Plasma and Scarring of Diabetic Ulcers
Condition: Chronic Skin Ulcer; Diabetic Foot; Plasma Protein; Transport Protein
MeSH terms: Diabetic Foot; Skin Ulcer; Ulcer
Study type: Interventional

Summary: 
      Diabetic foot is defined by World Health Organization as a syndrome in which the presence of
      neuropathy, ischemia and infection cause tissue damage or ulcers from minor trauma. This
      condition can be controlled in its early stages with conservative treatment, which is
      effective in preventing infections and amputations. However, even with the new knowledge
      acquired and the development of new therapies, the specialist often faces wounds that do not
      improve despite the proper treatment, so therapies have been sought to help the healing of
      these Ulcers. Growing evidence suggests that healing of chronic diabetic foot ulcers depends
      on growth factors and that the therapeutic use of these in wounds has the potential to
      accelerate their healing in conjunction with wound care Conventional. This study evaluates
      the effect of plasma rich in autologous growth factors on healing chronic ulcers of diabetic
      origin.

      This is a randomized clinical trial that evaluates two study groups. Control group in which
      advanced weekly healing will be performed while the post-advanced healing study group will be
      performed the intradermal application of plasma rich in growth factors. These manoeuvres will
      be performed once a week for four weeks and at the end of the study the results in both
      groups will be checked. Both groups will also evaluate, frequency and intensity of pain,
      quality of life, histological changes in ulcers and metabolic evaluation
    

Detailed description: 
      The diabetic foot is a devastating complication of diabetes mellitus that is accompanied by
      chest-evolving ulcers, which often lead to amputation. Annually more than a million people
      lose some limb stake because of this cause. Foot ulceration is the most common complication
      of diabetic patients and is thought to affect up to 15% of them throughout their lives (8,
      9).

      Ulcers that do not progress over a period of approximately 4 weeks should be reassessed.
      Several studies have also shown that chronic ulcers in general and diabetic foot ulcers in
      particular have alterations in fibroblast proliferation, the balance between growth factors,
      metalloproteinase (MMP) and remodeling extracellular matrix (2, 8, 9).

      Changes in matrix metalloproteinase levels, as well as Regulated on Activation , Normal
      T-cell Expressed and Secretated, interleukin and procalcitonin have been correlated with
      wound healing (10). Metalloproteinase and its regulators, matrix metalloproteinase tissue
      inhibitors (TIMPes) are necessary for a wound to be properly healed at an appropriate level,
      at the right location and for a precise period of time. These aspects, it is presumed, are
      not met in chronic wounds such as ulcers of diabetic origin (11).

      Autologous growth factor-rich plasma (PRFC), applied at the wound site releases dozens of
      growth factors, chemokines and cytokines that regulate angiogenesis, epithelialization and
      regeneration (15, 16). They are secreted by the granules and has been found to contribute to
      increase the speed of the natural healing process with a decrease in time (4).

      Platelet-rich plasma is defined as an autologous or allogeneic platelet derivative with a
      concentration of platelets higher than the baseline. Autologous growth factor-rich plasma
      (PRFC) is the fraction of plasma derived from autologous blood, which after being processed
      by centrifugation, has a higher concentration of platelets and higher than baseline growth
      factors (8). There are two obtaining techniques: closed and open. In the case of obtaining by
      "closed technique", the method used must follow the instructions described in the disposable
      kits used in each commercial system. The "open technique" allows, on the other hand, from
      lower blood volumes and simple equipment, to obtain a higher concentration of growth factors
      (25).

      Given its autologous nature, PRFC is a safe product, which by definition lacks the potential
      risk of disease transmission involving the use of donor blood material. With regard to the
      oncogenic potential of PRFC that some authors have suggested, there is no available evidence
      to support it.

