Product classes to combat wound infections

How infections are treated

The normal approach to treating infection is to kill the microbes using broad-spectrum antibiotics and antiseptics.

For internal body regions, e.g. the blood, an antimicrobial approach will be optimal because it will support the body’s efforts to keep these areas sterile and will often be able to resolve a systemic infection in a few days (e.g. Royer et al. 2018).

External body surfaces, however, are in constant contact with the environment and this makes it impossible to keep them sterile. The body has, therefore, chosen a different strategy and instead actively hosts a complex microbiome - i.e. an ecosystem of bacteria, fungi, viruses and mites – such that the pathogens, i.e. disease-causing microbes, will have difficulty getting a foothold and such that no single microbe readily can become dominant and take over control from the immune system. Over 1000 species of bacteria have been found to live on healthy human skin and being found from the upper surface of the skin to the deep dermal layers (Lloyd-Price et al. 2016). Studies have shown that the diversity of the microbiome often is reduced in skin diseases (Loesche et al. 2017; Ring et al. 2017). Therefore, approaches that further reduce this variability may consequently not be optimal.

To treat wound infection by reducing bacterial load – an assumption not backed by evidence

It is generally assumed, that wound infections have to be treated by antimicrobial approaches and a reduction in bacterial load is seen as a positive result. However, the fact that the body actively hosts a microbiome and uses this for its protection of the body means, that “bacterial load”, or the presence of a high number of bacteria, does not indicate the presence of an infection. A non-specific reduction in bacterial load is, consequently, more likely to have a negative impact on wound healing (and healthy skin) as this would disrupt the microbiome and thereby the body’s defences.

In a number of recent reviews (e.g. Dumville et al 2017; Norman et al. 2016, 2017; Storm-Versloot et al. 2010), the efficacy of antibiotics and antiseptics for the treatment of infected surgical wounds, pressure ulcers, venous leg ulcers, diabetic foot ulcers and burns were evaluated, and concluded that neither antibiotics nor antiseptics provide any consistent clinical benefits in these wound types.

These studies therefore show the limitations of antimicrobial approaches and that we, to fight wound infections, need to use a strategy different to killing off the bacteria.

 

MPPT

Antibiotic

Antiseptic

Un-treated

Clinical

3.0 ± 0.9

7.0 ± 1.2

8.0 ± 1.1

 

Preclinical

3.0 ± 0.4

7.0 ± 0.4

 

10.0 ± 0.4

Data from Bilyayeva et al. (2014, 2017)

The studies performed with MPPT lend support to this conclusion. The comparators in the MPPT clinical study were an antibiotic and an antiseptic and they displayed very similar clinical effects, even though there were significant differences between them. The study did not include an untreated control, which could be used to establish a baseline for healing, but a preclinical study, which used the same study design, parameters and comparators, did include an untreated control. The data are compared in the table above and, as can be seen, the effects of MPPT and the antibiotics were identical in the two studies and this would suggest that some level of comparison can be extended to the two remaining groups, i.e. the antiseptic and the untreated control. Accepting this, it can be seen that the antiseptic and the antibiotic groups have wound healing effects not very different from the untreated control group and this is in line with the findings of the mentioned review studies. In contrast, MPPT, which lacks antimicrobial actions, was able to support a much more rapid wound healing process, i.e. a 57% reduction relative to the antibiotic and a 70% reduction relative to the untreated control in time to reach a wound without infection and that had started healing. This represents a substantial improvement and supports the conclusion that antimicrobial approaches are not optimal for wound care.

Product classes

The most commonly used product classes to treat wound infections are:

  • Antibiotics and antifungals
  • Antiseptics
  • Normal honey and Manuka honey
  • DACC (Dialkylcarbomoyl chloride impregnated dressings)

The table below compares these based on:

  • Mode-of-action, i.e. how they obtain their effect.
  • Antimicrobial effects, i.e. whether they kill bacteria and fungi at the levels used in clinical wound care, i.e. the clinical concentration.
  • Cytotoxicity, i.e. whether they kill the body’s own cells at the clinical concentration used.
  • Effects on the microbiome.
  • Comparative effects relative to MPPT (data from clinical study, where available).

The table shows that all approaches, except MPPT, are based on antimicrobial principles, i.e. the killing of the microbes which will cause damage to the microbiome. Müller & Kramer (2008) measured the antimicrobial efficacy of a number of antiseptics on 2 strains of bacteria. They found that the antiseptics, at clinical concentrations, in general reduced bacterial survival by a factor 1,000 or more, e.g. if 10,000 bacteria are present only 10 will survive – this means that the antiseptics profoundly will disrupt the microbiome. Furthermore a number of studies have found that antiseptics and honey are cytotoxic, i.e. they kill the body’s own cells at the concentrations used clinically in wound care.. Below, each of these products will be briefly covered.

