Occult Pathologic Findings in Reduction Mammaplasty in 5781 Patients-An International Multicenter Study.

Breast cancer is among the most commonly diagnosed cancers in the world, affecting one in eight women in their lifetimes. The disease places a substantial burden on healthcare systems in developed countries and often requires surgical correction. In spite of this, much of the breast cancer pathophysiology remains unknown, allowing for the cancer to develop to later stages prior to detection. Many women undergo reduction mammaplasties (RM) to adjust breast size, with over 500,000 operations being performed annually. Tissue samples from such procedures have drawn interest recently, with studies attempting to garner a better understanding of breast cancer's development. A number of samples have revealed nascent cancer developments that were previously undetected and unexpected. Investigating these so-called "occult" findings of cancer in otherwise healthy patients may provide further insight regarding risk factors and countermeasures. Here, we detail occult findings of cancer in reduction mammaplasty samples provided from a cohort of over 5000 patients from 16 different institutions in Europe. Although the majority of our resected breast tissue specimens were benign, our findings indicate that there is a continued need for histopathological examination. As a result, our study suggests that preoperative imaging should be routinely performed in patients scheduled for RM, especially those with risk factors of breast cancer, to identify and enable a primary oncologic approach.

Electrotransfer parameters as a tool for controlled and targeted gene expression in skin.

Skin is an attractive target for gene electrotransfer. It consists of different cell types that can be transfected, leading to various responses to gene electrotransfer. We demonstrate that these responses could be controlled by selecting the appropriate electrotransfer parameters. Specifically, the application of low or high electric pulses, applied by multi-electrode array, provided the possibility to control the depth of the transfection in the skin, the duration and the level of gene expression, as well as the local or systemic distribution of the transgene. The influence of electric pulse type was first studied using a plasmid encoding a reporter gene (DsRed). Then, plasmids encoding therapeutic genes (IL-12, shRNA against endoglin, shRNA against melanoma cell adhesion molecule) were used, and their effects on wound healing and cutaneous B16F10 melanoma tumors were investigated. The high-voltage pulses resulted in gene expression that was restricted to superficial skin layers and induced a local response. In contrast, the low-voltage electric pulses promoted transfection into the deeper skin layers, resulting in prolonged gene expression and higher transgene production, possibly with systemic distribution. Therefore, in the translation into the clinics, it will be of the utmost importance to adjust the electrotransfer parameters for different therapeutic approaches and specific mode of action of the therapeutic gene.

Transdermal Nicotine Application Attenuates Cardiac Dysfunction after Severe Thermal Injury.

BACKGROUND: Severe burn trauma leads to an immediate and strong inflammatory response inciting cardiac dysfunction that is associated with high morbidity and mortality. The aim of this study was to determine whether transdermal application of nicotine could influence the burn-induced cardiac dysfunction via its known immunomodulatory effects. MATERIAL AND METHODS: A standardized rat burn model was used in 35 male Sprague Dawley rats. The experimental animals were divided into a control group, a burn trauma group, a burn trauma group with additional nicotine treatment, and a sham group with five experimental animals per group. The latter two groups received nicotine administration. Using microtip catheterization, functional parameters of the heart were assessed 12 or 24 hours after infliction of burn trauma. RESULTS: Burn trauma led to significantly decreased blood pressure (BP) values whereas nicotine administration normalized BP. As expected, burn trauma also induced a significant deterioration of myocardial contractility and relaxation parameters. After application of nicotine these adverse effects were attenuated. CONCLUSION: The present study showed that transdermal nicotine administration has normalizing effects on burn-induced myocardial dysfunction parameters. Further research is warranted to gain insight in molecular mechanisms and pathways and to evaluate potential treatment options in humans.

Numerical and structural chromosomal anomalies in undifferentiated pleomorphic sarcoma.

