Contribution of Topical Agents such as Hyaluronic Acid and Silver Sulfadiazine to Wound Healing and Management of Bacterial Biofilm.

Background and Objectives: Wound healing is commonly associated with critical bacterial colonization or bacterial infection, which induces prolonged inflammation, resulting in delayed re-epithelialization. An appropriate wound dressing requires a humid environment, which also functions as a barrier against bacterial contamination and will accelerate a regenerative response of the wound. Silver sulfadiazine (SSD) is used to prevent wound infection. Hyaluronic acid (HA) is an extracellular matrix component involved in tissue regeneration. This retrospective study was conducted to evaluate the effectiveness of cream and gauze pads based on hyaluronic acid at low molecular weight (200 kDa) and silver sulfadiazine 1% in the wound healing process. In addition, we examined SSD action on biofilms in vitro and on animal wounds, obtaining positive outcomes therefrom. Materials and Methods: We selected 80 patients with complicated chronic wounds of different etiologies, including diabetes mellitus (10), post-traumatic ulcers (45), burns (15), and superficial abrasion (10). Results: After 8 weeks, ulcer size was decreased in 95 ± 2% of the treated patients; a significant reduction in the inflammatory process was observed from day 14 onwards (p < 0.01 vs. baseline), considering improvement of the surrounding skin and reduction of the bacterial load. The SSD treatment decreased bacterial colony proliferation, both in planktonic state and in biofilm, in a dose-dependent manner on the wound but inhibited the development of tissue granulation at the highest dose (800 µg/wound). Conclusions: In conclusion, the combined action of SSD and HA is clinically effective in improving wound healing.

Intracellular Survival of Biofilm-Forming MRSA OJ-1 by Escaping from the Lysosome and Autophagosome in J774A Cells Cultured in Overdosed Vancomycin.

We investigated the drug-resistant mechanisms of intracellular survival of methicillin-resistant S. aureus (MRSA). Our established MRSA clinical strain, OJ-1, with high biofilm-forming ability, and a macrophage cell line, J774A, were used. After ingestion of OJ-1 by J774A, the cells were incubated for ten days with vancomycin at doses 30 times higher than the minimum inhibitory concentration. The number of phagocytosed intracellular OJ-1 gradually decreased during the study but plateaued after day 7. In J774A cells with intracellular OJ-1, the expression of LysoTracker-positive lysosomes increased until day 5 and then declined from day 7. In contrast, LysoTracker-negative and OJ-1-retaining J774A cells became prominent from day 7, and intracellular OJ-1 also escaped from the autophagosome. Electron microscopy also demonstrated that OJ-1 escaped the phagosomes and was localized in the J774A cytoplasm. At the end of incubation, when vancomycin was withdrawn, OJ-1 started to grow vigorously. The present results indicate that intracellular phagocytosed biofilm-forming MRSA could survive for more than ten days by escaping the lysosomes and autophagosomes in macrophages. Intracellular MRSA may survive in macrophages, and accordingly, they could be resistant to antimicrobial drug treatments. However, the mechanisms their escape from the lysosomes are still unknown. Additional studies will be performed to clarify the lysosome-escaping mechanisms of biofilm-forming MRSA.

A novel mouse wound model for scar tissue formation in abdominal muscle wall.

Hypertrophic scars found on the human body rarely develop in experimental animals, possibly due to their looser skin structure. This makes it difficult to understand the genesis of scar lesions. Therefore, appropriate animal models are urgently needed. In this study, we established a novel experimental model of a scar-forming wound by resecting a small portion of the abdominal muscle wall on the lower center of the abdomen in C57BL/6N mice, which are exposed to contractive forces by the surrounding muscle tissue. As a low-tension control, a back skin excision model was used with a splint fixed onto the excised skin edge, and granulation tissue formed on the muscle fascia supported by the back skeleton. One week after the resection, initial healing reactions, such as fibroblast proliferation, occurred in both models. However, after 21 days, lesions with collagen-rich granulation tissues, which were also accompanied by multiple nodular/spherical-like structures, developed only in the abdominal wall model. These lesions were analogous to scar lesions in humans. Therefore, the animal model developed in this study is unique in that fibrous scar tissues form under physiological conditions without using any artificial factors and is valuable for studying the pathogenesis and preclinical treatment of scar lesions.

