSS for IOs version 21.0 (SPSS, Inc. Chicago, IL, USA). The assumption of normality of all continuous variables was verified using the Kolmogorov-Smirnov test. Statistical differences between groups were calculated using the Chi-Square test and, where appropriate, Fisher's exact test for categorical variables. The Mann-Whitney U test was performed for abnormally distributed quantitative parameters, and Student's t-test was performed for quantitative variables that were distributed normally. The criteria of p \< 0.05 was accepted as statistically significant with a confidence interval of 95%. This study was conducted in accordance with the Declaration of Helsinki (2013 revision) and followed all local laws and regulations in clinical research investigations in patients.
Interventions
PROCEDUREUltrasound group
Every week during a six-week treatment period
PROCEDURESurgical group
Every week during a six-week treatment period
Sponsors
Universidad Complutense de Madrid
Francisco Javier Álvaro Afonso
David Sevillano Fernández
Yolanda García Álvarez
Irene Sanz Corbalan
Esther García Morales
Study design
Allocation
RANDOMIZED
Intervention model
PARALLEL
Primary purpose
TREATMENT
Masking
SINGLE (Subject)
Intervention model description
A randomized and controlled parallel clinical trial
Eligibility
Sex/Gender
ALL
Age
18 Years to No maximum
Healthy volunteers
No
Inclusion criteria
* • Male and female patients ≥18 years old * Type 1 or type 2 diabetes with levels of HbA1c≤85.8 mmol/mol (10%) within 30 days of the beginning of the study * Wound stages IB, IIB, ID, and IID according to the University of Texas Diabetic Wound Classification \[11\] * Wound duration of 1-24 months * Wound size between 1-30 cm2 after debridement * Clinical picture of wounds showing mild or moderate infection, according to the criteria of the Infectious Disease Society of America Guidelines \[12\] and the European Wound Management Association (EWMA) \[13\] * Ankle-brachial index (ABI) ≤0.9 and ankle systolic blood pressure (ASBP) ≥70mmHg, or toe systolic blood pressure (TSBP) ≥50mmHg, ABI\>0.9, TSBP ≥50mmHg and toe-brachial index (TBI) ≤0.7
Exclusion criteria
* • Chronic renal disease or dialysis * Non-treated osteomyelitis * Necrotizing soft tissue infections * Critical limb ischaemia patients with ABI≤0.5 and ASBP\<70mmHg or \<50mmHg * Life expectancy \<6 months due to malignant DFU * Pregnancy and lactation * Patients diagnosed with hepatitis or human immunodeficiency virus (HIV) * Patients showing local or systemic conditions that may impair tissue repair
Design outcomes
Primary
| Measure | Time frame | Description |
|---|---|---|
| Change From Baseline Neo-angiogenesis (Microvessel Density) at 6 Weeks | At week zero and week 6 | Sections of tissue were immunohistochemically-stained with the CD31 marker. Light microscopy was used to count the number of microvessels/endothelial cells in a standardized grid, with the results expressed as microvessel density (Leica DMD 800 morphometric system). Microvessel density was scored according to the following scale: 0 (absent), 1 (low, at least one microvessel), 2 (moderate) and 3 (more than two micro vessels). Higher scores mean a better outcome |
| Change From Baseline Collagen Formation (Collagen Content ) at 6 Weeks | At week zero and week 6 | Massons's trichome staining was used to differentiate collagen content from other components, such as muscle fibrin and erythrocytes, in tissue samples. Collagen content was scored according to the following scale: 0 (absent), 1 (mild), 2 (moderate) and 3 (severe). Higher scores mean a better outcome. |