BACKGROUND: Malignant fibrous histiocytoma (MFH) or undifferentiated pleomorphic sarcoma (UPS) is the most common soft-tissue sarcoma of late adult life. Further advances in genetic characterization are warranted. The aim of this study was to search for numerical and structural chromosomal anomalies in UPS. MATERIALS AND METHODS: We investigated five sarcoma-specific chromosomal translocations, five oncogene amplifications as well as the numerical karyotype of 19 UPS samples and one UPS/MFH cell line (U2197) using FISH probes on interphase nuclei. RESULTS: Our results demonstrate that chromosomal translocations involving CHOP, SYT, EWS, FUS and FKHR genes are absent. Furthermore, amplification of ERBB2 (10.5%) and MDM2 (10.5%) was observed whereas the EGFR, C-MYC and N-MYC genes were not amplified. Interestingly, predominant aneuploidies were found in eight chromosomes. CONCLUSION: The data demonstrate rarity of sarcoma-specific chromosomal breaks and oncogene amplifications in UPS, yet polysomic chromosomes appear more characteristically in this condition.

PLGA nanoparticles loaded with host defense peptide LL37 promote wound healing.

Wound treatment remains one of the most prevalent and economically burdensome healthcare issues in the world. Poly (lactic-co-glycolic acid) (PLGA) supplies lactate that accelerates neovascularization and promotes wound healing. LL37 is an endogenous human host defense peptide that modulates wound healing and angiogenesis and fights infection. Hence, we hypothesized that the administration of LL37 encapsulated in PLGA nanoparticles (PLGA-LL37 NP) promotes wound closure due to the sustained release of both LL37 and lactate. In full thickness excisional wounds, the treatment with PLGA-LL37 NP significantly accelerated wound healing compared to PLGA or LL37 administration alone. PLGA-LL37 NP-treated wounds displayed advanced granulation tissue formation by significant higher collagen deposition, re-epithelialized and neovascularized composition. PLGA-LL37 NP improved angiogenesis, significantly up-regulated IL-6 and VEGFa expression, and modulated the inflammatory wound response. In vitro, PLGA-LL37 NP induced enhanced cell migration but had no effect on the metabolism and proliferation of keratinocytes. It displayed antimicrobial activity on Escherichia coli. In conclusion, we developed a biodegradable drug delivery system that accelerated healing processes due to the combined effects of lactate and LL37 released from the nanoparticles.

Evaluation of human amniotic membrane as a wound dressing for split-thickness skin-graft donor sites.

Human amniotic membrane (HAM) has been used as a biomaterial in various surgical procedures and exceeds some qualities of common materials. We evaluated HAM as wound dressing for split-thickness skin-graft (STSG) donor sites in a swine model (Part A) and a clinical trial (Part B). Part A: STSG donor sites in 4 piglets were treated with HAM or a clinically used conventional polyurethane (PU) foil (n = 8 each). Biopsies were taken on days 5, 7, 10, 20, 40, and 60 and investigated immunohistochemically for alpha-smooth muscle actin (αSMA: wound contraction marker), von Willebrand factor (vWF: angiogenesis), Ki-67 (cell proliferation), and laminin (basement membrane integrity). Part B: STSG donor sites in 45 adult patients (16 female/29 male) were treated with HAM covered by PU foam, solely by PU foam, or PU foil/paraffin gauze (n = 15 each). Part A revealed no difference in the rate of wound closure between groups. HAM showed improved esthetic results and inhibitory effects on cicatrization. Angioneogenesis was reduced, and basement membrane formation was accelerated in HAM group. Part B: no difference in re-epithelialization/infection rate was found. HAM caused less ichor exudation and less pruritus. HAM has no relevant advantage over conventional dressings but might be a cost-effective alternative.

Skin electroporation of a plasmid encoding hCAP-18/LL-37 host defense peptide promotes wound healing.

Host defense peptides, in particular LL-37, are emerging as potential therapeutics for promoting wound healing and inhibiting bacterial growth. However, effective delivery of the LL-37 peptide remains limiting. We hypothesized that skin-targeted electroporation of a plasmid encoding hCAP-18/LL-37 would promote the healing of wounds. The plasmid was efficiently delivered to full-thickness skin wounds by electroporation and it induced expression of LL-37 in the epithelium. It significantly accelerated reepithelialization of nondiabetic and diabetic wounds and caused a significant VEGFa and interleukin (IL)-6 induction. IL-6 was involved in LL-37-mediated keratinocyte migration in vitro and IL-6 neutralizing antibodies delivered to mice were able to suppress the wound healing activity of the hCAP-18/LL-37 plasmid. In a hindlimb ischemia model, electroporation of the hCAP-18/LL-37 plasmid increased blood perfusion, reduced muscular atrophy, and upregulated the angiogenic chemokines VEGFa and SDF-1a, and their receptors VEGF-R and CXCR-4. These findings demonstrate that a localized gene therapy with LL-37 is a promising approach for the treatment of wounds.

