Cytocompatibility testing of cyclodextrin-functionalized antimicrobial textiles-a comprehensive approach.

Functionalized textiles can be used in wound management to reduce the microbial burden in the wound area, to prevent wound infections, and to avoid cross-contamination between patients. In the present study, a comprehensive in vitro approach to enable the assessment of antibacterial activity of functionalized textiles and cytotoxicity of cyclodextrin (CD)-complexes with chlorhexidine diacetate (CHX), iodine (IOD), and polihexanide (PHMB) is suggested to evaluate their properties for supporting optimal conditions for wound healing. For all ß-CD-antiseptic functionalized cotton samples a strong antibacterial effect on the Gram-positive bacteria Staphylococcus aureus and Staphylococcus epidermidis as well as on the Gram-negative bacteria Klebsiella pneumoniae and Escherichia coli was proven. In addition, ß-CD-CHX and ß-CD-PHMB were effective against the yeast Candida albicans. The growth of Pseudomonas aeruginosa could be reduced significantly by ß-CD-IOD and ß-CD-PHMB. The established comprehensive testing system for determination of biocompatibility on human HaCaT keratinocytes is suitable for obtaining robust data on cell viability, cytotoxicity and mode of cell death of the ß-CD-antiseptic-complexes. The promising results of the high antimicrobial activity of these functionalized textiles show the high potential of such materials in medical applications.

In vitro Evaluation of the Cleansing Effect of a Monofilament Fiber Debridement Pad Compared to Gauze Swabs.

BACKGROUND: Removal of nonvital tissue is an accepted method to eradicate biofilms and to stimulate wound healing. Debridement using a monofilament polyester fiber pad has clinically been shown to be effective as well as pain and trauma free. METHODS: For in vitro determination of the cleansing capacity of this product compared to gauze swabs, a wound debridement model was used with glass plates coated with a bovine serum albumin solution, stained with hematoxylin. Both products were moistened and fixed to a weight connected to a regulated motor and were then pulled over the holding device with the coated glass plate under standardized conditions (power = 0.067 N/cm2, velocity = 1.6 cm/s). RESULTS: At a low coating concentration (0.5%) both products were equally effective, but at a high concentration (1.5%) cleansing did not occur after 5 wipes. When wiping the plates 15 times, the debridement pad cleansed significantly (p < 0.001) better than gauze. When consecutively wiping 4 coated plates with a single debridement pad or swab, the pad exhibited and maintained a significantly higher cleansing capacity while gauze quickly lost its effect. CONCLUSION: Our in vitro test results indicated a higher cleansing capacity of the debridement pad compared to gauze swabs.

Antimicrobial functionalization of bacterial nanocellulose by loading with polihexanide and povidone-iodine.

Bacterial nanocellulose (BNC) is chemically identical with plant cellulose but free of byproducts like lignin, pectin, and hemicelluloses, featuring a unique reticulate network of fine fibers. BNC sheets are mostly obtained by static cultivation. Now, a Horizontal Lift Reactor may provide a cost efficient method for mass production. This is of particular interest as BNC features several properties of an ideal wound dressing although it exhibits no bactericidal activity. Therefore, BNC was functionalized with the antiseptics povidone-iodine (PI) and polihexanide (PHMB). Drug loading and release, mechanical characteristics, biocompatibility, and antimicrobial efficacy were investigated. Antiseptics release was based on diffusion and swelling according to Ritger-Peppas equation. PI-loaded BNC demonstrated a delayed release compared to PHMB due to a high molar drug mass and structural changes induced by PI insertion into BNC that also increased the compressive strength of BNC samples. Biological assays demonstrated high biocompatibility of PI-loaded BNC in human keratinocytes but a distinctly lower antimicrobial activity against Staphylococcus aureus compared to PHMB-loaded BNC. Overall, BNC loaded with PHMB demonstrated a better therapeutic window. Moreover, compressive and tensile strength were not changed by incorporation of PHMB into BNC, and solidity during loading and release could be confirmed.

pH influence on antibacterial efficacy of common antiseptic substances.

