[Reliability of the FFP classification in pelvic fractures]. / Anwendungsverlässlichkeit der FFP-Klassifikation bei Beckenfrakturen.

BACKGROUND: Pelvic fractures are typical for frail geriatric patients. They are characterized by increasing pain and loss of mobility. As geriatric pelvic fractures differ from the typical high velocity injuries, Rommens and Hofmann recommended a new classification for fragility fractures of the pelvis (FFP) in 2013. In addition to the location of the fracture they also assessed the degree of dislocation. OBJECTIVE: Compared to known fracture classifications of the pelvis, the FFP classification appears complex. Therefore, this study was designed to investigate the interobserver reliability of the FFP classification. MATERIAL AND METHODS: The members of the Section of Geriatric Traumatology (DGOU) were presented with 10 DICOM data sets with fractures of the pelvis for classification. As a reference the classification of P.M. Rommens, the author of the FFP, was established. RESULTS: In this study 24 consultants (47%) and 27 (53%) residents took part. Also, six radiologists were invited to participate. A total of 493 assessments were made. In 184 (37%) cases there was agreement with the reference, in 183 (37%) computed tomography images the fractures were classified lower, in 26 (26%) higher than the reference. This initially resulted in a Cohens κ coefficient of 0.36. This corresponds to a sufficient agreement (fair) according to Garbuz. With the reduction of the classification to four main groups, the Cohens κ coefficient increased to a satisfactory value. CONCLUSION: The reasons for the partly low agreement could be the participants' lack of experience and an inaccurate description of the classification levels. In the end, however, the FFP classification is the only one that meaningfully represents pelvic fractures of geriatric and frail patients. By simplifying to the four main groups, a better interobserver reliability is achieved. For a successful treatment, however, attention to the individual patient and the "fracture personality" is essential.

Appendix stump closure with titanium clips in laparoscopic appendectomy.

PURPOSE: Different techniques, including clips, have been used to close the stump in laparoscopic appendectomy. The aim was to investigate the results after application of a newly developed titanium clip for this operation. METHODS: From June 2008 to February 2010, 104 patients from two different hospitals undergoing laparoscopic appendectomy were included in this prospective study. Closure of the appendix base was generally intended with a titanium double-shanked clip (DS-Clip). The variables of interest were intra-and postoperative complications, operation time and hospital stay. Furthermore, an evaluation of the clip's practicability by the surgeon was performed using a standardised questionnaire. RESULTS: In 104 patients screened intraoperatively, four patients had to be excluded as the operating surgeon felt that the clip was not adequate for closing the stump, generally because of severe inflammation of the base of the appendix with involvement of the caecum. One patient developed an intra-abdominal abscess which had to be drained interventionally; no reoperations were necessary. The overall complication rate, the operation time and the hospital stay were well comparable with other devices for appendix stump closure such as staplers, loops or polymeric clips. The practicability of the clip was mainly rated as excellent or good by the operating surgeons. CONCLUSION: This study suggests that the presented titanium DS-Clip is a safe and cost-effective technique for securing the appendix base in laparoscopic appendectomy. The application is easy and can be learned quickly, making it a good option also for teaching hospitals.

An international study of caregiver-reported burden and quality of life in metachromatic leukodystrophy.

BACKGROUND: Metachromatic leukodystrophy (MLD) is an autosomal recessive lysosomal disorder caused by mutations in the arylsulfatase A gene. Until now, there has been little information on the burden of MLD on patients and their caregivers. This multinational study aims to quantify caregiver-related impacts of MLD across several key domains including symptoms, treatment burden, time investment, social and emotional well-being, and professional and financial impact. RESULTS: Data were collected through moderator-assisted web survey and telephone interviews. The survey was developed with extensive input from clinical experts and MLD patient advocacy groups. The EQ-5D-5L questionnaire was administered during follow-up interviews. The total sample consisted of parents of MLD patients in the US (n = 10), France (n = 10), Germany (n = 6), UK (n = 5), Belgium (n = 1), and Norway (n = 2). The impact of MLD is evident from the EQ-5D-5L scores, which indicate utility values for caregivers below respective national population norms and a higher proportion of caregivers reporting problems with anxiety/depression. Time involved for care was demonstrated by a mean of 4.1 inpatient and 29.6 outpatient hospital visits in the previous 12-month period. These commitments place stress on familial relationships with 50% of caregivers reporting their child's MLD diagnosis had negatively impacted their relationship with their spouse/partner. Professionally, 76.5% of caregivers stopped working or switched to part-time employment following their child's MLD diagnosis, and most acknowledged caring for their child had affected their potential for career progression or promotion. Differences are also observed based on late infantile versus juvenile onset MLD, time since diagnosis, and for transplanted patients versus those who received palliative care only. CONCLUSIONS: This multinational study demonstrates that MLD consistently negatively affects many aspects of caregivers' lives including health, relationships, and professional status, irrespective of location. We expect that the results of this study are generalizable to other countries. This study enhances our understanding of MLD caregiver impacts, which could improve patient care and assist in identifying support for individuals with MLD and their families.

