Synapse (Netherlands)

Natural hydrogels (e.g., collagen hydrogels) show good potential in understanding cell-matrix interaction and find application in tissue engineering. However, it remains challenging to bioprint cell-laden natural hydrogels with good printability, shape retention and stability. In this study, non-water-soluble short collagen type I microfibers (COL-I μFiber) were blended with water-soluble methacrylated collagen peptide (COPMA) and xanthan gum (XG), forming an interpenetrated network, and bioprinted into stable natural-derived COPMA-μFiber-XG constructs, followed by in situ stem cell proliferation and differentiation. First, to enhance the printability and the mechanical properties of COPMA, a COPMA-μFiber-XG bioink was developed, featuring rapid UV-curing and self-healing properties. The encapsulated human mesenchymal stem cells (hMSCs) spread along the COL-I μFibers in the bioprinted constructs, with increased metabolic activity and production of extracellular matrix and bioactive proteins (COL-I and scleraxis) in 28 days. The internal biophysical and biochemical signals provided by COL-I μFibers and the fibrous COPMA matrix synergistically interacted with exogenous biochemical signals (e.g., transforming growth factor-beta 3) to further promote stem cell differentiation. Overall, bioprinted fibrous COPMA-μFiber-XG constructs are biocompatible and bioactive matrices to support hMSCs proliferation and differentiation.

In thrombosis and haemostasis, coagulation and platelet activation pathways culminate to form solid fibrin clots, which can become vaso-occlusive or prevent excessive bleeding. We report a novel mechanism describing how developing fibrin clots prolong and modulate the reactivity of thrombin, an enzyme propagating platelet and coagulation activation and forming fibrin from fibrinogen. Using immunological and genetic approaches, we delineate how thrombin bound to the A and Baachains of fibrin E-domains regulates lateral fibrin fibre extension. Our data reveal that fibrin-bound thrombin remains active and is temporarily protected against inactivation by antithrombin-III. Immunological displacement of thrombin from fibrin profoundly lowered its capacity, whereas a peptide mimicking the AA-chain binding-site increased its reactivity. In a cohort of patients with congenital dysfibrinogenemia, carrying FGA, FGB or FGG mutations associated with bleeding or thrombosis phenotypes, we noticed a high thrombin capacity and suppressed thrombinantithrombin-III complex formation, pointing to a prolonged active thrombin lifetime, likely due to abnormal formation of thrombin-containing fibrin. In conclusion, the combination of an impaired clotting and increased thrombogenicity may explain the paradoxical bleeding and thrombotic complications observed in such patients. Development of fibrin-directed agents may offer new therapeutic opportunities to normalize hemostasis or prevent thrombosis.

ABSTRACT The intraneuronal aggregation of tau is a key driver of pathogenesis in tauopathies such as Alzheimer’s disease. Passive immunotherapy is a promising strategy for targeting tau, with several tau-specific antibodies having demonstrated the ability to reduce tau pathology and improve behavioural deficits in tau transgenic mouse models. Despite preclinical promise, however, conventional antibodies have limited access to the cell cytoplasm where tau pathology itself originates and accumulates to cause downstream neuronal dysfunction. As such, conventional antibodies are typically limited to targeting extracellular tau, failing to address the primary site of tau pathogenesis. This challenge can be overcome by intracellular antibodies or intrabodies, small antibody fragments that can be expressed within cells to target intracellular antigens like tau. Here, we have generated a single-chain variable fragment derived from the N-terminal tau-specific antibody, RNJ1, and investigated its potential as an intrabody to reduce tau pathology and restore neuronal function in a tau transgenic mouse model. The RNJ1 intrabody successfully engaged intracellular tau and reduced total tau and phosphorylated tau inclusions in brains of tau transgenic mice. Furthermore, treatment with the RNJ1 intrabody in female tau transgenic mice induced restoration of various protein pathways important for cellular homeostasis, thus promoting the restoration of neuronal function. Our findings underscore the therapeutic utility of targeting intracellular tau in disease, providing novel insights into the potential mechanisms by which intrabodies ameliorate tau pathology.