      Key factors released include platelet-derived growth factor (PDGF), beta transforming growth
      factor (TGF-b), platelet factor 4 (PF4), interleukin 1 (IL-1), the angiogenic factor derived
      from platelets (PDAF), vascular endothelial growth factor (VEGF), epidermal growth factor
      (EGF), platelet-derived endothelial growth factor (PDEGF), epithelial cell growth factor
      (ECGF), insulin (IGF-1), osteocalcin, osteonectin, fibrinogen, fibronectin and thrombospondin
      (TSP) (11,27). These proteins secreted by activated platelets influence many aspects of
      healing.

      PRFC has been used in different areas of Regenerative Medicine to improve wound healing
      processes with multiple clinical applications in different fields of medicine: General
      Surgery, Traumatology and Sports Medicine, Dentistry and Surgery Plastic and Reconstructive
      Surgery, Dermatology, Neurology and Neurosurgery (28, 8).

      The implementation of new strategies in the treatment of diabetic foot ulcers involving
      growth factors and platelet-rich plasma (15,16,17,20,27,28) has a history of basing it.

      Despite these solid biological bases, the treatment results of skin ulcers with
      platelet-derived growth factors are varied and there are relatively few controlled studies.
      There is confusion about the usefulness of platelet-rich plasma because the studies that have
      evaluated them use a wide range of products (different concentrations of platelets and
      leukocytes, different techniques and frequencies of application, different types of injuries
      to be treated, very heterogeneous and different endpoints) that make it almost impossible to
      compare data and draw conclusions (29, 30).

      There are reviews that evaluate the results that have been had so far on the effects that
      these biological supplements with platelets offer as adjuvants in wound healing and healing.
      A meta-analysis published in 2016, which included 10 randomized clinical trials evaluating
      the effect of PRFC on chronic wound healing. Three of these randomized clinical trials
      involved ulcers of diabetic origin and three studies involved venous leg ulcers. The overall
      analysis did not shed light on the effectiveness of PRFC, but the results indicated that
      autologous PRFC can improve the healing of diabetic ulcers compared to standard care,
      although the value of evidence is low (46).

      This is why we believe it is appropriate to carry out studies that allow to evaluate in a
      clear way and with a simple but complete description the effect that the use of PRFC has on
      the healing of chronic ulcers of diabetic origin of difficult management.

      Research hypothesis. The 4-week intradermal application of Plasma rich in growth factors
      (PRFC) reduces healing time of chronic ulcers of difficult-to-manage diabetic origin, the
      size of the injury, induces changes at the level histological and contributes to reducing
      pain and improving the patient's quality of life compared to the best treatment available.

      Material and methods. Randomized clinical trial conducted in patients with chronic ulcers
      (more than 4 weeks of evolution) of diabetic origin that come to the Wound Clinic of the
      Regional General Hospital of León for evaluation and care.

      The sample size was calculated in 20 patients per group (control group and study group), to
      detect at least a 50% difference in the reduction in the dimensions of the chronic ulcer at
      the end of the manoeuvre, with a statistical significance of 0.05% and a poten 80%. Patients
      will be selected by non-probabilistic sampling for convenience.

