A huge number of inbred mouse strains with different bone properties have become available for musculoskeletal research. C57Bl/6J and C3H/HeOuJ mice show a significant difference in their bone characteristics. Nevertheless, there is a lack of knowledge on the molecular basis of these strain differences. The aim of this study is to determine the gene expression of selected regulators expressed in the bone marrow as well as bone microstructure of C57Bl/6J and C3H/HeOuJ mice. Bone properties were investigated in 20-week-old female C57Bl/6J and C3H/HeOuJ mice. Total RNA was extracted from the bone marrow of the tibia and gene expression of the following genes was determined by quantitative real-time PCR: SOST, DKK1, OPN, FGF23, RANKL, IL6, TNF, IL17a, and OPG. The femur and third lumbar vertebral body (L3) were investigated by μCT. Bone histomorphometric evaluations were performed in tartrate-resistant acid phosphatase/toluidine blue stained fourth lumbar vertebral bodies (L4). C57Bl/6J and C3H/HeOuJ mice showed significant differences in the gene expression of DKK1, FGF23, IL-6, TNF, and OPG. When compared with C57Bl/6J mice, C3H/HeOuJ mice had a stronger cortical and trabecular bone microstructure at the femur. In contrast, at L3 bone volume/total volume (BV/TV) and trabecular number were significantly higher in C57Bl/6J than in C3H/HeOuJ mice. Bone histomorphometry of L4 revealed significantly higher BV/TV, trabecular number, and thickness in C57Bl/6J mice. Furthermore, the number of osteoblasts and the number of osteoclasts/bone perimeter were higher in the C57Bl/6J mice. This study shows that C57Bl/6J and C3H/HeOuJ mice exhibit a differential expression of cytokines present in the bone marrow. Bone properties differ not only between both strains but also in relation to the investigated bone region.
- Klíčová slova
- RNA, bone histomorphometry, bone metabolism, microCT, mouse strains, osteoimmunology,
- Publikační typ
- časopisecké články MeSH
2D materials have rapidly gained attention due to their exceptional properties like high surface area, flexibility, and tunable electronic characteristics. These attributes make them highly versatile for applications in energy storage, electronics, and biomedicine. Inspired by graphene's success, researchers are exploring other 2D materials from bulk crystals. Electrochemical exfoliation (ECE) is an efficient method for producing these materials, offering more sustainable mild conditions, quick processing, simple equipment, and high yields. While substantial progress has been made in the ECE of layered van der Waals (L-vdW) crystals, the exploration of layered non-van der Waals (L-NvdW) materials remains in its early stages. This review delves into using ECE to create 2D nanoplatelets from L-NvdW crystals. A comparative analysis of exfoliation techniques is provided for L-vdW and L-NvdW materials, followed by a comprehensive overview of recent advances in ECE methods applied to L-NvdW crystals. The discussion is organized around key categories, including the selective extraction of "M" and "A" layers respectively from MAX phases, decalcification of Zintl phases, and oxide delocalization from metal oxides. It is concluded by highlighting the potential applications of these 2D materials and discussing the challenges and future directions in this evolving field.
- Klíčová slova
- 2D materials, delocalization, electrochemical exfoliation, layered non‐van der Waals,
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
- Publikační typ
- tisková chyba MeSH
Rhomboid proteases are ubiquitous intramembrane serine proteases that can cleave transmembrane substrates within lipid bilayers. They exhibit many and diverse functions, such as but not limited to, growth factor signaling, immune and inflammatory response, protein quality control, and parasitic invasion. Human rhomboid protease RHBDL4 has been demonstrated to play a critical role in removing misfolded proteins from the endoplasmic reticulum and is implicated in severe diseases such as various cancers and Alzheimer's disease. Therefore, RHBDL4 is expected to constitute an important therapeutic target for such devastating diseases. Despite its critical role in many biological processes, the enzymatic properties of RHBDL4 remain largely unknown. To enable a comprehensive characterization of RHBDL4's kinetics, catalytic parameters, substrate specificity, and binding modality, we expressed and purified recombinant RHBDL4 and employed it in a Förster resonance energy transfer-based cleavage assay. Until now, kinetic studies have been limited mostly to bacterial rhomboid proteases. Our in vitro platform offers a new method for studying RHBDL4's enzymatic function and substrate preferences. Furthermore, we developed and tested potential inhibitors using our assay and successfully identified peptidyl α-ketoamide inhibitors of RHBDL4 that are highly effective against recombinant RHBDL4. We utilize ensemble docking and molecular dynamics simulations to explore the binding modality of substrate-derived peptides bound to RHBDL4. Our analysis focused on key interactions and dynamic movements within RHBDL4's active site that contributed to binding stability, offering valuable insights for optimizing the nonprime side of RHBDL4 ketoamide inhibitors. In summary, our study offers fundamental insights into RHBDL4's catalytic activities and substrate preferences, laying the foundation for downstream applications such as drug inhibitor screenings and structure-function studies, which will enable the identification of lead drug compounds for RHBDL4.