| Change From Baseline Myofibroblasts Formation (Myofibroblasts Content) at 6 Weeks | At week zero and week 6 | Actin staining was used to evaluate the presence of myofibroblasts involved in wound healing. These cells increase in number during wound healing. The number of stained cells was semi-quantitatively analyzed using a 0 - 3 scaling score (0= no myofibroblasts, 1= myofibroblasts in low quantity, 2= myofibroblasts in moderate quantity, 3= myofibroblasts in high quantity) |
Secondary
| Measure | Time frame | Description |
|---|---|---|
| Comparison of Quantitative Microbiological Analysis (Bacterial Counts Expressed Colony-forming Units Per Gram of Tissue) (CFU/g) | At week zero and week 6 | Tissue samples were weighed and mechanically homogenised in 0.5ml volumes of sterile phosphate buffered saline (PBS, Sigma Aldrich, St Louis, MO). Homogenates were diluted and plated onto Columbia agar (BD, Sparks, MD), Columbia agar supplemented with colistin and nalidixic acid (BD), MacConkey agar (BD), and Sabouraud dextrose agar (BD) using a spiral plater workstation (Don Whitley Scientific, Shipley, UK).The limit of detection was 10 colony-forming units (CFU). Results were expressed as CFU per gram of tissue (CFU/g). Isolated microorganisms were identified by standard criteria and the BBL Crystal identification system (BD). Susceptibility testing of Staphylococcus aureus isolates for oxacillin was performed according to Clinical and Laboratory Standards Institute (CLSI) guidelines, using a 30g cefoxitin disc and Mueller-Hinton agar . |
| Wound Score at 6 Weeks | Six weeks | Wound bed tissue was evaluated for presence, quality, and consistency of granulation tissue using a validated wound scoring system, with scores ranging between a minimum of zero points and maximum of seven points. Higher scores mean a better outcome. |
| Wound Size | 6 weeks | A planimetric measurements of wound size were conducted using Visitrak (Smith & Nephew, Hull, UK), with the area of the lesion determined with an approximation of ±5mm2 |
Countries
Spain
Participant flow
Recruitment details
An open-label randomized and controlled parallel clinical trial was performed involving outpatients with complicated Diabetic Foot Ulcer that were admitted to specialized diabetic foot unit between November 2017 to December 2019.
Pre-assignment details
51 patients met inclusion criteria and were were randomized to treatment
Participants by arm
| Arm | Count |
|---|---|
| Ultrasound Group (UAW Group) UAW debridement was performed using an UAW SONOCA 185 device (Söring GmbH, Germany). The UAW device generates an ultrasound low frequency of 25kHz and is equipped with three UAW instruments with different sonotrode shapes. The choice of sonotrode depends on wound depth, which ranges from superficial to deep. The UAW instrument piezoelectrically transforms the electrical energy delivered from the UAW device into mechanical oscillations in the sonotrode tip. For most wounds in the UAW group, a two-minute treatment with 40% intensity was performed by holding the sonotrode in contact mode, holding it perpendicular to the wound bed and moving it across in an up-and-down pattern.
Ultrasound group: For most wounds in the UAW group, a two-minute treatment with 40% intensity was performed by holding the sonotrode in contact mode, holding it perpendicular to the wound bed and moving it across in an up-and-down pattern. For wounds measuring \>15cm2, the debridement procedure was increased to three minutes. In addition to UAW debridement, a scalpel was used for careful tissue removal, but only if periwound skin exhibited calluses and maceration. | 27 |
| Surgical Group All debridement procedures were performed by the same surgeon (J.L.M.), who is specialist in diabetic foot surgery with more than 20 years of experience.
Surgical debridement involved removal of all necrotic and devitalized tissue that was incompatible with healing, as well as surrounding callus.