Human ß-defensin-3 correlates with muscle fibre degeneration in idiopathic inflammatory myopathies.

Sporadic inclusion body myositis (sIBM) and polymyositis (PM) are characterized by muscle inflammation, with sIBM showing additional degenerative alterations. In this study we investigated human beta defensins and associated TLRs to elucidate the role of the innate immune system in idiopathic inflammatory myopathies (IIM), and its association with inflammatory and degenerative alterations. Expression levels of human beta-defensin (HBD)-1, HBD-2, HBD-3 and TLR2, 3, 4, 7 and 9 were analysed by quantitative real-time PCR in skeletal muscle tissue. Localization of HBD-3, collagen 6, dystrophin, CD8-positive T-cells, CD-68-positive macrophages, ß-amyloid, the autophagy marker LC3, and TLR3 were detected by immunofluorescence and co-localization was quantified. HBD-3 and all TLRs except for TLR9 were overexpressed in both IIM with significant overexpression of TLR3 in sIBM. HBD-3 showed characteristic intracellular accumulations near deposits of ß-amyloid, LC3 and TLR3 in sIBM, and was detected in inflammatory infiltrations and macrophages invading necrotic muscle fibres in both IIM. The characteristic intracellular localization of HBD-3 near markers of degeneration and autophagy, and overexpression of endosomal TLR3 in sIBM hint at different pathogenetic mechanisms in sIBM compared with PM. This descriptive study serves as a first approach to the role of the innate immune system in sIBM and PM.

Proteome analysis reveals antiangiogenic environments in chronic wounds of diabetes mellitus type 2 patients.

In contrast to normal healing wounds, chronic wounds commonly show disturbances in proteins regulating wound healing processes, particularly those involved in cell proliferation and protein degradation. Multidimensional protein identification technology MS/MS was conducted to investigate and compare the protein composition of chronic diabetic foot exudates to exudates from split-skin donor sites of burn victims otherwise healthy. Spectral counting revealed 188 proteins differentially expressed (more than twofold and p-value <0.05) in chronic wounds. Most were involved in biological processes including inflammation, angiogenesis, and cell mortality. Increased expression of the inflammatory response stimulating S100 proteins, predominantly S100A8 and S100A9 (almost tenfold), was identified. Matrix metalloproteinases (MMPs) MMP1, MMP2, and MMP8 were identified to be elevated in chronic wounds with significant impact on collagen degradation and tissue destruction. Further, proteins with antiangiogenic properties were found at higher expression levels in chronic wounds. Reduced angiogenesis leads to drastic shortage in nutrition supply and causes increased cell death, demonstrated by Annexin A5 exclusively found in chronic wound exudates. However, excessive nucleic and cytosolic material infers cell death occurring not only by apoptosis but also by necrosis. In conclusion, mass spectrometric investigation of exudates from chronic wounds demonstrated dramatic impairment in wound repair with excessive inflammation, antiangiogenic environment, and accelerated cell death.

Innate immune response after adenoviral gene delivery into skin is mediated by AIM2, NALP3, DAI and mda5.

Methods for human skin gene therapy requires efficient and stable introduction of genes into skin cells. Transient cutaneous gene therapy is an attractive approach in the treatment of skin diseases. The 'Achilles heel' of adenoviral gene therapy is its immunogenicity and many aspects of adenovirus induced cutaneous immune reaction still remain unanswered, particularly the role of keratinocytes. Therefore, human keratinocytes were transfected with adenoviral DNA and cytokine expression was analyzed. Moreover, adenoviral transduction of full-skin was performed ex vivo and in vivo. We observed cytokine induction after cytoplasmatic internalization of adenoviral DNA into epidermal cells. Inhibition of AIM2, NALP3, DAI or mda5 downregulated the cytokine response. Transduction of immunocompetent mice led to a detectable transgene expression for 12 days. Re-application of the vector led to a decrease in intensity and duration of transgene expression limited to 4 days and an increased cytokine expression. In contrast, immunodeficient mice showed a reduced expression of cytokines after DNA internalization. AIM2, NALP3, DAI and mda5 are essential in the induction of an innate immune response towards adenoviral DNA. This immune reaction leads to a decrease in transduction efficiency of the vector after re-application and modulation of these receptor systems stabilizes transgene expression.