The mouse S100A15 ortholog parallels genomic organization, structure, gene expression, and protein-processing pattern of the human S100A7/A15 subfamily during epidermal maturation.

The calcium-binding proteins of the human S100A7/A15 (hS100A7/A15) subfamily are differentially expressed in normal and pathological epidermis. The hS100A7 (psoriasin) and S100A15 reside in a chromosomal cluster of highly similar paralogs. To exploit the power of mouse models for determining functions of gene products, the corresponding S100A7/A15 ortholog was cloned and examined in murine skin. The single mouse S100A15 (mS100A15) gene encodes a protein of 104 amino acids with a predicted molecular weight of 12,870 Da and two EF-hand calcium binding sites. Using gene-specific primers and specific antibodies, expression of mS100A15 in both skin and isolated keratinocytes is confined to differentiating granular and cornified epidermal cells. Immunoblotting of epidermal extracts revealed a series of high molecular weight bands that are also recognized by an antibody for transglutaminase-mediated protein crosslinks. mS100A15 expression is upregulated in cultured keratinocytes induced to differentiate by calcium or phorbol esters. Maximal induction occurs concordantly with expression of late differentiation markers. Induction is enhanced in keratinocytes overexpressing protein kinase Calpha and is dependent on activator protein-1 transcription factors. The regulation, expression pattern and crosslinking of mS100A15 are consistent with the characteristics of the human orthologs, providing a valid surrogate model to study changes in these proteins associated with cutaneous pathologies.

Expression and tissue distribution of mouse Hax1.

HAX1 is an ubiquitously expressed human gene. Though a number of cellular and viral proteins are known to interact with HAX1, its function is still not completely understood. On the basis of these identified interaction partners, HAX1 seems to play a role in apoptosis and the organization of the cytoskeleton. The cDNAs for human and mouse Hax1 share 86% identity and 80% identity at the protein level, suggesting a similar functional importance. To date, no conclusive data on the tissue specific expression of the murine Hax1 are available and only one interaction partner has been identified. Here, we show a detailed expression analysis for the murine ortholog by RT-PCR, Northern and Western blot. Furthermore, the distribution of Hax1 within different mouse tissues was studied by immunohistochemistry (IHC). In general, we found a good correlation between the results obtained from different detection techniques. Similar to its human counterpart, mouse Hax1 seems to be ubiquitously expressed. At the RNA level, we found the highest expression of Hax1 in liver, kidney and testis. In sharp contrast to the human HAX1 which is highly expressed in skeletal muscle, the mouse ortholog was detected only at very low levels. Using a specific antibody, we detected Hax1 in the majority of mouse tissues by IHC. Interestingly, the most prominent expression of Hax1 was found in epithelial, endothelial and muscle cells. Surprisingly, thymus, spleen and pancreas did not show detectable immunostaining. Furthermore, we have studied the subcellular localisation of Hax1 in a keratinocyte and a neuronal cell line by immunofluorescence. We found Hax1 to be localised mainly in the cytoplasm and detected a partial colocalisation with mitochondria. The results presented here summarize for the first time the expression of the murine Hax1 in different tissues and two cell lines. Further studies will elucidate the functional importance of this protein in individual cell types with respect to structural aspects, cell mobility and apoptosis.

Arginase-1 overexpression induces cationic amino acid transporter-1 in psoriasis.

Regulated uptake of extracellular l-arginine by cationic amino acid transporters (CATs) is required for inducible nitric oxide synthase and arginase activity. Both enzymes were recently recognized as important in the pathophysiology of psoriasis because of their contribution to epidermal hyperproliferation. We here characterize the expression pattern of CATs in psoriatic skin compared to healthy skin. CAT-1 mRNA expression was strongly upregulated in lesional and nonlesional areas of psoriatic skin compared to healthy skin, whereas expression of CAT-2A and the inducible isoform CAT-2B was unaltered in psoriatic skin. Furthermore, we tested the hypothesis that arginase-1 overexpression regulates CAT expression via intracellular l-arginine concentration. In in vitro experiments with arginase-1 overexpressing HaCaT cells, CAT-1 mRNA expression was increased. Likewise, this occurs in l-arginine-starved HaCaT cells. Both CAT-2 isoforms were not affected. Arginase-1 overexpression limits the synthesis of NO at physiological, but not supraphysiological, l-arginine levels. Plasma l-arginine concentration was diminished in psoriasis patients and the arginase product l-ornithine was significantly increased compared to healthy controls. In summary, arginase-1 overexpression leads to upregulated CAT-1 expression in psoriatic skin, which is due to lowered intracellular l-arginine levels and limits NO synthesis at physiological l-arginine concentrations.