Collagen-derived dipeptide Pro-Hyp administration accelerates muscle regenerative healing accompanied by less scarring after wounding on the abdominal wall in mice.

Collagens act as cellular scaffolds in extracellular matrixes, and their breakdown products may also have important biological functions. We hypothesize that collagen dipeptide Pro-Hyp induces favorable healing activities and examined the effects of Pro-Hyp administered via different routes on wound healing using our novel murine model, in which an advanced fibrosis-prone scar lesion was developed in the abdominal muscle wall under the skin. After excising a part of the abdominal wall, a free-drinking experiment was performed using solutions with casein (CS), high molecular weight collagen peptides (HP), and low molecular weight collagen peptides including Pro-Hyp and Hyp-Gly (LP), in addition to water (HO). On day 21 of the study, when compared to the HO and CS groups, muscle regeneration in the LP group was significantly advanced in the granulation tissue, which was associated with a decrease in fibrosis. To clarify the effects of Pro-Hyp, daily intraperitoneal administration of pure Pro-Hyp was performed. Pro-Hyp administration induced many myogenically differentiated cells, including myogenin-positive myoblasts and myoglobin-positive myocytes, to migrate in the granulation tissue, while scar tissue decreased. These results indicated that Pro-Hyp administration accelerates muscle regenerative healing accompanied by less scarring after wounding on the abdominal wall.

Immunology of Acute and Chronic Wound Healing.

Skin wounds greatly affect the global healthcare system, creating a substantial burden on the economy and society. Moreover, the situation is exacerbated by low healing rates, which in fact are overestimated in reports. Cutaneous wounds are generally classified into acute and chronic. The immune response plays an important role during acute wound healing. The activation of immune cells and factors initiate the inflammatory process, facilitate wound cleansing and promote subsequent tissue healing. However, dysregulation of the immune system during the wound healing process leads to persistent inflammation and delayed healing, which ultimately result in chronic wounds. The microenvironment of a chronic wound is characterized by high quantities of pro-inflammatory macrophages, overexpression of inflammatory mediators such as TNF-α and IL-1ß, increased activity of matrix metalloproteinases and abundance of reactive oxygen species. Moreover, chronic wounds are frequently complicated by bacterial biofilms, which perpetuate the inflammatory phase. Continuous inflammation and microbial biofilms make it very difficult for the chronic wounds to heal. In this review, we discuss the role of innate and adaptive immunity in the pathogenesis of acute and chronic wounds. Furthermore, we review the latest immunomodulatory therapeutic strategies, including modifying macrophage phenotype, regulating miRNA expression and targeting pro- and anti-inflammatory factors to improve wound healing.

Collagen-Derived Di-Peptide, Prolylhydroxyproline (Pro-Hyp): A New Low Molecular Weight Growth-Initiating Factor for Specific Fibroblasts Associated With Wound Healing.

