Steffen Ulrich Eisenhardt

Autosomal recessive nonsyndromic epidermal differentiation disorders (AR‐nEDDs), also known as autosomal recessive congenital ichthyosis (ARCI), are rare genetic skin diseases that lack curative treatments and can only be managed symptomatically. Their study is hampered by the limited availability of biological samples and donor tissues. Artificial skin models offer a valuable alternative for experimental research. Here, we generated epidermal skin equivalents (ESE) using primary keratinocytes from patients with AR‐nEDDs carrying pathogenic variants in ALOX12B , CYP4F22 , and CERS3 , as well as immortalized N/TERT‐2G cells. Whereas primary cells undergo senescence, N/TERT‐2G cells offer a promising alternative to overcome this limitation. Histological staining and qPCR were applied to assess key epidermal proteins and AR‐nEDD‐related gene expression in patient‐ and control‐derived models. Patient‐derived ESE reproduced the major histopathological features and protein expression patterns characteristic of AR‐nEDDs. In contrast, N/TERT‐2G‐based models showed earlier expression of 12R‐LOX, CYP4F22, and CERS3 proteins in epidermal layers compared to healthy donor equivalents. These findings suggest that N/TERT‐2G cells represent a reproducible platform for future gene‐editing approaches of modelling epidermal differentiation disorders. However, as they do not inherently carry pathogenic variants, they are particularly suitable for gene‐editing approaches such as CRISPR/Cas9‐mediated disease modelling. Conversely, patient‐derived keratinocyte models remain indispensable for validating disease mechanisms and evaluating therapeutic strategies.

Background: This study evaluates the time efficiency and safety of Wide Awake Local Anesthesia No Tourniquet (WALANT) compared to general/plexus anesthesia in emergency hand surgery. Methods: This single-center, retrospective study analyzed 812 emergency category 2b hand surgeries performed between 2018 and 2024. Patients were divided into 2 groups: local anesthesia (LA), including WALANT, and general/plexus anesthesia. In collaboration with the Institute for Medical Statistics and Biometry, “time to intervention” and postoperative surgical complications were prospectively defined as co-primary endpoints. In addition, a descriptive analysis of the study population was conducted, and a matched-pair analysis was performed to account for injury complexity. Statistical analysis included t -tests, Cohen’s d, and Gart and Nam’s method for risk difference. Results: The mean “time to intervention” was significantly shorter in the LA group (2.2 hours) compared to that in the general/plexus anesthesia group (4.1 hours). Postoperative surgical complication rates were comparable between the LA (8.5%) and general/plexus anesthesia (8.2%) groups, with no significant difference observed. WALANT showed a lower complication rate (5.7%) than LA without epinephrine (9.8%). A matched-pair analysis confirmed these findings. Among the general/plexus anesthesia cases, 16.2% exceeded the recommended 6-hour treatment window, compared to 6.6% in the LA group. Conclusions: Wide Awake Local Anesthesia No Tourniquet facilitates faster treatment of emergency hand surgery cases while maintaining equivalent quality of care compared to general/plexus anesthesia. This study highlights the time efficiency and safety of WALANT in emergency settings, supporting its broader application.

Introduction: Scapholunate instability is a common and challenging wrist pathology with no universally accepted surgical solution. The all-dorsal scapholunate reconstruction technique restores the dorsal ligament with an extensor carpi radialis brevis tendon autograft. The aim of this study was to assess outcomes in patients with a chronic scapholunate ligament rupture. Methods: We retrospectively analysed 31 patients. Outcomes were assessed with the Disabilities of the Arm, Shoulder and Hand (DASH) score and a visual analogue scale score for pain. Clinical examination included wrist range of motion and grip strength. The radiological parameters measured were scapholunate (SL) angle and gap. Follow-up averaged 39 months (range 12–84) for all outcomes. Results: Postoperatively median pain scores improved from 5 to 2 and disability scores from 58 to 48, although changes were not statistically significant. Median wrist extension at final follow-up measured 50° (IQR 42.5 to 70) and flexion 45° (IQR 39 to 60). Radiographically, there was persistent carpal malalignment with a median SL angle of 66° (IQR 58 to 73), a static SL gap of 3 mm (IQR 2.2 to 4) and a dynamic SL gap of 4.8 mm (IQR 3.3 to 5.4). Median grip strength improved from 28 kg (IQR 20 to 40) preoperatively to 29 kg (20–37) postoperatively. Five patients required salvage procedures owing to progressive symptomatic osteoarthritis. Complications included complex regional pain syndrome ( n = 3), transient nerve symptoms ( n = 2), anchor-site osteolysis ( n = 13) and isolated scaphocapitate joint osteoarthritis ( n = 5). Conclusion: The relative simplicity and preservation of wrist function with the all-dorsal reconstruction technique justify its continued use, despite high complication and revision incidences. Level of Evidence: Level IV