 

Mode-of-action

Anti-microbial

Cytotoxic to human and animal cells

Damages microbiome

Preserves and balances microbiome

Days to non-infected wound (mean±SD)*

Antibiotics

Pharmacological

+

-

+

-

7.0 ± 1.2

Antiseptics

Chemical

+

+

+

-

8.0 ± 1.1

Normal honey

H2O2

+

+

+

-

ND

Manuka honey

Chemical

+

+

+

-

ND

DACC

Physio-chemical

+

-

+

-

ND

MPPT

Micro-absorption

-

-

-

+

3.0 ± 0.9

Comparison of antibiotics, antiseptics and MPPT. *Data from Bilyayeva et al. (2017). The number of days to reach a non-infected wound, i.e. free of necrosis, pus and fibrinogenous thickenings, were recorded for MPPT (n=88), a topical antibiotic (gentamicin) (n=90) and an antiseptic (iodine combined with DMSO) (n=88). ND: Not determined.

Antibiotic and antifungal drugs

These drugs have been developed to selectively affect or block processes that occur only in bacteria or fungi, e.g. to prevent normal cell wall synthesis. Antibiotic and antifungal drugs will, therefore, selectively kill bacteria and fungi without affecting human or animal cells. The majority of these drugs have been developed to be “broad-spectrum”, i.e. affecting a wide range of types of bacteria and fungi, which means that they will kill most of the bacteria and fungi that they come into contact with. However, antibiotics and antifungals are usually unable to penetrate biofilm, which means that they cannot reach microbes hiding therein and this will limit their efficacy.

In relation to the microbiome, the antibiotics and antifungals will kill a large number of the microbes that make up the microbiome and will thereby prevent the immune system from being able to control its composition. As the microbes are killed based on whether they are sensitive to the drugs or not and not whether they are needed for the protection of the skin or the wound, the result will be a disrupted, chaotic area which will be open to invasion by pathogenic species or to uncontrolled growth by resident species, i.e. the exact situation the immune system is trying to prevent.

Antibiotics and antifungals have huge benefits in the treatment of internal infections, but they also have some disadvantages:

  • Microbes can develop resistance to the drugs.
  • Some individuals become sensitive to certain drug classes, e.g. penicillin.
  • Prolonged use can cause side-effects, e.g. kidney damage.

Antiseptics

Antiseptics kill bacteria and fungi by a chemical action, i.e. they are essentially a poison to bacteria and often also fungi and viruses. In contrast to antibiotics, they have not been specifically developed to only affect bacteria and fungi, but will also affect human and animal cells. For most antiseptics, their mode-of-action, i.e. how they kill the bacteria, is not understood.

The table below lists the most commonly used antiseptics. From left to right, it shows

  1. Clinical concentration normally used in wound care;
  2. Cytotoxic IC50, i.e. the concentration at which they kill 50% of the cells in cell culture experiments;
  3. Therapeutic Index, i.e. IC50 divided by clinical concentration. This is the safety factor between the concentration with a clinical effect vs. the concentration causing side-effects. Normally, for pharmaceutical drugs the safety factor must be 10 or more. If the index is below zero, it means that the antiseptic acts as a poison to human and animal cells at lower concentrations than the one used in the clinic.
 

Clinical concentration %

IC50*

Therapeutic Index

Products
Chlorhexidine

0.5

0.0087

0.017

Chlorhexidine Gauze Dressing BP

Bactigras

Iodine

0.5

    Iodozyme
Povidone Iodine

0.35

0.475 (0.1*)

1.4 (0.3)

Povitulle

Inadine

Cadexomer Iodine

0.9

0.45*

0.5

Iodosorb

Iodoflex Dressing

Colloidal silver

0.05 – 0.5

0.0538

1.1 – 0.11 Urgosorb Silver
Silver sulfadiazine

1

0.006

0.006

Silvadene
Octenidine

0.05

0.0038

0.08

Octenilin Wound Irrigation Solution

Octenilin Wound Gel

PHMB

0.1 to 0.5

0.0136 (0.002**)

0.14 to 0.027 (0.02)

Prontosan

Suprasorb X+PHMB

Tielle PHMB Dressing

ActivHeal PHMB Foam Adhesive and Non-adhesive

Kerlix AMD Antimicrobial

Excilon AMD IV Sponges

The IC50 values originated from Muller & Kramer (2008) and were based on the effects of the antiseptics on murine fibroblasts. Later studies have shown greater sensitivity of cells to certain antiseptics and these findings have been included in parenthesis. *: Liu et al. 2017; **: Creppy et al. 2014.