Many cells and soluble factors are involved in the wound healing process, which can be divided into inflammatory, proliferative, and remodeling phases. Fibroblasts play a crucial role in wound healing, especially during the proliferative phase, and show heterogeneity depending on lineage, tissue distribution, and extent of differentiation. Fibroblasts from tissue stem cells rather than from healthy tissues infiltrate wounds and proliferate. Some fibroblasts in the wound healing site express the mesenchymal stem cell marker, p75NTR. In the cell culture system, fibroblasts attached to collagen fibrils stop growing, even in the presence of protein growth factors, thus mimicking the quiescent nature of fibroblasts in healthy tissues. Fibroblasts in wound healing sites proliferate and are surrounded by collagen fibrils. These facts indicate presence of new growth-initiating factor for fibroblasts attached to collagen fibrils at the wound healing site, where the collagen-derived peptide, prolyl-hydroxyproline (Pro-Hyp), is generated. Pro-Hyp triggers the growth of p75NTR-positive fibroblasts cultured on collagen gel but not p75NTR-negative fibroblasts. Thus, Pro-Hyp is a low molecular weight growth-initiating factor for specific fibroblasts that is involved in the wound healing process. Pro-Hyp is also supplied to tissues by oral administration of gelatin or collagen hydrolysate. Thus, supplementation of gelatin or collagen hydrolysate has therapeutic potential for chronic wounds. Animal studies and human clinical trials have demonstrated that the ingestion of gelatin or collagen hydrolysate enhances the healing of pressure ulcers in animals and humans and improves delayed wound healing in diabetic animals. Therefore, the low molecular weight fibroblast growth-initiating factor, Pro-Hyp, plays a significant role in wound healing and has therapeutic potential for chronic wounds.

A Novel Autologous Micrografts Technology in Combination with Negative Pressure Wound Therapy (NPWT) for Quick Granulation Tissue Formation in Chronic/Refractory Ulcer.

Negative pressure wound therapy (NPWT) has been commonly used over the years for a wide range of chronic/refractory lesions. Alternatively, autologous micrografting technology is recently becoming a powerful modality for initiating wound healing. The case presented is of a patient with a lower leg ulcer that had responded poorly to NPWT alone for three weeks. Consequently, the patient was put on a combination therapy of NPWT and micrografting. After injection of a dermal tissue micrografts suspension into the entire wound bed, NPWT was performed successively for two weeks, resulting in fresh granulation tissue formation. Thereafter, the autologous skin graft was taken well. This case study indicates that for a chronic/refractory ulcer patient with poor NPWT outcome, combination therapy using micrografting treatment and NPWT could rapidly initiate and enhance granulation tissue formation, creating a favorable bedding for subsequent skin grafting.

The Effects of Silver Sulfadiazine on Methicillin-Resistant Staphylococcus aureus Biofilms.

Methicillin-resistant Staphylococcus aureus (MRSA), the most commonly detected drug-resistant microbe in hospitals, adheres to substrates and forms biofilms that are resistant to immunological responses and antimicrobial drugs. Currently, there is a need to develop alternative approaches for treating infections caused by biofilms to prevent delays in wound healing. Silver has long been used as a disinfectant, which is non-specific and has relatively low cytotoxicity. Silver sulfadiazine (SSD) is a chemical complex clinically used for the prevention of wound infections after injury. However, its effects on biofilms are still unclear. In this study, we aimed to analyze the mechanisms underlying SSD action on biofilms formed by MRSA. The antibacterial effects of SSD were a result of silver ions and not sulfadiazine. Ionized silver from SSD in culture media was lower than that from silver nitrate; however, SSD, rather than silver nitrate, eradicated mature biofilms by bacterial killing. In SSD, sulfadiazine selectively bound to biofilms, and silver ions were then liberated. Consequently, the addition of an ion-chelator reduced the bactericidal effects of SSD on biofilms. These results indicate that SSD is an effective compound for the eradication of biofilms; thus, SSD should be used for the removal of biofilms formed on wounds.

Recent Advances in the Controlled Release of Growth Factors and Cytokines for Improving Cutaneous Wound Healing.