Repair of oronasal fistulae by interposition of multilayered amniotic membrane allograft.

BACKGROUND: Oronasal fistulas are a frequent complication after cleft palate surgery. Numerous repair methods have been described, but wound-healing problems occur often. The authors investigated, for the first time, the suitability of multilayered amniotic membrane allograft for fistula repair in a laboratory experiment (part A), a swine model (part B), and an initial patient series (part C). METHODS: In part A, one-, two-, and four-layer porcine and human amniotic membranes (n = 20 each) were fixed in a digital towing device and the force needed for rupture was determined. In part B, iatrogenic oronasal fistulas in 18 piglets were repaired with amniotic membrane allograft, autofetal amniotic membrane, or small intestinal submucosa (n = 6 each). Healing was evaluated by probing and visual inflammation control (no/moderate/strong) on postoperative days 3, 7, 10, and 76. Histological analysis was performed to visualize tissue architecture. In part C, four patients (two women and two men, ages 21 to 51 years) were treated with multilayered amniotic membrane allograft. RESULTS: In part A, forces needed for amniotic membrane rupture increased with additional layers (p < 0.001). Human amniotic membrane was stronger than porcine membrane (p < 0.001). In part B, fistula closure succeeded in all animals treated with amniotic membrane with less inflammation than in the small intestinal submucosa group. One fistula remained persistent in the small intestinal submucosa group. In part C, all fistulas healed completely without inflammation. CONCLUSIONS: Amniotic membrane is an easily available biomaterial and can be used successfully for oronasal fistula repair. The multilayer technique and protective plates should be utilized to prevent membrane ruptures. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, V.

Growth rate of late passage sarcoma cells is independent of epigenetic events but dependent on the amount of chromosomal aberrations.

Soft tissue sarcomas (STS) are characterized by co-participation of several epigenetic and genetic events during tumorigenesis. Having bypassed cellular senescence barriers during oncogenic transformation, the factors further affecting growth rate of STS cells remain poorly understood. Therefore, we investigated the role of gene silencing (DNA promoter methylation of LINE-1, PTEN), genetic aberrations (karyotype, KRAS and BRAF mutations) as well as their contribution to the proliferation rate and migratory potential that underlies "initial" and "final" passage sarcoma cells. Three different cell lines were used, SW982 (synovial sarcoma), U2197 (malignant fibrous histiocytoma (MFH)) and HT1080 (fibrosarcoma). Increased proliferative potential of final passage STS cells was not associated with significant differences in methylation (LINE-1, PTEN) and mutation status (KRAS, BRAF), but it was dependent on the amount of chromosomal aberrations. Collectively, our data demonstrate that these fairly differentiated/advanced cancer cell lines have still the potential to gain an additional spontaneous growth benefit without external influences and that maintenance of increased proliferative potential towards longevity of STS cells (having crossed senescence barriers) may be independent of overt epigenetic alterations.

The effectiveness of pedicled groin flaps in the treatment of hand defects: results of 49 patients.