 

The table shows that all the antiseptics are toxic to human and animal cells at the concentration used in wound care, which means that all antiseptics in addition to killing bacteria, also kills the cells in the body. It has, for example, been shown that PHMB at a concentration of 0.0002% accumulates internally in cells (Chindera et al. 2016) and that PHMB, over a period of 3 hours (Creppy et al. 2014), will kill almost all cells in concentrations of 0.002% to 0.01%. These concentrations are far below those used clinically in wound care.

The IC50 values in the table above were determined when exposing the cells to the antiseptic for 30 min. However, Yabes et al. (2017) in human cells measured the toxic effects of Manuka Honey and PHMB on keratinocytes, fibroblasts and osteoblasts (key cells types for wound healing) as a function of concentration and time of exposure. The level of cytotoxicity was expressed as the number of surviving cells after exposure to Manuka or PHMB relative to the number that survived when exposed to saline for the same duration of time, i.e. saline was equal to 100%.

They found (see table above) that the cytotoxicity became much more pronounced if the time of exposure was increased. For example, they found, for Manuka honey, that the survival of fibroblasts was 98% when exposing the cells to 40% Manuka honey for 30 min but if the exposure time was increased to 24 hours, the survival rate fell to 50%. For PHMB, at 0.01%, i.e. 10 times below the concentration used clinically, they found that only 18% of the osteoblasts, i.e. the cells involved in bone formation and repair, survived. This picture was seen for both Manuka Honey and PHMB at all concentrations, i.e. if the exposure time was increased the survival rate fell sharply. Most antiseptics are in contact with the wound 24 hours a day for several days, which means that the cytotoxic effects estimated earlier will have been underestimated.

A valid question is whether data from cell cultures are relevant to wound healing in an individual. Carroll et al. (2017) used a porcine (pig) model of wound healing and the goal was to determine whether antiseptics could be used in combination with NPWT (negative pressure wound therapy). NPWT itself has limited effects on infection and the idea was to infuse water containing an antiseptic to reduce the infection while at the same time applying negative pressure to the wound surface to assist the healing. Many places already use saline in combination with NPWT. They measured the thickness of the granulation layer and found that silver, octenidine and PHMB all inhibited the formation of granulation tissue. The results are summarised in their graph below:

Recently, a clinical case emerged where PHMB was used on a wound that was free of infection and that was healing. The patient (see case story) in less than 10 days developed a strong adverse reaction to the gel containing 0.1% PHMB, i.e. the lowest concentration used normally in wound care and cosmetic products. The adverse events included the emergence of holes in exposed bone, the appearance of gas production, which is indicative of an increase in anaerobic bacteria, wound regression, which included damage to granulation tissue, fascia and epithelialising wound edges and a pain syndrome. It was necessary to terminate the use of PHMB.

These studies (in animals and humans) have confirmed that the findings in cell culture studies are relevant to wound healing in an individual and that the cytotoxic effects seen in culture do interfere with healing. A relevant question is, why these effects have not been seen earlier considering that antiseptics have been used for many years. To some extent they have, because antiseptics have minimal effects on wound healing. However, part of the explanation may also be that most studies with antiseptics have been conducted in infected wounds that required treatment to heal. In these cases, the infection will already be inhibiting the formation of new tissue and it will therefore be difficult to determine whether an antiseptic used to fight the infection also interferes with the formation of new tissuel. To see these cytotoxic effects, studies in wounds without infection are required and when these are performed, as shown above, the toxic effects become apparent.

In summary, all antiseptics

  • Are toxic to human and animal cells at the concentrations used clinically;
  • Will damage the microbiome;
  • Have limited clinical efficacy.

Honey and Manuka Honey

Honey has been used for centuries in wound care, but detailed analysis has also found that all honeys are not the same (Islam et al. 2014). Honey provides a number of actions, including the removal of wound exudate by osmosis, but the main focus has been on its antimicrobial effects.