Bioengineered materials are widely utilized due to their biocompatibility and degradability, as well as their moisturizing and antibacterial properties. One field of their application in medicine is to treat wounds by promoting tissue regeneration and improving wound healing. In addition to creating a physical and chemical barrier against primary infection, the mechanical stability of the porous structure of biomaterials provides an extracellular matrix (ECM)-like niche for cells. Growth factors (GFs) and cytokines, which are secreted by the cells, are essential parts of the complex process of tissue regeneration and wound healing. There are several clinically approved GFs for topical administration and direct injections. However, the limited time of bioactivity at the wound site often requires repeated drug administration that increases cost and may cause adverse side effects. The tissue regeneration promoting factors incorporated into the materials have significantly enhanced wound healing in comparison to bolus drug treatment. Biomaterials protect the cargos from protease degradation and provide sustainable drug delivery for an extended period of time. This prolonged drug bioactivity lowered the dosage, eliminated the need for repeated administration, and decreased the potential of undesirable side effects. In the following mini-review, recent advances in the field of single and combinatorial delivery of GFs and cytokines for treating cutaneous wound healing will be discussed.

Acceleration of Skin Wound-Healing Reactions by Autologous Micrograft Tissue Suspension.

Background and objectives: Skin grafting is a method usually used in reconstructive surgery to accelerate skin regeneration. This method results frequently in unexpected scar formations. We previously showed that cutaneous wound-healing in normal mice is accelerated by a micrograft (MG) technique. Presently, clinical trials have been performed utilizing this technology; however, the driving mechanisms behind the beneficial effects of this approach remain unclear. In the present study, we focused on five major tissue reactions in wound-healing, namely, regeneration, migration, granulation, neovascularization and contraction. Methods: Morphometrical analysis was performed using tissue samples from the dorsal wounds of mice. Granulation tissue formation, neovascularization and epithelial healing were examined. Results: The wound area correlated well with granulation sizes and neovascularization densities in the granulation tissue. Vascular distribution analysis in the granulation tissue indicated that neovessels extended and reached the subepidermal area in the MG group but was only halfway developed in the control group. Moreover, epithelialization with regeneration and migration was augmented by MG. Myofibroblast is a known machinery for wound contraction that uses α-smooth muscle actin filaments. Their distribution in the granulation tissue was primarily found beneath the regenerated epithelium and was significantly progressed in the MG group. Conclusions: These findings indicated that MG accelerated a series of wound-healing reactions and could be useful for treating intractable wounds in clinical situations.

Sequential Delivery of Cryogel Released Growth Factors and Cytokines Accelerates Wound Healing and Improves Tissue Regeneration.

Growth factors and cytokines that are secreted by cells play a crucial role in the complex physiological reaction to tissue injury. The ability to spatially and temporally control their actions to maximize regenerative benefits and minimize side effects will help accelerate wound healing and improve tissue regeneration. In this study, the sequential targeted delivery of growth factor/cytokine combinations with regulatory functions on inflammation and tissue regeneration was examined using an internal splint wound healing model. Four examined growth factors and cytokines were effectively incorporated into a novel chitosan-based cryogel, which offered a controlled and sustained release of all factors while maintaining their biological activities. The cryogels incorporated with inflammation modulatory factors (IL-10 and TGF-ß) and with wound healing factors (VEGF and FGF) were placed on the wound surface on day 0 and day 3, respectively, after wound initiation. Although wound area gradually decreased in all groups over time, the area in the cryogel group with growth factor/cytokine combinations was significantly reduced starting on day 7 and reached about 10% on day 10, as compared to 60-65% in the control groups. Sequential delivery of inflammation modulatory and wound healing factors enhanced granulation tissue formation, as well as functional neovascularization, leading to regenerative epithelialization. Collectively, the chitosan-based cryogel can serve as a controlled release system for sequential delivery of several growth factors and cytokines to accelerate tissue repair and regeneration.

Inappropriate direct oral anticoagulant dosing in atrial fibrillation patients is associated with prescriptions for outpatients rather than inpatients: a single-center retrospective cohort study.