BACKGROUND: Wound infection plays an important role in compromised wound healing. A high bioburden impairs healing and leads to formation of a chronic wound. Distinctly higher pH values were observed in chronic wounds compared to acute wounds. However, there is only limited knowledge of pH dependency on the antibacterial efficacy of common antimicrobial substances. METHODS: This study investigated the pH influence on the antimicrobial efficacy of povidone (PVP)-iodine, silver nitrate, chlorhexidine, octenidine and polihexanide against Staphylococcus aureus and Pseudomonas aeruginosa using the agar diffusion test and microplate laser nephelometry. RESULTS: The bactericidal activity of chlorhexidine and octenidine was mainly pH-independent in a pH range of 5.0-9.0. In contrast, polihexanide showed a significant efficacy increase at a higher pH. It was also found that the influence of the pH on antiseptics differs among species of bacteria. For instance, S. aureus exhibited an increasing sensitivity against silver nitrate with rising pH whereas the effect on P. aeruginosa was found to be distinctly decreased. The antimicrobial effect of PVP-iodine was strongly diminished with rising pH. CONCLUSIONS: The shift towards higher pH values in chronic wounds compared to acute wounds makes it imperative to know whether the antimicrobial efficacy of applied antimicrobial substances is altered by different pH levels. The results suggest that application of polihexanide might be advantageous for the management of wound infections, as both S. aureus and P. aeruginosa exhibited an increased susceptibility with rising pH.

Application of a drainage film reduces fibroblast ingrowth into large-pored polyurethane foam during negative-pressure wound therapy in an in vitro model.

Negative-pressure wound therapy (NPWT) is an advantageous treatment option in wound management to promote healing and reduce the risk of complications. NPWT is mainly carried out using open-cell polyurethane (PU) foams that stimulate granulation tissue formation. However, growth of wound bed tissue into foam material, leading to disruption of newly formed tissue upon dressing removal, has been observed. Consequently, it would be of clinical interest to preserve the positive effects of open-cell PU foams while avoiding cellular ingrowth. The study presented analyzed effects of NPWT using large-pored PU foam, fine-pored PU foam, and the combination of large-pored foam with drainage film on human dermal fibroblasts grown in a collagen matrix. The results showed no difference between the dressings in stimulating cellular migration during NPWT. However, when NPWT was applied using a large-pored PU foam, the fibroblasts continued to migrate into the dressing. This led to significant breaches in the cell layers upon removal of the samples after vacuum treatment. In contrast, cell migration stopped at the collagen matrix edge when fine-pored PU foam was used, as well as with the combination of PU foam and drainage film. In conclusion, placing a drainage film between collagen matrix and the large-pored PU foam dressing reduced the ingrowth of cells into the foam significantly. Moreover, positive effects on cellular migration were not affected, and the effect of the foam on tissue surface roughness in vitro was also reduced.

Novel bioactive amino-functionalized cellulose nanofibers.

Amino-cellulose-based nanofibers are prepared by electrospinning of blended solutions of 6-deoxy-6-trisaminoethyl-amino (TEAE) cellulose and polyvinyl alcohol (PVA). The TEAE cellulose with a degree of substitution of 0.67 is synthesized via a nucleophilic displacement reaction starting from cellulose-p-toluenesulfonic acid ester. Several solution characteristics such as polymer concentration, electrical conductivity, and surface tension as well as setup parameters are investigated to optimize the ability of nanofiber formation. These parameters are evaluated using the rheological studies of the solutions. The nanofibers obtained are characterized by scanning electron microscopy (SEM) and show a high antimicrobial activity against Staphylococcus aureus and Klebsiella pneumoniae.

A superabsorbent polymer-containing wound dressing efficiently sequesters MMPs and inhibits collagenase activity in vitro.

Superabsorbent polymer (SAP)-containing wound dressings present a valuable and unique category of wound management products. An in vitro approach was used to assess the effects of a new SAP dressing in treatment of non-healing wounds. It was shown that the SAP dressing possesses a significant binding capacity for MMP-2 and MMP-9 in vitro (P\0.001). The inclusion of the bound proteases was so strong that no MMP-2 and only marginal amounts of MMP-9 were released from the dressing samples in a subsequent elution step. In addition, the SAP dressing was able to take up collagenase and reduce its activity in vitro. However, collagenase was not completely inactivated upon binding and enzyme-mediated substrate turnover could be observed at the dressings. In conclusion, in vitro data confirm the positive effect of the SAP wound dressing observed in vivo. The findings suggest that it should be specifically useful for highly exuding wounds with an elevated proteolytic activity that needs to be reduced to support healing.