      Procedure. Patients will be incorporated into the study through the Wound Clinic and the
      Hematology department of the Regional General Hospital of León where they will be invited to
      participate in it, after explanation of the procedure and those who accept, they will be
      invited will request to sign informed consent. Patients who agree to participate in the study
      will have metabolic and haematological evaluation by the Hematology service of the same
      hospital. Laboratory studies (hematic biometrics, blood chemistry, HbA1C, lipid profile and
      coagulation times) will be performed. Patients will be cited 1 time per week at the Wound
      Clinic and advanced ulcer cure will be performed in both groups consisting of healing with
      physiological and antiseptic solution, biofilm debridement, as well as devitalized tissue,
      occluding with non-absorbent gauze, dried gauze and bandage in both groups. The study group
      will also have the intradermal application of PRFC after healing. The study group will also
      be taken as a biopsy by punch (5mm x 5mm, considering a small part) in the first week (before
      the application of PRFC) and at the end of treatment (1 week after the last application of
      prFC). In patients where a complete ulcer closure is achieved, no final biopsy taken to
      prevent further ulceration from occurring. This in order to determine the effects of Plasma
      rich in growth factors at the histological level on ulcers of diabetic origin. Such
      procedures shall be carried out by the student. During each appointment, prior to the
      completion of the treatment, the measurement of the ulcer in its major and smallest diameters
      will be performed and the area will be calculated through computerized planimetry. This will
      be done by the surgeon general and responsible for the Wound Clinic who will be blinded which
      group each patient belongs to). Each patient will also be evaluated through the Visual Pain
      Analog Scale (EVA) and the SF-12 Lifestyle Questionnaire, each week. Both scales with
      validity in force (49, 50). During the duration of the study, weekly photographic monitoring
      of ulcers will be carried out to assess their evolution and response to treatment in each
      patient. Both the evaluations and the photographic follow-up will be carried out by the
      student.

      Technical of centrifugation and obtaining PRFC. After 8 hours fasting, blood sampling (20 cc)
      will be performed by ulnar artery puncture, which will be collected in sterile tubes with
      sodium citrate at 3.8% as anticoagulant. A sample will be used to perform a
      pre-centrifugation platelet count. The rest of the blood sample will be centrifuged at 3200
      revolutions per minute (rpm) for 15 minutes at room temperature; separation by pipetting of
      the obtained plasma (approximately 3-4 ml) with a variable platelet content, which will be
      centrifuged again for 8 minutes at 1800 rpm, at room temperature. Subsequently by meticulous
      pipetting, the fractions obtained from the plasma shall be separated from above, and
      collected in sterile tubes separated in such a way that the last fraction obtained
      (approximately 1-5 to 2 ml and located in the lower portion of the tube) corresponding to the
      plasma rich in growth factors or PRFC (26). A 0.5 ml sample will be taken from this fraction
      of PRFC to perform a platelet count and determine post-centrifugation platelet performance in
      each patient and application. This value will be recorded to determine the increase in
      platelet concentration in each patient. The remaining fraction (1.5 ml) will be applied
      through intradermal injection immediately upon collection. This procedure will be performed
      in the Laboratory of the General Hospital of León by the same person trained and familiar
      with the procedure throughout the study. Calcium citrate will not be added as a activator as
      it is considered according to the literature that the activation of growth factors takes
      place in soft tissues once PRFC comes into contact with collagen fibers.

      TECHNICAL APPLICATION OF PRFC Once the ulcer has been cured in the manner described above and
      before the application of the gauze, the study group will be applied to 2 thirds of the PRFC
      obtained through intradermal injection in tod or the perimeter of the ulcer with 3cc syringe
      and hypodermic needle. The remaining third will be applied to the ulcer bed by drip
      technique. Subsequently, non-absorbent gauze will be placed on it and finally covered with
      dried gauze and bandage. These will be removed until the next PRFC application (1 week).

      Statistical Analysis. Continuous variables will be reported as means +/- standard deviations
      and categorical ones as frequencies and percentages. We will perform Student T for the
      initial comparison of the study groups and X2 according to the type of variable. To test our
      hypothesis we will perform ANOVA of repeated measurements and post hoc test of Tukey. A
      Multiple Correlation analysis will determine the possible association of other factors that
      influence the healing time of ulcers.
    

Intervention name(s): Application of platelet rich plasma in chronic wounds of diabetic origin
Intervention type(s): Other
Intervention description(s): Application of plasma rich in autologous growht factors after advanced healing
Other intervention name(s): Advanced cure
Allocation: Randomized
Intervention model: Parallel Assignment
Masking: Single (Outcomes Assessor)
Primary purpose: Supportive Care
Keywords: Platelet; Diabetic foot