BACKGROUND: Inappropriate dosing of direct oral anticoagulants (DOACs) has been associated with clinical safety and efficacy; however, little is known about clinical data associated with an inappropriate DOAC dosing in Japan. In addition, there is no report in which the appropriateness of DOAC dosing between prescription for inpatients and for outpatients was examined. In this study, we aimed to investigate the prevalence and factors associated in the inappropriate dosing of DOACs in patients with atrial fibrillation (AF). METHODS: The retrospective cohort study was conducted at a single Japanese university hospital. Both inpatients and outpatients, who were diagnosed with AF and for whom treatment with either dabigatran, rivaroxaban, apixaban, or edoxaban was initiated between April 1, 2014 and March 31, 2018, were enrolled in the study. Appropriateness of DOAC dosing was assessed according to the manufacturer's labeling recommendations (dose reduction criteria) of each DOAC. Inappropriate reduced dose, namely, underdosing, was defined as prescription of a reduced dose of DOAC despite the patient not meeting the dose reduction criteria. Inappropriate standard dose, namely, overdosing, was defined as prescription of a standard dose of DOAC despite the patient meeting the dose reduction criteria. Inappropriate DOAC dosing was defined as a deviation of the recommended dose (both underdosing and overdosing). RESULTS: A total of 316 patients (dabigatran, 28; rivaroxaban, 107; apixaban, 116; and edoxaban, 65) were included, with a median (interquartile range) age of 75 (66-81) years and 62.3% male. DOACs were prescribed at an appropriate standard dose in 39.2% of patients, an appropriate reduced dose in 36.7%, an inappropriate standard dose in 2.5%, and an inappropriate reduced dose in 19.3%. Multivariate analysis revealed that the inappropriate dosing of DOACs was significantly associated with prescriptions for outpatients (vs. inpatients; odds ratio [OR] 2.87, 95% confidence interval [CI] 1.53-5.62, p < 0.001) and those with higher HAS-BLED scores (OR 1.87, 95% CI 1.42-2.51, p < 0.001). CONCLUSIONS: Our results demonstrated that the inappropriate dosing of DOACs occurred in approximately 20% of AF patients, and was more frequent in outpatients (vs. inpatients) and in those with a higher risk of bleeding. It is recommended that pharmacists play a greater role in assisting in the prescription process to help physicians make better decisions.

Change in the Antimicrobial Resistance Profile of Extended-Spectrum ß-Lactamase-Producing Escherichia coli.

BACKGROUND: This study aimed to investigate the trends and antimicrobial resistance profile of extended-spectrum ß-lactamase-producing Escherichia coli (ESBL-EC) clinical isolates. METHODS: A total of 1,303 E. coli isolates from January 2012 to December 2017 at Fukuoka University Chikushi Hospital, Japan, were analyzed. The rate of resistance to cefmetazole (CMZ), flomoxef (FMOX), imipenem (IPM), meropenem (MEPM), amikacin (AMK), gentamicin (GM), minocycline (MINO), ciprofloxacin (CPFX), and levofloxacin (LVFX) was compared between non-ESBL-producing E. coli (non-ESBL-EC) and ESBL-EC. RESULTS: The proportion of ESBL-EC among all the E. coli isolates was 24.6% (320/1,303), and the proportion remained stable throughout the study period. There was no difference in the rate of resistance to CMZ, FMOX, IPM, MEPM, and AMK between non-ESBL-EC and ESBL-EC; however, the rate of resistance to GM, MINO, CPFX, and LVFX was higher in ESBL-EC than in non-ESBL-EC (17.5% vs. 10.0%, 19.1% vs. 7.7%, 87.5% vs. 24.2%, and 87.5% vs. 23.5%, respectively; P < 0.01). The rate of resistance to CPFX and LVFX in ESBL-EC increased throughout the study course. The rate of E. coli isolates susceptible to all the antibiotics was significantly higher in non-ESBL-EC than in ESBL-EC (68.2% vs. 7.5%; P < 0.01), and this rate decreased significantly from 10.0% in 2012 to 3.8% in 2017 in ESBL-EC (P < 0.01). CONCLUSIONS: Our findings indicate a changing antimicrobial resistance profile of ESBL-EC, particularly to fluoroquinolones. Determination of the prevalence and antimicrobial resistance of ESBL-EC will help physicians in selecting the initial empirical treatment for patients with ESBL-EC infections.