PURPOSE: Despite the growing number of free and local flaps used for repairing defects of the hand, groin flaps are also still widely used. The aims of this study were to evaluate the outcome of a large series of patients whose defects were covered by pedicled groin flaps, and to find out whether it is still indicated in replacing damaged soft tissue of the hand in the era of microsurgery. METHODS: From 1982 to 2009, we treated 85 patients with soft tissue defects on the hand and distal forearm with pedicled groin flaps in our department and recorded them in a prospective database. We interviewed and examined 49 patients in this cohort. RESULTS: The mean age of the 85 patients was 33 years, the male/female ratio was 4:1, the mean hospital stay was 29 ± 13 days, and the mean follow-up was 9 years. The duration to flap division was 24 ± 5 days. Altogether, we performed a mean of 4.6 operations per patient, including thinning of the flap, deepening of the interdigital fold, and stump and flap revisions. One flap loss occurred. Of the 49 patients, results were mostly classified as good, and 82% of patients would undergo the procedure again. The mean Disabilities of the Arm, Shoulder, and Hand score value was 23 ± 17. The Vancouver Scar Scale showed nearly normal height and vascularity of the groin flap (0.2 ± 0.4 and 0.3 ± 0.6, respectively), pigmentation was slightly abnormal (0.8 ± 0.6), and pliability was evaluated between "supple" and "yielding" (1.5 ± 1.2). CONCLUSIONS: Results achieved with the groin flaps were positive. Most patients were satisfied with the results, and the operation was easily performed when McGregor's recommendations were followed. Nevertheless, considering the high number of secondary operations, the long hospital stay, and immobilization of the arm, groin flaps should be used only when free flaps or regional pedicle flaps are either not feasible or not indicated. TYPE OF STUDY/LEVEL OF EVIDENCE: Therapeutic III.

Innate defense regulator peptide 1018 in wound healing and wound infection.

Innate defense regulators (IDRs) are synthetic immunomodulatory versions of natural host defense peptides (HDP). IDRs mediate protection against bacterial challenge in the absence of direct antimicrobial activity, representing a novel approach to anti-infective and anti-inflammatory therapy. Previously, we reported that IDR-1018 selectively induced chemokine responses and suppressed pro-inflammatory responses. As there has been an increasing appreciation for the ability of HDPs to modulate complex immune processes, including wound healing, we characterized the wound healing activities of IDR-1018 in vitro. Further, we investigated the efficacy of IDR-1018 in diabetic and non-diabetic wound healing models. In all experiments, IDR-1018 was compared to the human HDP LL-37 and HDP-derived wound healing peptide HB-107. IDR-1018 was significantly less cytotoxic in vitro as compared to either LL-37 or HB-107. Furthermore, administration of IDR-1018 resulted in a dose-dependent increase in fibroblast cellular respiration. In vivo, IDR-1018 demonstrated significantly accelerated wound healing in S. aureus infected porcine and non-diabetic but not in diabetic murine wounds. However, no significant differences in bacterial colonization were observed. Our investigation demonstrates that in addition to previously reported immunomodulatory activities IDR-1018 promotes wound healing independent of direct antibacterial activity. Interestingly, these effects were not observed in diabetic wounds. It is anticipated that the wound healing activities of IDR-1018 can be attributed to modulation of host immune pathways that are suppressed in diabetic wounds and provide further evidence of the multiple immunomodulatory activities of IDR-1018.

Experimental murine model of primary high grade undifferentiated pleomorphic sarcoma not otherwise specified.

BACKGROUND: Undifferentiated pleomorphic sarcoma not otherwise specified (NOS) is a malignant neoplasm of uncertain origin arising both in the soft tissue and the bone. The WHO classified this tumour in 2002 but controversy has plagued this entity due to limited availability of tissue for study. The aim of this study was to establish a reproducible xenograft model of primary human undifferentiated pleomorphic sarcoma NOS. MATERIALS AND METHODS: Primary human sarcoma samples were divided into tumour fragments and transplanted subcutaneously in mice. Sarcoma xenografts were analysed histolomorphologically (light/electron-microscopy; immunohistochemistry). RESULTS: All tumours resulted in viable sarcoma NOS xenografts demonstrating similar histological patterns. In both the original tumours and the xenografts, tumour necrosis was found ranging from 15% to 25%. The background stroma of the xenografts was hyalinised like the primary sarcoma. Electron microscopical analyses showed good maintenance of ultrastructure. CONCLUSION: Implantation of intact tumor fragments yielded in a complete tumor take rate. The development of new cancer therapeutics requires animal models that closely resemble the human patient. This study provides ideal animal models for the research of pathogenesis and pathobiology of primary human undifferentiated pleomorphic sarcoma NOS.

Bacterial burden in the operating room: impact of airflow systems.