- Klíčová slova
- endoplasmic reticulum stress, endoplasmic-reticulum-associated protein degradation, enzyme inhibitor, enzyme kinetics, enzyme purification, enzyme structure, protein misfolding, rhomboid protease, serine protease,
- MeSH
- kinetika MeSH
- lidé MeSH
- membránové proteiny * metabolismus chemie genetika antagonisté a inhibitory MeSH
- rekombinantní proteiny chemie metabolismus genetika MeSH
- rezonanční přenos fluorescenční energie MeSH
- serinové endopeptidasy * chemie metabolismus genetika MeSH
- substrátová specifita MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- membránové proteiny * MeSH
- rekombinantní proteiny MeSH
- serinové endopeptidasy * MeSH
Syphilis is a multistage sexually transmitted disease caused by Treponema pallidum ssp. pallidum (TPA). This study analyzed clinical samples collected from patients with a diagnosed syphilis infection from 2004-2022, isolated in the Czech Republic. Mucocutaneous swab samples (n = 543) from 543 patients were analyzed, and from these samples, 80.11 % (n = 435) were PCR positive, and 19.89 % (n = 108) were PCR negative for TPA DNA. Swabs were more often positive when collected from syphilis patients in the primary and secondary stages, compared to the latent or unknown stage. There was no significant difference in PCR positivity between the primary and secondary stages (p = 0.099). In IgM-positive patients, a statistically significant association with PCR-positivity was found in samples from seropositive (p = 0.033) and serodiscrepant (RPR negative) patients (p = 0.0006). When assessing our laboratory-defined cases of syphilis, the RPR, IgM, and PCR tests were similarly effective (within the range of 80.1-86.1 %). However, parallel testing with these methods was even more effective, i.e., RPR + PCR was 96.1 % effective and RPR + IgM + PCR was 97.8 % effective. A combination of RPR + PCR, or a combination of all three tests (RPR, IgM, and PCR) can therefore be used to reliably detect active syphilis cases, including reinfections. Our findings show that the reverse algorithm for detecting syphilis could be substantially improved by adding IgM and PCR testing.