Silver-loaded seaweed-based cellulosic fiber improves epidermal skin physiology in atopic dermatitis: safety assessment, mode of action and controlled, randomized single-blinded exploratory in vivo study.

BACKGROUND: The epidermal part of the skin is the major interface between the internal body and the external environment. The skin has a specific physiology and is to different degrees adapted for protection against multiple exogenous stress factors. Clothing is the material with the longest and most intensive contact to human skin. It plays a critical role especially in inflammatory dermatoses or skin conditions with an increased susceptibility of bacterial and fungal infections like atopic dermatitis. Previously, we have shown a dose-dependent antibacterial and antifungal activity of silver-loaded seaweed-based cellulosic fibres. AIM OF THE STUDY: We studied the mode of action of silver-loaded seaweed-based cellulosic fiber and performed a broad safety assessment. The principal aim was to analyse the effects of wearing the textile on epidermal skin physiology in 37 patients with atopic dermatitis in a controlled, randomized single-blinded in vivo study. Furthermore, the sensitization potential was tested in a patch test in 111 panellists. RESULTS: We could demonstrate in vitro a dose-dependent scavenging of induced reactive oxygen species by silver-loaded seaweed-based cellulosic fibers. Safety assessment of these fibres showed no detectable release of silver ions. Furthermore, ex vivo assessment after 24 h application both in healthy volunteers and patients with atopic dermatitis by sequential tape stripping and subsequently raster electron microscopy and energy dispersive microanalysis analysis revealed no detectable amounts of silver in any of stratum corneum layers. Serum analysis of silver showed no detectable levels. The in vivo patch testing of 111 volunteers revealed no sensitization against different SeaCell Active (SeaCell GmbH, Rudolstadt, Germany) containing fabrics. The in vivo study on 37 patients with known atopic dermatitis and mild-to-moderate eczema on their arms were randomly assigned to either silver-loaded seaweed fibre T-shirts or to cotton T-shirts for 8 weeks. A significant reduction in Staphylococcus aureus colonization was detectable for the silver T-shirts compared with cotton T-shirts without any changes in non-pathogenic surface bacteria colonization. Furthermore, a more pronounced improvement in barrier function (transepidermal water loss) was observed in mildly involved eczema areas during the first 4 weeks of the study. Stratum corneum hydration and surface pH improved in both treatment groups over time. CONCLUSION: The tested silver-loaded seaweed fibre can be regarded as safe and seams to be suited for application in bio-active textiles in atopic dermatitis based on its positive in vivo activity.

Comparative in vitro study on cytotoxicity, antimicrobial activity, and binding capacity for pathophysiological factors in chronic wounds of alginate and silver-containing alginate.

Chronic wounds contain elevated levels of proteases, proinflammatory cytokines, and free radicals. The presence of bacteria further exaggerates the tissue-damaging processes. For successful treatment, the wound dressing needs to manage wound exudates, create a moist environment, inhibit infection, bind pathophysiological factors that are detrimental to wound healing, and provide thermal isolation. Furthermore, it has to relieve pain, be easy to use, show no allergic potency, and not release toxic residues. The present study suggests a comprehensive in vitro approach to enable the assessment of wound dressings to support optimal conditions for wound healing. Three alginate-based wound dressings: alginate alone, alginate containing ionic silver, and alginate with nanocrystalline silver, were tested for biocompatibility, antimicrobial activity, and influence on chronic wound parameters such as elastase, matrix metalloproteases-2, tumor necrosis factor-alpha, interleukin-8, and free radical formation. Alginate was found to bind considerable amounts of elastase, reduce the concentration of proinflammatory cytokines and inhibit the formation of free radicals. Furthermore, alginate showed antibacterial activity and high biocompatibility. Incorporation of silver into alginate fibers increased antimicrobial activity and improved the binding affinity for elastase, matrix metalloproteases-2, and the proinflammatory cytokines tested. Addition of silver also enhanced the antioxidant capacity. However, a distinct negative effect of silver-containing alginates on human HaCaT keratinocytes was noted in vitro.