Molecular cloning and characterization of alternatively spliced mRNA isoforms from psoriatic skin encoding a novel member of the S100 family.

In an effort to identify psoriasis-associated genes, we compared gene expression in normal and psoriatic skin, using differential display RT-PCR technique. Sequence analysis of a 650-bp cDNA fragment (clone 110) that was highly up-regulated in lesional skin revealed homology to a noncoding cDNA (NICE-2). By subsequent cDNA cloning, using RNA from psoriatic skin, we have identified two alternatively spliced mRNA-isoforms (0.5 and 4.4 kb), which differ in composition of their untranslated regions. By sequence comparison, we have mapped the novel gene, named S100A15, to the S100 gene cluster within the epidermal differentiation complex (chromosome 1q21). Analysis of the deduced amino acid sequence revealed a protein of 101 amino acids containing two potential EF-hand motifs with high homology to the S100A7. Northern blot hybridization and semiquantitative RT-PCR analysis confirmed the S100A15 overexpression in psoriasis, showing different levels of expression of the S100A15 mRNA isoforms. In situ hybridization of the S100A15 revealed a markedly increased staining of basal and suprabasal epidermal layers of psoriatic skin compared with healthy tissue. Our data suggest an involvement of the novel S100A15 in epidermal differentiation and inflammation and might therefore be important for the pathogenesis of psoriasis and other diseases.

HAX-1, identified by differential display reverse transcription polymerase chain reaction, is overexpressed in lesional psoriasis.

Psoriasis is a chronic inflammatory disease characterized by epidermal hyperplasia and an inflammatory infiltrate. The normal differentiation from basal to granular keratinocytes with subsequent apoptosis and cornification is disturbed in the akanthotic epidermis. This could be due to both an excess of mitogenic stimuli with hyperproliferation and/or resistance to apoptosis. By mRNA differential display we found HAX-1 to be overexpressed in lesional psoriatic skin. The overexpression of HAX-1 was verified at the mRNA level by Northern blot and in situ hybridization, as well as at the protein level by Western blot and immunohistochemistry. Detection of HAX-1 in mRNA from different tissues showed strong expression in the brain, pancreas, skeletal muscle, and heart. In contrast to primary keratinocytes and melanocytes we found HAX-1 highly expressed in human immortalized keratinocytes (HaCaT) and different melanoma cell lines. In HaCaT cells as a model for psoriatic keratinocytes we found an increased ultraviolet-induced apoptosis after expression of HAX-1 antisense mRNA. In psoriasis, the epidermal differentiation could be disturbed due to the increased expression of HAX-1 and hence a prolonged resistance to terminal differentiation. Antiapoptotic mechanisms are an emerging concept for the understanding of the pathogenesis of this disease possibly leading to clinical applications.

Human S100A15 splice variants are differentially expressed in inflammatory skin diseases and regulated through Th1 cytokines and calcium.

The human calcium-binding protein (hS100A15) was first identified in inflamed hyperplastic psoriatic skin, where the S100A15 gene is transcribed into two mRNA splice variants, hS100A15-S and hS100A15-L. To compare the contribution of the human S100A15 (hS100A15) isoforms in skin inflammation and differentiation, we determined the expression, distribution and regulation of hS100A15-S and hS100A15-L in psoriasis and chronic atopic eczema compared with normal skin. We found that both hS100A15 transcripts were mainly distributed in the epidermis of normal and inflamed skin with hS100A15-L being the predominantly expressed mRNA isoform in both psoriasis and atopic eczema. In cultured keratinocytes, IL-1beta and Th1 cytokines significantly induced hS100A15-L compared with hS100A15-S. In contrast, Th2-derived cytokines had no influence on the expression of either hS100A15 splice variant. Differentiation of human keratinocytes induced by 1.2 mm calcium resulted in the upregulation of both hS100A15 mRNA isoforms. Our data show that both hS100A15 splice variants are differentially regulated and expressed with epidermal differentiation and skin inflammation. Overexpression of hS100A15 in chronic inflammatory skin diseases and regulation by inflammatory cytokines and calcium suggest that hS100A15 is involved in Th1-associated epithelial responses and epidermal maturation in normal and diseased human skin.