FF-10501 induces caspase-8-mediated apoptotic and endoplasmic reticulum stress-mediated necrotic cell death in hematological malignant cells.

FF-10501 is a novel inhibitor of inosine monophosphate dehydrogenase (IMPDH). Clinical trials of FF-10501 for myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML) are currently being conducted in the United States. Although it has been shown that FF-10501 induces apoptosis in hematological malignant cells, the intracellular mechanisms of this effect have not been characterized. We conducted an in vitro study to elucidate the mechanisms of FF-10501-induced cell death using 12 hematological malignant cell lines derived from myeloid and lymphoid malignancies. FF-10501 suppressed the growth of each cell line in a dose-dependent manner. However, the clinically relevant dose (40 µM) of FF-10501 induced cell death in three cell lines (MOLM-13, OCI-AML3, and MOLT-3). Investigation of the cell death mechanism suggested that FF-10501 induces both apoptotic and necrotic cell death. FF-10501-induced apoptosis was mediated by caspase-8 activation followed by activation of the mitochondrial pathway in MOLM-13 and MOLT-3 cells. FF-10501 induced necrotic cell death via endoplasmic reticulum stress in OCI-AML3 cells. The present study is the first to identify intracellular pathways involved in FF-10501-induced cell death.

Inhibitory effects of polysorbate 80 on MRSA biofilm formed on different substrates including dermal tissue.

Methicillin-resistant Staphylococcus aureus (MRSA) forms biofilms on necrotic tissues and medical devices, and causes persistent infections. Surfactants act on biofilms, but their mode of action is still unknown. If used in the clinic, cytotoxicity in tissues should be minimized. In this study, we investigated the inhibitory effect of four different surfactants on MRSA biofilm formation, and found that a nonionic surfactant, polysorbate 80 (PS80), was the most suitable. The biofilm inhibitory effects resulted from the inhibition of bacterial adhesion to substrates rather than biofilm disruption, and the effective dose was less cytotoxic for 3T3 fibroblasts. However, the effects were substrate-dependent: positive for plastic, silicon, and dermal tissues, but negative for stainless-steel. These results indicate that PS80 is effective for prevention of biofilms formed by MRSA on tissues and foreign bodies. Therefore, PS80 could be used in medical practice as a washing solution for wounds and/or pretreatment of indwelling catheters.

Acceleration Mechanisms of Skin Wound Healing by Autologous Micrograft in Mice.

A micrograft technique, which minces tissue into micro-fragments >50 µm, has been recently developed. However, its pathophysiological mechanisms in wound healing are unclear yet. We thus performed a wound healing study using normal mice. A humanized mouse model of a skin wound with a splint was used. After total skin excision, tissue micro-fragments obtained by the Rigenera protocol were infused onto the wounds. In the cell tracing study, GFP-expressing green mice and SCID mice were used. Collagen stains including Picrosirius red (PSR) and immunohistological stains for α-smooth muscle actin (αSMA), CD31, transforming growth factor-ß1 (TGF-ß1) and neutrophils were evaluated for granulation tissue development. GFP-positive cells remained in granulation tissue seven days after infusion, but vanished after 13 days. Following the infusion of the tissue micrograft solution onto the wound, TGF-ß1 expression was transiently upregulated in granulation tissue in the early phase. Subsequently, αSMA-expressing myofibroblasts increased in number in thickened granulation tissue with acceleration of neovascularization and collagen matrix maturation. On such granulation tissue, regenerative epithelial healing progressed, resulting in wound area reduction. Alternative alteration after the micrograft may have increased αSMA-expressing myofibroblasts in granulation tissue, which may act on collagen accumulation, neovascularization and wound contraction. All of these changes are favorable for epithelial regeneration on wound.