Effect of non-adhering dressings on promotion of fibroblast proliferation and wound healing in vitro.

Non-adhering dressings are commonly used during granulation, tissue formation, and re-epithelialization. Elucidating cytotoxic effects and influence on proliferation/migration capacity of cells like fibroblasts is of interest. Dressings' effects were investigated by comprehensive in vitro approach: (1) MTT assay measuring cell viability after direct contact, (2) ATP assay determining effects on cell proliferation, and (3) scratch wound assay featuring an in vitro wound healing model. One cotton-based dressing with vaseline (vas) was included in the study and four polyester dressings containing vas and technology-lipido-colloid matrix (TLC), carboxymethylcellulose (CMC), hydrocolloid (HC), or glycerin (gly) as additives. A polyamide dressing with vas + CMC and three silicone-based dressings (AT, CC, M) were tested. Polyester + vas + CMC did not negatively affect cell viability or proliferation but it was found that fibroblast layers appeared more irregular with decreased F-actin network structure and tubulin density possibly leading to hampered scratch closure. Silicone AT, polyester + gly and polyamide + vas + CMC caused distinct cell damage. The latter two further reduced cell viability, proliferation and scratch healing. From the overall results, it can be concluded that cotton + vas, polyester + TLC, polyester + vas + HC and the silicone dressings CC and M have the potential to prevent damage of newly formed tissue during dressing changes and positively influence wound healing.

Hemostatic wound dressings: Predicting their effects by in vitro tests.

BACKGROUND: Application of controlled in vitro techniques can be used as a screening tool for the development of new hemostatic agents allowing quantitative assessment of overall hemostatic potential. MATERIALS AND METHODS: Several tests were selected to evaluate the efficacy of cotton gauze, collagen, and oxidized regenerated cellulose for enhancing blood clotting, coagulation, and platelet activation. RESULTS: Visual inspection of dressings after blood contact proved the formation of blood clots. Scanning electron microscopy demonstrated the adsorption of blood cells and plasma proteins. Significantly enhanced blood clot formation was observed for collagen together with ß-thromboglobulin increase and platelet count reduction. Oxidized regenerated cellulose demonstrated slower clotting rates not yielding any thrombin generation; yet, led to significantly increased thrombin-anti-thrombin-III complex levels compared to the other dressings. As hemostyptica ought to function without triggering any adverse events, induction of hemolysis, instigation of inflammatory reactions, and initiation of the innate complement system were also tested. Here, cotton gauze provoked high PMN elastase and elevated SC5b-9 concentrations. CONCLUSIONS: A range of tests for desired and undesired effects of materials need to be combined to gain some degree of predictability of the in vivo situation. Collagen-based dressings demonstrated the highest hemostyptic properties with lowest adverse reactions whereas gauze did not induce high coagulation activation but rather activated leukocytes and complement.

Antifungal and antibacterial properties of a silver-loaded cellulosic fiber.

The skin is the interface between the body and the environment. Each skin type has a specific skin physiology and is more or less adapted for protection against multiple stress factors. Textiles on the other hand are the tissues with the longest contact to the human skin. They play a critical role especially in skin conditions with an increased rate of bacterial and fungal infections like atopic dermatitis and hyperhidrosis, and in diabetic patients and aged skin. The present study demonstrates the antifungal and antibacterial effects of SeaCell Active in an in vitro test system against Candida albicans (DSM 11225), Candida tropicalis (ATCC 1169), and Candida krusei (ATCC 6258). Furthermore, the antibacterial activity of fibers with different amounts of SeaCell Active fibers in a dose-dependent manner against Staphylococcus aureus (ATCC 22923) and Escherichia coli (ATCC 35218) could be demonstrated. If this fiber seems to be suited for bioactive textiles in specific anatomical regions and skin conditions with a susceptibility for fungal and bacterial infections due to Candida species, namely Staphylococcus aureus and Escherichia coli, must be examined by means of further investigations, especially in vivo tests in human, considering allergic and toxic effects of the fiber.