Collagen peptide (COP) is water soluble, bioactive, and tends to be a promising alternative to collagen for tissue regeneration. However, its low viscosity and lack of readily polymerizable groups hinder its bioprinting and limit its wide applications in tissue engineering. In this study, methacrylated collagen peptide‐xanthan gum (COPMA‐XG) bioinks with interpenetrating networks are developed for bioprinting stable constructs, followed by stem cell differentiation. First, self‐healing COPMA hydrogels are developed with rapid UV‐curing and tunable mechanical properties. To increase the printability and the mechanical properties of COPMA, XG is mixed to create a set of COPMA‐XG bioinks. COPMA‐XG hydrogels show self‐healing properties, optimal printability, and stable morphology in the medium. The bioprinted human bone marrow mesenchymal stem cells (hMSCs) laden COPMA‐XG constructs are biocompatible and bioactive, with increased production of extracellular matrix, collagen type I, and scleraxis over 28 days. Overall, bioprinted COPMA‐XG constructs are versatile matrices to support hMSCs proliferation and differentiation with potential for ligament tissue engineering.

The hemostatic process relies on platelet and coagulation activation, with additional roles of red blood cells and the vessel wall. By systematic screening of databases for gene-linked information on hemostasis, we collected phenotypic profiles of 3474 orthologous human and mouse genes regarding bleeding, arterial thrombosis, thrombophilia, platelet traits, coagulation, and erythrocytes. Comparisons showed that defects in 252 mouse genes led to increased bleeding combined with platelet dysfunction or thrombocytopenia, in addition to 150 human orthologs that are registered for familial bleeding disorders, based on panel sequencing. Additionally, 139 mouse genes contributed to arterial thrombosis without bleeding phenotype. To further investigate the role of platelets in hemostasis, we integrated multiple genome-wide RNA-sequencing transcriptomes and proteomes from healthy subjects and C57BL/6 mice. This provided reference levels for 54 790 (54 247) transcripts and 6379 (4563) proteins in human (mouse) platelets. Orthologous transcripts in human and mouse platelets correlated with R=0.75, whereas orthologous platelet proteins correlated with R=0.87. Comparison with the phenotypic analysis revealed the following: (i) overall high qualitative similarity of human and mouse platelets regarding composition and function; (ii) presence of transcripts in platelets for most of the 3474 phenotyped genes; (iii) preponderance of syndromic platelet-expressed genes; and (iv) 20-40% overlap with genes from genome-wide association studies. For 42 mouse genes, among which receptors, signaling proteins, and transcription regulators (ASXL1, ERG, GATA2, MEIS1, NFE2, and TAL1), we confirmed novel links with human platelet function or count. This interspecies comparison can serve as a valuable resource for researchers and clinicians studying the genetics of blood-borne hemostasis and thrombosis.

ABSTRACTThe classical psychedelic drug (+)‐lysergic acid diethylamide (LSD) continues to attract considerable multidisciplinary interest, and over the last eight decades, many derivatives and analogs of LSD have been synthesized. One site on the ergoline scaffold of LSD that has been frequently modified is the N1‐position, with the N1‐acylated LSD derivative 1‐acetyl‐LSD (1A‐LSD, ALD‐52) being one of the earliest examples. In more recent years, several other alkylcarbonyl‐ and cycloalkylcarbonyl‐substituted LSD derivatives have been evaluated, including several distributed as research chemicals. Although N1‐substitution is detrimental for the activity of LSD at the 5‐HT2A receptor (the primary site of action of psychedelic drugs), N1‐acylated LSD derivatives are rapidly hydrolyzed in vivo and are believed to act as prodrugs for LSD. Recently, 1‐(thiophene‐2‐carbonyl)‐LSD (1T‐LSD, SYN‐L‐021) was detected as a new recreational drug, signaling a move towards N1‐acyl groups with an aromatic character. The present study was conducted to investigate the analytical profile and pharmacology of 1‐(2‐furoyl)‐lysergic acid diethylamide (1F‐LSD, SYN‐L‐005), a novel analog of 1T‐LSD. The binding of 1F‐LSD to the 5‐HT2A receptor and other monoamine sites was assessed using radioligand binding. Furthermore, the in vivo activities of 1F‐LSD and 1T‐LSD were assessed in C57BL/6 J mice by comparing their biotransformation to LSD and effects on the head‐twitch response (HTR), a 5‐HT2A‐mediated behavior. Both 1F‐LSD and 1T‐LSD induced the HTR in mice and were hydrolyzed to LSD after in vivo administration, indicating that both substances exhibit LSD‐like properties and may serve as prodrugs for LSD.