Expression of the human Cathepsin L inhibitor hurpin in mice: skin alterations and increased carcinogenesis.

The serine protease inhibitor (serpin) hurpin (serpin B13) is a cross class-specific inhibitor of the cysteine protease Cathepsin (Cat) L. Cat L is involved in lysosomal protein degradation, hair follicle morphogenesis, epidermal differentiation and epitope generation of antigens. Hurpin is a 44 kDa protein which is expressed predominantly in epidermal cells. In psoriatic skin samples, hurpin was strongly overexpressed when compared with normal skin. Keratinocytes overexpressing hurpin showed increased resistance towards UVB-induced apoptosis. To further analyse the functional importance of this inhibitor, we have generated transgenic mice with deregulated Cat L activity by expressing human hurpin in addition to the endogenous mouse inhibitor. The three independent transgenic lines generated were characterized by identical effects excluding insertional phenotypes. Macroscopically, mice expressing human hurpin are characterized by abnormal abdominal fur. The number of apoptotic cells and caspase-3 positive cells was reduced after UV-irradiation in transgenic animals compared with wild-type mice. Interestingly, after chemical carcinogenesis, transgenic mice showed an increased susceptibility to develop skin cancer. Array analysis of gene expression revealed distinct differences between wild-type and hurpin-transgenic mice. Among others, differentially expressed genes are related to antigen presentation and angiogenesis. These results suggest an important role of Cat L regulation by hurpin which might be of clinical relevance in human skin diseases.

S100A15, an antimicrobial protein of the skin: regulation by E. coli through Toll-like receptor 4.

E. coli is a gram-negative bacterium rarely found on human skin. We investigated whether direct interaction of E. coli with keratinocytes might induce an innate immune response through recognition by pattern recognition receptors. The capacity of E. coli to activate innate immune responses and IL-8 induction was investigated. We found that E. coli significantly induced human S100A7 and S100A15 transcript abundance and IL-8 release in cultured primary human keratinocytes. S100A15 is a member of the S100 protein family with previously unknown function. E. coli induced effects could be inhibited by neutralizing Toll-like receptor 4 (TLR4) antibodies, suggesting that E. coli-induced IL-8 and S100A15 expression in human keratinocytes are TLR4 dependent. TLR4-/- mice lacked elevated mS100A15 expression after infection with E. coli in contrast to wild-type mice. In vitro, human S100A15 displayed antimicrobial activity against E. coli. Our findings suggest that E. coli modulates S100A15 and IL-8 expression of keratinocytes by recognition through TLR4.

Expression of hurpin, a serine proteinase inhibitor, in normal and pathological skin: overexpression and redistribution in psoriasis and cutaneous carcinomas.

Hurpin was identified by differential display analysis studying UV-repressible genes in the keratinocyte cell line HaCaT. We have previously reported that hurpin mRNA is overexpressed in psoriatic skin compared to non-lesional or normal skin; hurpin inhibits cathepsin L and that, after overexpression in keratinocytes, hurpin decreases UV-induced apoptosis. To further study the expression of hurpin, we have isolated monoclonal antibodies against hurpin and analyzed its expression in normal and diseased skin by immunohistochemistry (IHC). In the epidermis of normal skin, we found hurpin to be mainly expressed in the stratum basale. In contrast, we found an enhanced expression of hurpin in the stratum spinosum and stratum granulosum in the majority of diseased skin samples. Within the dermis of normal and diseased skin, hurpin was detected in sebaceous and sweat glands, hair follicles, and endothelial cells of blood vessels. Hurpin was localized in the cytoplasm in normal and diseased skin. Additionally to IHC, we analyzed hurpin expression in selected skin diseases by semiquantitative reverse-transcription polymerase chain reaction. We found overexpression of hurpin mRNA in psoriasis, squamous cell carcinoma (SCC), and actinic keratosis. In contrast, expression of hurpin in melanoma and basal cell carcinoma was comparable to that in normal skin. Overall, the strongest overexpression was observed in SCC and psoriasis. Individual differences in hurpin expression between patients were observed. The increased expression and redistribution of hurpin in diseased skin suggests its possible involvement in inflammatory processes or the regulation of endogenous or pathogen-derived proteinase activity. Additional studies will elucidate the physiological role of hurpin.