Synthesis of Novel Cellulose Carbamates Possessing Terminal Amino Groups and Their Bioactivity.

Cellulose phenyl carbonates are an excellent platform to synthesize a broad variety of soluble and functional cellulose carbamates. In this study, the synthesis of cellulose carbamates with terminal amino groups, namely ω-aminoethylcellulose- and ω-aminoethyl-p-aminobenzyl-cellulose carbamate, is discussed. The products are well soluble and their structures can be clearly described by NMR spectroscopy. The cellulose carbamates exhibit a bactericide and fungicide activity in vitro. The ω-aminoethylcellulose carbamate possesses a strong activity against Candida albicans and Staphylococcus aureus (IC50 of 0.02 mg mL(-1) and 0.05 mg mL(-1)). The antimicrobial activity and cytotoxicity can be improved by p-amino-benzylamine (ABA) as an additional substituent. The mixed cellulose carbamate exhibits a high biocompatibility (LC50 of 3.18 mg mL(-1)) and forms films on cotton and PES, which exhibit a strong activity against S. aureus and Klebsiella pneumoniae.

Influence of selected wound dressings on PMN elastase in chronic wound fluid and their antioxidative potential in vitro.

Exudates from non-healing wounds contain elevated levels of proteolytic enzymes, like elastase from polymorphonuclear granulocytes (PMN elastase), reactive oxygen species (ROS) and reactive nitrogen species (RNS). The overproduction of proteolytic enzymes leads to reduced concentrations of growth factors and proteinase inhibitors, resulting in an imbalance between degradation and remodelling processes. Thus, the reduction of protein-degrading enzymes and scavenging of ROS and RNS seem to be suitable ways to support the healing process of chronic stagnating wounds. The aim of this study was to test selected wound dressings from different biomaterials (collagen, oxidized regenerated cellulose (ORC) and ORC/collagen mixture), regarding their antioxidative potential in vitro and their influence on the concentration and activity of PMN elastase in chronic wound fluid. Antioxidant capacity of the investigated wound dressing was determined by a pholasin-based chemiluminescent assay. PMN elastase concentration was determined by means of ELISA. Enzyme activities could be measured by a fluorescence assay. As the presented data demonstrates, all tested materials showed antioxidant capacity. In addition, the investigated materials were able to reduce the concentration and activity of PMN elastase. Beside other aspects, such as biocompatibility, biodegradability, fluid absorption and clinical effects (e.g. angiogenesis and microcirculation), the understanding of these properties may help to support the further refinement of wound dressings for improved wound healing.

Nanoparticle Formulation of AEA and BAEA Cellulose Carbamates Increases Biocompatibility and Antimicrobial Activity.

Antimicrobially active polymers with a cationic moiety gain increasing interest in health care providing a biocompatible alternative to commonly used products. They adhere to the polyanionic bacterial surface, disrupt cell membranes and kill bacteria. An efficient path to obtain amino group-containing cellulose is nucleophilic displacement reaction of p-toluenesulfonic acid ester and amines leading to 6-deoxy-6-(ω-aminoalkyl) aminocellulose carbamates. Spherical nanoparticles of the obtained 6-deoxy-6-(2-aminoethyl) amino (AEA) and 6-deoxy-6-{2-bis[N',N'-(2-aminoethyl)]aminoethyl}amino (BAEA) cellulose carbamate with a size range of 80-120 nm were produced. The AEA and BAEA cellulose carbamates exhibited significant antimicrobial activity with moderate cell compatibility. Nanoparticle formulation enhanced biocompatibility.

6-Deoxy-6-aminoethyleneamino cellulose: synthesis and study of hemocompatibility.

Hemocompatibility of aqueous solutions of antimicrobial 6-deoxy-6-aminoethyleneamino (AEA) cellulose with different degrees of substitution (DS, 0.54-0.92) was investigated in vitro. The AEA cellulose derivatives were synthesized by tosylation of cellulose and subsequent nucleophilic substitution with 1,2-diaminoethane. The structure was confirmed by elemental analysis as well as by FTIR and NMR spectroscopies. Markers for coagulation (thrombin generation, aPTT, PT, blood clotting, thrombocyte activation) and membrane integrity (hemolysis) were measured in human whole blood, human platelet-rich plasma, human pooled plasma, and erythrocytes suspension. AEA cellulose with a low DS of 0.54 showed the highest hemocompatibility in vitro, suggesting the possibility of biomedical applications.