Ultrasound group: For most wounds in the UAW group, a two-minute treatment with 40% intensity was performed by holding the sonotrode in contact mode, holding it perpendicular to the wound bed and moving it across in an up-and-down pattern. For wounds measuring \>15cm2, the debridement procedure was increased to three minutes. In addition to UAW debridement, a scalpel was used for careful tissue removal, but only if periwound skin exhibited calluses and maceration. | 24 |
| Total | 51 |
Withdrawals & dropouts
| Period | Reason | FG000 | FG001 |
|---|---|---|---|
| Overall Study | Chronic renal disease, critical limb ischemia and heart attack | 1 | 2 |
Baseline characteristics
| Characteristic | Surgical Group | Ultrasound Group (UAW Group) | Total |
|---|---|---|---|
| Age, Categorical <=18 years | 0 Participants | 0 Participants | 0 Participants |
| Age, Categorical >=65 years | 2 Participants | 3 Participants | 5 Participants |
| Age, Categorical Between 18 and 65 years | 22 Participants | 24 Participants | 46 Participants |
| Age, Continuous | 58 years STANDARD_DEVIATION 5.4 | 64.1 years STANDARD_DEVIATION 12.4 | 61.1 years STANDARD_DEVIATION 8.9 |
| Duration of Diabetes | 10.3 years STANDARD_DEVIATION 5 | 22 years STANDARD_DEVIATION 12.9 | 16.15 years STANDARD_DEVIATION 8.95 |
| Glycated hemoglobin | 51 mmol/L STANDARD_DEVIATION 4.5 | 57 mmol/L STANDARD_DEVIATION 9.9 | 54 mmol/L STANDARD_DEVIATION 9.4 |
| Race (NIH/OMB) American Indian or Alaska Native | 0 Participants | 0 Participants | 0 Participants |
| Race (NIH/OMB) Asian | 0 Participants | 0 Participants | 0 Participants |
| Race (NIH/OMB) Black or African American | 0 Participants | 0 Participants | 0 Participants |
| Race (NIH/OMB) More than one race | 0 Participants | 0 Participants | 0 Participants |
| Race (NIH/OMB) Native Hawaiian or Other Pacific Islander | 0 Participants | 0 Participants | 0 Participants |
| Race (NIH/OMB) Unknown or Not Reported | 0 Participants | 0 Participants | 0 Participants |
| Race (NIH/OMB) White | 24 Participants | 27 Participants | 51 Participants |
| Region of Enrollment Spain | 24 participants | 27 participants | 51 participants |
| Sex: Female, Male Female | 0 Participants | 3 Participants | 3 Participants |
| Sex: Female, Male Male | 24 Participants | 24 Participants | 48 Participants |
| Texas Classification IB | 4 Participants | 4 Participants | 8 Participants |
| Texas Classification ID | 8 Participants | 12 Participants | 20 Participants |
| Texas Classification II B | 8 Participants | 3 Participants | 11 Participants |
| Texas Classification IID | 4 Participants | 8 Participants | 12 Participants |
| Type Diabetes Mellitus Type 1 | 0 Participants | 5 Participants | 5 Participants |
| Type Diabetes Mellitus Type 2 | 24 Participants | 22 Participants | 46 Participants |
Adverse events
| Event type | EG000 affected / at risk | EG001 affected / at risk |
|---|---|---|
| deaths Total, all-cause mortality | 0 / 27 | 1 / 24 |
| other Total, other adverse events | 0 / 27 | 0 / 24 |
| serious Total, serious adverse events | 0 / 27 | 0 / 24 |
Outcome results
Primary
Change From Baseline Collagen Formation (Collagen Content ) at 6 Weeks
Massons's trichome staining was used to differentiate collagen content from other components, such as muscle fibrin and erythrocytes, in tissue samples. Collagen content was scored according to the following scale: 0 (absent), 1 (mild), 2 (moderate) and 3 (severe). Higher scores mean a better outcome.
Time frame: At week zero and week 6
| Arm | Measure | Value (MEAN) | Dispersion |
|---|---|---|---|
| Ultrasound Group (UAW Group) | Change From Baseline Collagen Formation (Collagen Content ) at 6 Weeks | 2.81 score on a scale | Standard Deviation 0.48 |
| Surgical Group | Change From Baseline Collagen Formation (Collagen Content ) at 6 Weeks | 1.20 score on a scale | Standard Deviation 0.26 |
p-value: <0.01Wilcoxon (Mann-Whitney)
Primary
Change From Baseline Myofibroblasts Formation (Myofibroblasts Content) at 6 Weeks
Actin staining was used to evaluate the presence of myofibroblasts involved in wound healing. These cells increase in number during wound healing. The number of stained cells was semi-quantitatively analyzed using a 0 - 3 scaling score (0= no myofibroblasts, 1= myofibroblasts in low quantity, 2= myofibroblasts in moderate quantity, 3= myofibroblasts in high quantity)
Time frame: At week zero and week 6
