Editorial
Recently, so called “Autologous Biomaterials”, as
naturally occurring substances are considered to have a potential for enhance
the bone and soft tissue healing and help fight infection[1]. From these biomaterials the Platelet-Rich Plasma
(PRP) - a preparation of concentrated autologous platelets containing many
bioactive factorsis widely used in clinical practice, whereas antimicrobial
neutrophil extract (AMP) is taken into account as a new autologous preparation for
enhance of tissue repair. PRP was previously used in animals and humans in
clinical therapies of many disorders in dentistry, neurosurgery,
cardiothoracic, and maxillofacial surgery, in orthopedics and traumatology [2]. Neutrophil-derived antimicrobial peptides, in
turn, were applied in treatment of osteomyelitis. However, to date combination
of both preparations has not been used in clinical practice. We assumed that
both autologous preparations used together could be applied locally in surgical
and orthopedic conditions to enhance healing. Therefore, the research preceding
clinical local use of both these preparations in veterinary practice has been
started. For this purpose, PRP rich in growth factors and with restricted
amount of WBC has been prepared. Next, we evaluated the method for preparation
of AMP neutrophil extract containing natural antimicrobial peptides.Moreover, in order to achieve a better understanding of the
contribution of inflammatory cells in the healing process the influence of AMP extract on the activity of neutrophils and
macrophages in vitrowas also assessed [3].
Cationic antimicrobial peptides are very common in
domesticated animals and vary considerably in their composition, structure and
distribution among species. These peptides, especially their influence on
inflammatory cells and regulation of inflammation as well as repair process in
multiple tissues are currently of great interest. Since antimicrobial peptides
and proteins can be offered as possible replacement of antibiotics which lost
their effectiveness due to microbial resistance, the alternative means are of
great importance. Among antimicrobial peptides cathelicidins are effector
molecules of innate immunity with diverse functions, including antimicrobial
activity, modulation of wound repair and inflammation by influence on
inflammatory cells. Within these cells, macrophages initially contribute to the
elimination of pathogens and the elicitation of an inflammatory reaction. When
the infection is resolved their function may be directed towards resolution of
inflammation and repair process. Macrophages can thus exhibit pro and
anti-inflammatory properties, and as reported previously antimicrobial peptides
i.e. defensins and cathelicidins may participate in this regulation [4].
In the light of fact that the major challenge for many
analyses of physiologic fluids is to identify optimal procedures for sample
preparation,in both cases reliable qualitative method to detection the
components of these bioactive preparations is necessary. In many cases only
very low amounts of biological specimen are available for the experiment.
MALDI-TOF MS is the method that provides rapid determination of molecular
masses and the heterogeneity of small amounts of peptides and proteins. The
limits of detection for MALDI-TOF MS depend on many variables that may change
detection sensitivity. However, for small peptides, under optimal conditions,
limits of detection can extend to 1 nmol/l solution in 1 ml. Another advance of MALDI-TOF MS is that peptides
smaller than 7000 Da are not detected by standardtechniques of 2-dimensional electrophoresis
because they are below the limits of size resolution, and small components may
not be fixed in gel and produce lower staining intensity per mole of peptide.
Most electrophoretic and chromatographic techniques preferentially detect large
molecules,because absorbance or intensity of staining per molar equivalent of
protein generally increases in proportion to size. Consequently, MALDI-TOF MS
has served as a useful tool to identification and characterization of small
peptides as well as to open up peptidomic analysis [5].
There are many protocols to obtain PRP, without one
standardized method. In case of AMP extract, in turn, we described the method
to obtain this preparation. Due to a small volume of the obtained blood the
MALDI TOF analysis is sufficient for preliminary qualitative detection of some
bioactive factors during the standardization of method of preparation of PRP
and AMP extract in many species. The modified method permits direct analysis of
biological samples without previous preparation steps that involve significant
time, expense, and potential losses of selected components. Using MALDI TOF for
qualitative determination of bioactive components it is possible to asses these
factors even if ELISA tests are unreliable because of species differences [6].
Our method permits to evaluate two autologous
biomaterials for topical application; PRP rich in growth factors and AMP
neutrophil extract which comprisesantimicrobial peptides; cathelicidins and
defensins with a broad spectrum of antimicrobial activity. These blood-derived
products can be used to enhance healing and ensure antimicrobial activity
without side effects induced by intact neutrophils. Therefore, the crude
neutrophil extract could be used for improvement of antimicrobial activities of
PRP without the enlargement of the WBC number. The addition of AMP extract into
cultures of neutrophils and macrophages revealed that this extract decreased
superoxide anion in culture of macrophages and inhibited degranulation and
respiratory burst in neutrophils, thus acts by decrease of proinflammatory
effect on studied subpopulations of white blood cells[3].
Detailed study
concerning the process of macrophages activation and its regulation during tissue repair should be useful in the introduction of new therapies based on manipulation ofmacrophages function for enhance healing[7].The first part of these studies concerning the role of antimicrobial
peptides in activation of monocyte-derived macrophages during implantation of
titanium tibial implants has been just finished[8].Response
of activated macrophages to stimulation by AMP extract may have
therapeutic role in enhancement of tissue repair
and remodeling, especially, for
patients with impaired healing and regenerative capacity.The modification of phenotype of macrophages and their products for therapeutic
purpose create new strategy for safe and effectivemethod of improve healing and repair with reduced
risk of complications.
editorial