- Klíčová slova
- Nontreponemal tests, PCR syphilis detection, RPR, Syphilis, Syphilis diagnostics, Syphilis serology, Treponemal tests,
- MeSH
- DNA bakterií genetika MeSH
- dospělí MeSH
- imunoglobulin M * krev MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladý dospělý MeSH
- polymerázová řetězová reakce * metody MeSH
- protilátky bakteriální krev MeSH
- senzitivita a specificita MeSH
- sérologická diagnostika syfilis metody MeSH
- syfilis * diagnóza mikrobiologie MeSH
- Treponema pallidum * genetika izolace a purifikace imunologie MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Česká republika epidemiologie MeSH
- Názvy látek
- DNA bakterií MeSH
- imunoglobulin M * MeSH
- protilátky bakteriální MeSH
The integration of 3D printing into the pharmaceutical sciences opens new possibilities for personalized medicine. Poly(lactide) (PLA), a biodegradable and biocompatible polymer, is highly suitable for biomedical applications, particularly in the context of 3D printing. However, its processability often requires the addition of plasticizers. This study investigates the use of phase diagram modeling as a tool to guide the rational selection of plasticizers and to assess their impact on the thermodynamic and kinetic stability of PLA-based amorphous solid dispersions (ASDs) containing active pharmaceutical ingredients (APIs). Thermodynamic stability against API recrystallization was predicted based on the API solubility in PLA and Plasticizer-PLA carriers using the Conductor-like Screening Model for Real Solvents (COSMO-RS), while the kinetic stability of the ASDs was evaluated by modeling the glass transition temperatures of the mixtures. Two APIs, indomethacin (IND) and naproxen (NAP), with differing glass-forming abilities (i.e., recrystallization tendencies), and three plasticizers, triacetin (TA), triethyl citrate (TEC), and poly(L-lactide-co-caprolactone) (PLCL), were selected for investigation. The physical stability of ASD formulations containing 9 wt% API and plasticizer to PLA in two ratios, 10:81 and 20:71 w/w %, was monitored over time using differential scanning calorimetry and X-ray powder diffraction and compared with phase diagram predictions. All formulations were predicted to be thermodynamically unstable; however, those containing no plasticizer or with TEC and TA at 10 wt% were predicted to exhibit some degree of kinetic stability. Long-term physical studies corroborated these predictions. The correlation between the predicted phase behavior and long-term physical stability highlights the potential of phase diagram modeling as a tool for the rational design of ASDs in pharmaceutical 3D printing.
- Klíčová slova
- 3D printing, Active pharmaceutical ingredient, Amorphous solid dispersion, COSMO-RS, PLA, Personalized medicine, Phase diagram, Plasticizers,
- MeSH
- 3D tisk * MeSH
- citráty chemie MeSH
- diferenciální skenovací kalorimetrie metody MeSH
- farmaceutická chemie metody MeSH
- farmaceutická technologie metody MeSH
- indomethacin * chemie MeSH
- krystalizace MeSH
- naproxen chemie MeSH
- polyestery * chemie MeSH
- rozpouštědla chemie MeSH
- rozpustnost * MeSH
- stabilita léku MeSH
- termodynamika MeSH
- tranzitní teplota MeSH
- triacetin chemie MeSH
- změkčovadla * chemie MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- citráty MeSH
- ethyl citrate MeSH Prohlížeč
- indomethacin * MeSH
- naproxen MeSH
- poly(lactide) MeSH Prohlížeč
- polyestery * MeSH
- rozpouštědla MeSH
- triacetin MeSH
- změkčovadla * MeSH
Toll-like receptors (TLRs) play important roles in innate immunity and developmental processes. Due to their nature as molecular pattern recognition receptors, their genetic diversity may reflect the effects of various pathogen pressures. Here, the extent of variability in the TLR1-6-10 gene cluster in three geographically and historically distinct breeds of horses was analysed. A genetically diverse group of representatives of 14 other horse breeds provided additional information on the variability of this gene cluster in the domestic horse. Altogether, 25 SNPs were identified in the TLR6-1-10 gene cluster across the 4 equine breed groups studied, of which 7 were synonymous and 18 non-synonymous. Twenty-eight inferred SNPs and 22 in silico translated amino acid haplotypes were identified. A predominant major haplotype present in all breed groups along with several group-specific haplotypes were identified. Strong linkage disequilibrium was detected for several SNPs, as well as effects of pervasive, site-specific selection. The existence of a major haplotype suggests it may confer a selective advantage across breeds. Less frequent breed-specific haplotypes may represent variability required or beneficial for responses to local pathogen pressures. Purifying site-specific selection was detected in the TIR domain and its vicinity in TLR6, whereas AA sites under diversifying selection were located in LRR domains and/or their surroundings in TLR1. Population structure models based on immune-related TLR6-1-10 markers did not distinguish between breed groups, whereas in models based on neutral microsatellite markers, breed groups clustered separately. This supports the assumption that the diversity of the TLR6-1-10 cluster is of adaptive value. The TLR6-1-10 alleles and haplotypes identified represent potential candidate markers for disease association studies.