Most honeys contain the enzyme glucose oxidase. This generates hydrogen peroxide, which is antimicrobial, i.e. kills bacteria and fungi. Hydrogen peroxide has been discovered to be used by the body in a number of physiological functions, e.g. for the killing of bacteria by neutrophils (Wittmann et al. 2012) and the body therefore has the necessary enzymes for its degeneration. When the body uses hydrogen peroxide it is applied locally and short-term, in contrast to the way it is used in the clinic, where it is applied to large areas and normally for longer periods of time. Studies have shown that hydrogen peroxide increases in toxicity the longer it is applied (Tatnall et al. 1991) and depending upon dose, it will therefore damage the microbiome and potentially cause cytotoxicity. Data suggest that it, with benefit, can be used short-term, i.e. for some minutes for the cleaning of wounds, but should not be left in contact with the wound.

Manuka Honey, in contrast, obtains its antimicrobial effects from the chemical methylglyoxal (Mavric et al. 2008). Methylglyoxal is a potent protein-glycating agent and an important precursor of advanced glycation end products (AGEs), i.e. it damages proteins and produces toxic compounds. Methylglyoxal is produced in our bodies during cell metabolism, glucose oxidation and lipid peroxidation or by degradation of carbohydrates, but abnormally high levels have been linked to diseases such as diabetes, Parkinson’s Disease, and Alzheimer’s Disease. It has been found that Manuka Honey contains very high levels of methylglyoxal and its presence can be traced back to the specific flowers the bees visit; it is well known that this will affect the properties of the honey (Islam et al. 2014). In relation to Manuka Honey it has been found:

  • The concentration of methylglyoxal in Manuka honeys is up to 100-fold higher than in conventional honeys (Majtan 2011).
  • Methylglyoxal and AGEs play a role in the pathogenesis of impaired diabetic wound healing. Berlanga et al. (2005) showed that methylglyoxal, when given orally to rats, causes microvascular damage and other diabetes-like complications, which are linked to delayed wound healing.
  • Yabes et al. (2017) showed that Manuka honey is toxic to cells involved in wound healing (kertinocytes, fibroblasts and osteoblasts). The data are shown above.
  • De Simone et al. (2017) demonstrated that methylglyoxal is a potent neurotoxin, i.e. it kills nerve cells.
  • Retamal et al. (2016), using primary cultures of human fibroblasts, found that soluble methylglyoxal is highly cytotoxic and induces cell death through apoptosis.
  • Liu et al. (2017) found a higher incidence of Parkinson’s Disease in diabetics and diabetics have high level of methlyglyoxal as a result of type 2 diabetes.
  • Xiu et al. (2017) found that methylglyoxal caused increased dopamine release, i.e. the hallmark of Parkinson’s Disease.
  • Angeloni et al. (2014) reported that methylglyoxal is strictly correlated with an increase of oxidative stress in Alzheimer’s disease and that many studies show that methylglyoxal and methylglyoxal-derived AGEs play a key role in the etiopathogenesis of Alzheimer's disease.
  • There is a lack of studies demonstrating clear wound healing effects of Manuka honey (Jull et al. 2015; Carter et al. 2016).

In conclusion, honey and Manuka honey are similar to the antiseptics with respect to their chemical mode-of-action, cytotoxicity and clinical effects. As an example, Shukrimi et al. (2008) compared the use of normal honey to povidone iodine in treatment of diabetic foot ulcers and found that they had similar effects. This is consistent with both groups being antimicrobials that disrupt the wound microbiome.

 

DACC

DACC (Dialkylcarbamoyl chloride) is a hydrophobic ester, i.e. it repels water. The membrane of all cells, i.e. bacteria, fungi and animals, is made of a lipid bi-layer which also is hydrophobic, i.e. repels water. This lipid bilayer will bind to the DACC ester and if the binding is sufficiently strong, the cell will be bound to the DACC. Due to the distribution of charges of the DACC and the bacterial membrane, the DACC should favour the attachment of bacteria over animal and human cells, although there appears to be some interactions between DACC and human cells as well (Falk and Ivarsson 2012). For use in wound care, the DACC is applied to the surface of a dressing, the dressing is placed on the wound surface, where the bacteria and fungi, if they come into contact with the dressing, will be bound to the DACC layer and, with each dressing change, the bound organisms will be removed. This will reduce the bacterial load in wounds.

Totty et al. (2017) reviewed the clinical data available on DACC and wrote: "We identified 17 studies with a total of 3408 patients which were included in this review. The DACC-coating was suggested to reduce postoperative surgical site infection rates and result in chronic wounds that subjectively looked cleaner and had less bacterial load on microbiological assessments." The findings were, therefore, comparable to other reviews of wound care products based on antimicrobial principles, namely that they may reduce the bacterial load, but that this effect has not been shown to have any substantial effect on wound healing.

 

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