BACKGROUND: Wound infections present one of the most prevalent and frequent complications associated with surgical procedures. This study analyzes the impact of currently used ventilation systems in the operating room to reduce bacterial contamination during surgical procedures. METHODS: Four ventilation systems (window-based ventilation, supported air nozzle canopy, low-turbulence displacement airflow, and low-turbulence displacement airflow with flow stabilizer) were analyzed. Two hundred seventy-seven surgical procedures in 6 operating rooms of 5 different hospitals were analyzed for this study. RESULTS: Window-based ventilation showed the highest intraoperative contamination (13.3 colony-forming units [CFU]/h) followed by supported air nozzle canopy (6.4 CFU/h; P = .001 vs window-based ventilation) and low-turbulence displacement airflow (3.4 and 0.8 CFU/h; P < .001 vs window-based ventilation and supported air nozzle canopy). The highest protection was provided by the low-turbulence displacement airflow with flow stabilizer (0.7 CFU/h), which showed a highly significant difference compared with the best supported air nozzle canopy theatre (3.9 CFU/h; P < .001). Furthermore, this system showed no increase of contamination in prolonged durations of surgical procedures. CONCLUSION: This study shows that intraoperative contamination can be significantly reduced by the use of adequate ventilation systems.

A novel xenograft model with intrinsic vascularisation for growing undifferentiated pleomorphic sarcoma NOS in mice.

PURPOSE: Preclinical development of antisarcoma therapy is primarily based on the subcutaneous transplantation of sarcoma xenografts. Tumour cell survival remains a hurdle of current models, which has been attributed to the hypoxic conditions following transplantation. We hypothesised that sarcoma models with an intrinsic tissue-engineered vascular supply are easily reproducible. The aim of this study was to establish a novel vascularised xenograft model. MATERIALS AND METHODS: Primary human soft tissue sarcomas were transplanted into a silicon chamber and placed around the superficial epigastric vessels of nude mice. Sarcoma xenograft samples were assessed histomorphologically. RESULTS: All sarcoma xenografts engrafted, leading to solid tumours. Histological, immunohistochemical staining and light/electron microscopy confirmed the xenografts as identical high-grade pleomorphic sarcomas (NOS) compared with the original patients' tumours. CONCLUSION: This novel sarcoma xenograft model with an intrinsic vascular supply could be of high value for studying human soft tissue sarcomas and their therapy.

Genotoxic and cytotoxic activity of host defense peptides against human soft tissue sarcoma in an in vitro model.

Soft tissue sarcomas (STSs) are a heterogeneous group of rare, mesenchymal tumors. Treatment with common chemotherapeutic drugs is consistently associated with low response rates and high rates of adverse toxic effects. Host defense peptides (HDPs) are used as part of innate immunity, and many of them act by directly lysing the target cell membrane. Studies have demonstrated high selectivity of HDP analogs against malignant cells because of a relative abundance of negative charges in malignant cell membranes, compared to normal cells. Our aim was to assess the toxic efficacy of [D]-K(6)L(9), [D]-K(3)H(3)L(9), and Protegrin-1 against the fibrosarcoma cell line, HT1080, and primary human fibroblasts to analyze the potential of these peptides as therapeutic options against STSs. Cell proliferation of the fibrosarcoma cell line, HT1080, and human fibroblasts was determined in vitro after treatment with [D]-K(6)L(9), [D]-K(3)H(3)L(9), and Protegrin-1. Genotoxicity was examined on the basis of the mild alkali version of single-cell gel electrophoresis (comet assay). Doxorubicin, a commonly used STS chemotherapeutic agent, served as the control. The native HDP, Protegrin-1, could show a cytotoxic tendency against malignant cells, but no selectivity in genotoxic trials. The synthetic peptide, [D]-K(6)L(9), could not show any selective oncolytic activity against sarcoma cells. [D]-K(3)H(3)L(9) has shown a tendency for toxic selectivity against malignant cells. There is a potential of developing suitable oncolytic candidates with selectivity against malignant cells. [D]-K(3)H(3)L(9) showed the first promising results, but there has to be further investigation to improve the therapeutic properties of HDPs.