Active wound dressings based on bacterial nanocellulose as drug delivery system for octenidine.

Although bacterial nanocellulose (BNC) may serve as an ideal wound dressing, it exhibits no antibacterial properties by itself. Therefore, in the present study BNC was functionalized with the antiseptic drug octenidine. Drug loading and release, mechanical characteristics, biocompatibility, and antimicrobial efficacy were investigated. Octenidine release was based on diffusion and swelling according to the Ritger-Peppas equation and characterized by a time dependent biphasic release profile, with a rapid release in the first 8h, followed by a slower release rate up to 96 h. The comparison between lab-scale and up-scale BNC identified thickness, water content, and the surface area to volume ratio as parameters which have an impact on the control of the release characteristics. Compression and tensile strength remained unchanged upon incorporation of octenidine in BNC. In biological assays, drug-loaded BNC demonstrated high biocompatibility in human keratinocytes and antimicrobial activity against Staphylococcus aureus. In a long-term storage test, the octenidine loaded in BNC was found to be stable, releasable, and biologically active over a period of 6 months without changes. In conclusion, octenidine loaded BNC presents a ready-to-use wound dressing for the treatment of infected wounds that can be stored over 6 months without losing its antibacterial activity.

Poly(ethyleneimines) in dermal applications: biocompatibility and antimicrobial effects.

Cationic polyamines, such as poly(ethyleneimines) (PEIs), may recommend themselves for antimicrobial applications as they can interact with microbial membranes resulting in their disruption. The purpose of the study was the assessment of biocompatibility and antibacterial activity of PEIs with different architectures (branched (b) and linear (l)) and molar masses (0.8-750 kDa). lPEI and bPEI exhibited a strong antibacterial activity against Staphylococcus aureus and Escherichia coli with a more pronounced effect on the Gram-positive bacteria. lPEIs further demonstrated a higher antibacterial efficacy compared to bPEIs but no significant differences between 5 and 25 kDa were observed. In accordance, antibacterial activity of bPEI did not specifically depend on molar mass. Only slightly lower minimal inhibitory concentrations (MIC) were observed at 5 kDa (S. aureus) and 25 kDa (E. coli) in the tests. As PEIs are compelling candidates for use in antimicrobial treatment, two basic aspects have to be investigated: treatment effectiveness and safety. PEIs clearly induced molecular weight dependent cytotoxic effects in vitro. PEIs with low molecular weight (0.8 and 5 kDa) exhibited higher biocompatibility. Nonetheless, the results confirmed a low genotoxic potential of lPEI and bPEIs. In conclusion, 2.5 kDa-lPEI and 0.8 kDa-bPEI can be recommended for use as antimicrobial polymers in dermal applications due to their high biocompatibility with concomitant antibacterial efficacy.

A new silver-loaded cellulosic fiber with antifungal and antibacterial properties.

The skin is the interface between the body and the environment. Each skin type has a specific skin physiology and is more or less adapted for protection against multiple stress factors. Textiles on the other hand are the tissues with the longest contact with the human skin. They play a critical role especially in skin conditions with an increased rate of bacterial and fungal infections like atopic dermatitis or hyperhidrosis, diabetic patients and aged skin. The present study demonstrates the antifungal and antibacterial effect of Sea Cell(R) Active in an in vitro test system against Candida albicans (DSM 11225), Candida tropicalis (ATCC 1169) and Candida krusei (ATCC 6258). Furthermore, the antibacterial activity of fibers with different amounts of Sea Cell Active fibers could be demonstrated in a dose-dependent manner against Staphylococcus aureus (ATCC 22923) and Escherichia coli (ATCC 35218). Whether this fiber seems to be suited for bioactive textiles in specific anatomical body regions and skin conditions with a susceptibility to fungal and bacterial infections, namely Candida species, S. aureus and E. coli, must be examined by means of further investigations, especially human in vivo tests considering allergic and toxic effects of the fiber.