Increased drug resistance of meticillin-resistant Staphylococcus aureus biofilms formed on a mouse dermal chip model.

PURPOSE: Meticillin-resistant Staphylococcus aureus (MRSA) biofilm formation in humans is of serious clinical concern. Previous in vitro studies have been performed with biofilms grown only on inorganic substrates; therefore, we investigated the vancomycin (VCM) resistance of MRSA biofilms grown on skin tissue. METHODOLOGY: We established a novel tissue substrate model, namely MRSA grown on segments of mouse skin tissue (dermal chips, DCs), and compared its resistance capacity against VCM with that of MRSA biofilms grown on plastic chips (PCs).Results/Key findings. For one MRSA isolate, we found that the VCM MIC was identical (1.56 µg ml-1) for planktonic cultures and for biofilms-formed on PCs (PC-BF), although the minimum bactericidal concentration (MBC) increased to 6.25 µg ml-1 in PC-BF. On the contrary, the MIC and MBC for biofilms formed on DCs (DC-BF) significantly increased (25 and 50 µg ml-1, respectively). Furthermore, the minimum biofilm-eradicating concentration was higher for DC-BF (100 µg ml-1) than for PC-BF (25 µg ml-1). Using six MRSA strains, we found that in PC-BF, the c.f.u. number decreased with increasing VCM concentration, whereas in DC-BF, it greatly increased until the MIC was reached, accompanied by the formation of large colonies, thicker bacterial walls and the presence of many mitotic cells. CONCLUSION: Our results indicate that the VCM resistance of MRSA was greater in DC-BF. We conclude that DCs may provide a specific environment for MRSA that enhances bacterial growth under cytotoxic VCM concentrations, and might be useful for the study of skin wound infections and the effects of antimicrobial drugs.

A Novel Skin Splint for Accurately Mapping Dermal Remodeling and Epithelialization During Wound Healing.

The mouse excisional dorsal full-thickness wound model with a silicon splint fixed on the skin has been widely used to mimic human wound healing. However, the method cannot accurately quantify dermal remodeling, since the initial point of epithelialization on the wound surface is unclear. To overcome this limitation, we have developed a novel mouse excisional wound model to assess the degree of epithelial extension and regeneration, using a plastic ring-shaped splint fixed beneath the surrounding epidermal tissue. At the end of the experiment, tissue samples were fixed in formalin, the splint was excised, and paraffin sections were prepared. Splint holes, corresponding to the prior location of the splint, were evident on the tissue cross-sections, and the epidermis above the holes was considered the initial excision site. The epidermal contraction and epithelial regeneration, as independent essential tissue alterations in wound healing, could be distinguishable and quantified. Compared with previous splint models, this method provides an accurate evaluation of epidermal processes in wound healing, and can be a platform to assess the effects of various wound healing factors. J. Cell. Physiol. 232: 1225-1232, 2017. © 2016 Wiley Periodicals, Inc.

Increased biofilm formation ability and accelerated transport of Staphylococcus aureus along a catheter during reciprocal movements.

Staphylococcus spp. is a major cause of device-related infections. However, the mechanisms of deep-tissue infection by staphylococci from the skin surface remain unclear. We performed in vitro experiments to determine how staphylococci are transferred from the surface to the deeper layers of agar along the catheter for different strains of Staphylococcus aureus with respect to bacterial concentrations, catheter movements, and biofilm formation. We found that when 5-mm reciprocal movements of the catheter were repeated every 8h, all catheter samples of S. aureus penetrated the typical distance of 50mm from the skin to the epidural space. The number of reciprocal catheter movements and the depth of bacterial growth were correlated. A greater regression coefficient for different strains implied faster bacterial growth. Enhanced biofilm formation by different strains implied larger regression coefficients. Increased biofilm formation ability may accelerate S. aureus transport along a catheter due to physical movements by patients.