| Arm | Measure | Value (MEAN) | Dispersion |
|---|---|---|---|
| Ultrasound Group (UAW Group) | Change From Baseline Myofibroblasts Formation (Myofibroblasts Content) at 6 Weeks | 2.56 score on a scale | Standard Error 0.89 |
| Surgical Group | Change From Baseline Myofibroblasts Formation (Myofibroblasts Content) at 6 Weeks | 2.00 score on a scale | Standard Error 0.1 |
p-value: <0.01Wilcoxon (Mann-Whitney)
Primary
Change From Baseline Neo-angiogenesis (Microvessel Density) at 6 Weeks
Sections of tissue were immunohistochemically-stained with the CD31 marker. Light microscopy was used to count the number of microvessels/endothelial cells in a standardized grid, with the results expressed as microvessel density (Leica DMD 800 morphometric system). Microvessel density was scored according to the following scale: 0 (absent), 1 (low, at least one microvessel), 2 (moderate) and 3 (more than two micro vessels). Higher scores mean a better outcome
Time frame: At week zero and week 6
| Arm | Measure | Value (MEAN) | Dispersion |
|---|---|---|---|
| Ultrasound Group (UAW Group) | Change From Baseline Neo-angiogenesis (Microvessel Density) at 6 Weeks | 2.82 score on a scale | Standard Deviation 0.52 |
| Surgical Group | Change From Baseline Neo-angiogenesis (Microvessel Density) at 6 Weeks | 1.77 score on a scale | Standard Deviation 1.3 |
p-value: <0.01Wilcoxon (Mann-Whitney)
Secondary
Comparison of Quantitative Microbiological Analysis (Bacterial Counts Expressed Colony-forming Units Per Gram of Tissue) (CFU/g)
Tissue samples were weighed and mechanically homogenised in 0.5ml volumes of sterile phosphate buffered saline (PBS, Sigma Aldrich, St Louis, MO). Homogenates were diluted and plated onto Columbia agar (BD, Sparks, MD), Columbia agar supplemented with colistin and nalidixic acid (BD), MacConkey agar (BD), and Sabouraud dextrose agar (BD) using a spiral plater workstation (Don Whitley Scientific, Shipley, UK).The limit of detection was 10 colony-forming units (CFU). Results were expressed as CFU per gram of tissue (CFU/g). Isolated microorganisms were identified by standard criteria and the BBL Crystal identification system (BD). Susceptibility testing of Staphylococcus aureus isolates for oxacillin was performed according to Clinical and Laboratory Standards Institute (CLSI) guidelines, using a 30g cefoxitin disc and Mueller-Hinton agar .
Time frame: At week zero and week 6
| Arm | Measure | Value (MEAN) | Dispersion |
|---|---|---|---|
| Ultrasound Group (UAW Group) | Comparison of Quantitative Microbiological Analysis (Bacterial Counts Expressed Colony-forming Units Per Gram of Tissue) (CFU/g) | 2.11 Log10 CFU/g | Standard Deviation 0.8 |
| Surgical Group | Comparison of Quantitative Microbiological Analysis (Bacterial Counts Expressed Colony-forming Units Per Gram of Tissue) (CFU/g) | 4.39 Log10 CFU/g | Standard Deviation 1.24 |
Comparison: We analyzed 51 patients (24 in surgical group and 27 in UAW group) with a statistical power of 0.80 and an alpha of 0.05, with a power of the clinical di↵erence of 37% to detect a statistically significant between groups.p-value: 0.01t-test, 2 sided
Secondary
Wound Score at 6 Weeks
Wound bed tissue was evaluated for presence, quality, and consistency of granulation tissue using a validated wound scoring system, with scores ranging between a minimum of zero points and maximum of seven points. Higher scores mean a better outcome.
Time frame: Six weeks
| Arm | Measure | Value (MEAN) | Dispersion |
|---|---|---|---|
| Ultrasound Group (UAW Group) | Wound Score at 6 Weeks | 5.4 score on a scale | Standard Deviation 1.5 |
| Surgical Group | Wound Score at 6 Weeks | 5.6 score on a scale | Standard Deviation 0.7 |
p-value: 0.93t-test, 2 sided
Secondary
Wound Size
A planimetric measurements of wound size were conducted using Visitrak (Smith & Nephew, Hull, UK), with the area of the lesion determined with an approximation of ±5mm2
Time frame: 6 weeks
| Arm | Measure | Value (MEAN) | Dispersion |
|---|---|---|---|
| Ultrasound Group (UAW Group) | Wound Size | 1.0 Centimeter square | Standard Deviation 1.22 |
| Surgical Group | Wound Size | 0.88 Centimeter square | Standard Deviation 1.04 |
Comparison: We analyzed 51 patients (24 in surgical group and 27 in UAW group) with a statistical power of 0.80 and an alpha of 0.05, with a power of the clinical difference of 37% to detect a statistically significant between groups.p-value: 0.711t-test, 2 sided