The pentameric C-reactive protein (pCRP), an acute-phase protein, binds to lysophosphatidylcholine (LPC) displayed on the surface of dying cells and microorganisms to activate the complement system and to opsonize immune cells via Fcγ-receptors (FcγRs). Members of the FcγR family are characterized by the recognition of the Fc part of IgG antibodies. We utilized a mouse thymoma BW5147 reporter cell panel stably expressing chimeric human FcγR-CD3ζ-chain receptors to define the molecular requirements for FcγR crosslinking by C-reactive protein (CRP). Applying this approach, we show a robust activation of CD64/FcγRI and CD32a/FcγRIIa by immobilized CRP isoforms as well as triggering of inhibitory CD32b/FcγRIIb. Of note, activation of FcγRIIa was restricted to the 131R allelic variant but not observed with 131H. In contrast, FcγRIII isoforms CD16aF, CD16aV and CD16b were not activated by pCRP, although binding of CRP isoforms to FcγRIII was detectable. Activation of FcγRs by free pCRP in solution phase was considerably lower than with immobilized pCRP on hydrophilic plastic surfaces and readily abolished by IgG at serum level concentrations, whereas it was enhanced by the addition of streptococci. The types of FcγRs mainly responding to pCRP in solution phase (CD64/FcγRI and CD32aR/FcγRIIaR) clearly differed from FcγRs responding to soluble multimeric IgG complexes (i.e., CD16aV/FcγRIIIaV and CD32aH/FcγRIIaH). Compared to pCRP, monomeric CRP (mCRP) showed lower levels of activation in those selective FcγRs. FcγR activation was linked to recognition by conformation-dependent CRP antibodies. Unmasking of the mAb 9C9-defined neoepitope in pCRP* correlated with the triggering of FcγRs, indicating that pCRP* is the major FcγR-activating CRP conformation. The assay provides a novel, scalable approach to determine the molecular properties of CRP as a physiological ligand of FcγR-mediated bioactivities. Scope statement: Fcγ receptors (FcγRs) are important immune regulators that come in different variants and combinations, making it difficult to predict which components will ultimately lead to immunological effector functions. Classical FcγRs are defined by their recognition of IgG-Fc, while other ligands, such as C-reactive protein (CRP), are often neglected. Circulating concentrations of CRP, an acute phase protein, are elevated during inflammatory responses. As a pattern recognition receptor, CRP binds to lysophosphatidylcholine expressed on the surface of dying cells and microbes in order to activate the complement system via C1q. We have established a reporter cell assay platform that goes beyond ligand binding and takes a deeper look at the activation outcome(s) by CRP compared with IgG-Fc. This is the first comprehensive study defining CRP-responsive vs non-responsive FcγRs and investigating the interaction of FcγRs with CRP isoforms (pCRP/pCRP*/mCRP). We distinguish binding from receptor triggering using reporter cells stably expressing a chimeric FcγR-CD3ζ chain, thereby defining the molecular requirements for FcγR cross-linking by CRP. The assay provides a novel, sensitive and scalable approach to the properties of CRP as a ligand inducing FcγR-mediated bioactivities.

Fibrosis is the pathophysiologic hallmark of chronic rejection after lung transplantation and the foremost hurdle to long-term recipient survival. Several murine lung transplantation models are available for the study of chronic rejection. However, they display heterogeneous results regarding fibrotic changes of the graft, and the histologic extent of fibrosis is mostly reported qualitatively. Therefore, a spatially resolved approach that allows for statistical analysis can aid in evaluating fibrosis in these models. This study presents Picrosirius Red staining for a semiquantitative evaluation of collagen organization in murine lung allografts and compares it to standard Hematoxylin and Eosin, Masson's Trichrome, and Herovici's stains. Staining was performed on sections from two different murine transplantation models based on minor and major histocompatibility complex (MHC) mismatches. The method was established for semiquantitative analysis of collagen organization in whole lung sections. Thus, it can serve as a tool for murine experimental models of fibrotic lung diseases.

BACKGROUND: CRP (C-reactive protein) is a prototypical acute phase reactant. Upon dissociation of the pentameric isoform (pCRP [pentameric CRP]) into its monomeric subunits (mCRP [monomeric CRP]), it exhibits prothrombotic and proinflammatory activity. Pathophysiological shear rates as observed in aortic valve stenosis (AS) can influence protein conformation and function as observed with vWF (von Willebrand factor). Given the proinflammatory function of dissociated CRP and the important role of inflammation in the pathogenesis of AS, we investigated whether shear stress can modify CRP conformation and induce inflammatory effects relevant to AS. METHODS: To determine the effects of pathological shear rates on the function of human CRP, pCRP was subjected to pathophysiologically relevant shear rates and analyzed using biophysical and biochemical methods. To investigate the effect of shear on CRP conformation in vivo, we used a mouse model of arterial stenosis. Levels of mCRP and pCRP were measured in patients with severe AS pre- and post-transcatheter aortic valve implantation, and the presence of CRP was investigated on excised valves from patients undergoing aortic valve replacement surgery for severe AS. Microfluidic models of AS were then used to recapitulate the shear rates of patients with AS and to investigate this shear-dependent dissociation of pCRP and its inflammatory function. RESULTS: Exposed to high shear rates, pCRP dissociates into its proinflammatory monomers (mCRP) and aggregates into large particles. Our in vitro findings were further confirmed in a mouse carotid artery stenosis model, where the administration of human pCRP led to the deposition of mCRP poststenosis. Patients undergoing transcatheter aortic valve implantation demonstrated significantly higher mCRP bound to circulating microvesicles pre-transcatheter aortic valve implantation compared with post-transcatheter aortic valve implantation. Excised human stenotic aortic valves display mCRP deposition. pCRP dissociated in a microfluidic model of AS and induces endothelial cell activation as measured by increased ICAM-1 (intercellular adhesion molecule 1) and P-selectin expression. mCRP also induces platelet activation and TGF-β (transforming growth factor beta) expression on platelets. CONCLUSIONS: We identify a novel mechanism of shear-induced pCRP dissociation, which results in the activation of cells central to the development of AS. This novel mechanosensing mechanism of pCRP dissociation to mCRP is likely also relevant to other pathologies involving increased shear rates, such as in atherosclerotic and injured arteries.