- Klíčová slova
- equine, haplotype, innate immunity, toll‐like receptor,
- MeSH
- genetická variace * MeSH
- haplotypy * MeSH
- jednonukleotidový polymorfismus * MeSH
- koně genetika MeSH
- multigenová rodina MeSH
- přirozená imunita * genetika MeSH
- toll-like receptor 6 genetika MeSH
- toll-like receptory genetika MeSH
- vazebná nerovnováha MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- toll-like receptor 6 MeSH
- toll-like receptory MeSH
During GABAergic synaptic transmission, G protein-coupled GABAB receptors (GBRs) activate K+ channels that prolong the duration of inhibitory postsynaptic potentials (IPSPs). We now show that KCTD16, an auxiliary GBR subunit, anchors hyperpolarization-activated cyclic nucleotide-gated (HCN) channels containing HCN2/HCN3 subunits to GBRs. In dopamine neurons of the VTA (DAVTA neurons), this interaction facilitates activation of HCN channels via hyperpolarization during IPSPs, counteracting the GBR-mediated late phase of these IPSPs. Consequently, disruption of the GBR/HCN complex in KCTD16-/- mice leads to prolonged optogenetic inhibition of DAVTA neuron firing. KCTD16-/- mice exhibit increased anxiety-like behavior in response to stress - a behavior replicated by CRISPR/Cas9-mediated KCTD16 ablation in DAVTA neurons or by intra-VTA infusion of an HCN antagonist in wild-type mice. Our findings support that the retention of HCN channels at GABAergic synapses by GBRs in DAVTA neurons provides a negative feedback mechanism that restricts IPSP duration and mitigates the development of anxiety.
- Klíčová slova
- GABA-A, GABA-B, HCN2, Hyperpolarization-activated cyclic nucleotide-gated channels, Late IPSP, Optogenetic inhibition, Slow IPSP,
- MeSH
- dopaminergní neurony * metabolismus MeSH
- hyperpolarizační iontové kanály řízené cyklickými nukleotidy * metabolismus genetika MeSH
- inhibiční postsynaptické potenciály fyziologie účinky léků MeSH
- myši inbrední C57BL MeSH
- myši knockoutované MeSH
- myši MeSH
- receptory GABA-B * metabolismus MeSH
- tegmentum mesencephali - area ventralis * metabolismus MeSH
- úzkost * metabolismus MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- hyperpolarizační iontové kanály řízené cyklickými nukleotidy * MeSH
- receptory GABA-B * MeSH
Restricted range size brings about noteworthy genetic consequences that may affect the viability of a population and eventually its extinction. Particularly, the question if an increase in inbreeding can avert the accumulation of genetic load via purging is hotly debated in the conservation genetic field. Insular populations with limited range sizes represent an ideal setup for relating range size to these genetic factors. Leveraging a set of eight differently sized populations of Galápagos mockingbirds (Mimus), we investigated how island size shaped effective population size (Ne), inbreeding and genetic load. We assembled a genome of M. melanotis and genotyped three individuals per population by whole-genome resequencing. Demographic inference showed that the Ne of most populations remained high after the colonisation of the archipelago 1-2 Mya. Ne decline in M. parvulus happened only 10-20 Kya, whereas the critically endangered M. trifasciatus showed a longer history of reduced Ne. Despite these historical fluctuations, the current island size determines Ne in a linear fashion. In contrast, significant inbreeding coefficients, derived from runs of homozygosity, were identified only in the four smallest populations. The index of additive genetic load suggested purging in M. parvulus, where the smallest populations showed the lowest load. By contrast, M. trifasciatus carried the highest genetic load, possibly due to a recent rapid bottleneck. Overall, our study demonstrates a complex effect of demography on inbreeding and genetic load, providing implications in conservation genetics in general and in a conservation project of M. trifasciatus in particular.
- Klíčová slova
- conservation genetics, demographic inference, genetic diversity, genetic load,
- MeSH
- genetická zátěž * MeSH
- genom genetika MeSH
- genotyp MeSH
- hustota populace * MeSH
- inbreeding * MeSH
- ostrovy * MeSH
- Passeriformes genetika MeSH
- populační genetika * MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Geografické názvy
- Ekvádor MeSH
- ostrovy * MeSH
