Search Results (12)

Nile tilapia (Oreochromis niloticus) and European sea bass (Dicentrarchus labrax) are economically significant species in Mediterranean countries, serving essential roles in the aquaculture industry due to high market demand and nutritional value. They experience substantial losses from bacterial pathogens Vibrio anguillarum and Streptococcus iniae, particularly at the onset of the summer season. The immune mechanisms involved in fish infections by V. anguillarum and S. iniae remain poorly understood. This study investigated their impact through experiments with control and V. anguillarum- and S. iniae-infected groups for each species. Blood samples were collected at 1, 3, and 7 days post bacterial injection to assess biochemical and immunological parameters, including enzyme activities (AST and ALT), oxidative markers (SOD, GPX, CAT, and MDA), and leukocyte counts. Further analyses included phagocyte activity, lysozyme activity, IgM levels, and complement C3 and C4 levels. Muscle tissues were sampled at 1, 3, and 7 days post injection to assess mRNA expression levels of 18 immune-relevant genes. The focus was on cytokines and immune-related genes, including pro-inflammatory cytokines (TNF-α, TNF-β, IL-2, IL-6, IL-8, IL-12, and IFN-γ), major histocompatibility complex components (MHC-IIα and MHC-IIβ), cytokine receptors (CXCL-10 and CD4-L2), antimicrobial peptides (Pleurocidin and β-defensin), immune regulatory peptides (Thymosin β12, Leap 2, and Lysozyme g), and Galectins (Galectin-8 and Galectin-9). β-actin was used as the housekeeping gene for normalization. Significant species-specific responses were observed in N. Tilapia and E. Sea Bass when infected with V. anguillarum and S. iniae, highlighting differences in biochemical, immune, and gene expression profiles. Notably, in N. Tilapia, AST levels significantly increased by day 7 during S. iniae infection, reaching 45.00 ± 3.00 (p < 0.05), indicating late-stage acute stress or tissue damage. Conversely, E. Sea Bass exhibited a significant rise in ALT levels by day 7 in the S. iniae group, peaking at 33.5 ± 3.20 (p < 0.05), suggesting liver distress or a systemic inflammatory response. On the immunological front, N. Tilapia showed significant increases in respiratory burst activity on day 1 for both pathogens, with values of 0.28 ± 0.03 for V. anguillarum and 0.25 ± 0.02 for S. iniae (p < 0.05), indicating robust initial immune activation. Finally, the gene expression analysis revealed a pronounced peak of TNF-α in E. Sea Bass by day 7 post V. anguillarum infection with a fold change of 6.120, suggesting a strong species-specific pro-inflammatory response strategy. Understanding these responses provides critical insights for enhancing disease management and productivity in aquaculture operations. Full article

Background: The sixty-day effects of initial composite interventions for the treatment of severely and critically ill patients with COVID-19 are not fully assessed. Methods: Using a Bayesian piecewise exponential model, we analyzed the 60-day mortality, health-related quality of life (HRQoL), and disability in 1082 severely and critically ill patients with COVID-19 between 8 December 2022 and 9 February 2023 in Shanghai, China. The final 60-day follow-up was completed on 10 April 2023. Results: Among 1082 patients (mean age, 78.0 years, 421 [38.9%] women), 139 patients (12.9%) died within 60 days. Azvudine had a 99.8% probability of improving 2-month survival (adjusted HR, 0.44 [95% credible interval, 0.24–0.79]), and Paxlovid had a 91.9% probability of improving 2-month survival (adjusted HR, 0.71 [95% credible interval, 0.44–1.14]) compared with the control. IL-6 receptor antagonist, baricitinib and a-thymosin each had a high probability of benefit (99.5%, 99.4%, and 97.5%, respectively) compared to their controls, while the probability of trail-defined statistical futility (HR > 0.83) was high for therapeutic anticoagulation (99.8%; HR, 1.64 [95% CrI, 1.06–2.50]) and glucocorticoid (91.4%; HR, 1.20 [95% CrI, 0.71–2.16]). Paxlovid, Azvudine, and therapeutic anticoagulation showed a significant reduction in disability (p < 0.05) Conclusions: Among severely and critically ill patients with COVID-19 who received 1 or more therapeutic interventions, treatment with Azvudine had a high probability of improved 60-day mortality compared with the control, indicating its potential in a resource-limited scenario. Treatment with an IL-6 receptor antagonist, baricitinib, and a-thymosin also had high probabilities of benefit in improving 2-month survival, among which a-thymosin could improve HRQoL. Treatment with Paxlovid, Azvudine, and therapeutic anticoagulation could significantly reduce disability at day 60. Full article

Liver fibrosis is the most common feature of liver disease, and activated hepatic stellate cells (HSCs) are the main contributors to liver fibrosis. Thus, finding key targets that modulate HSC activation is important to prevent liver fibrosis. Previously, we showed that thymosin β4 (Tβ4) influenced HSC activation by interacting with the Hedgehog pathway in vitro. Herein, we generated Tβ4 conditional knockout (Tβ4-flox) mice to investigate in vivo functions of Tβ4 in liver fibrosis. To selectively delete Tβ4 in activated HSCs, double-transgenic (DTG) mice were generated by mating Tβ4-flox mice with α-smooth muscle actin (α-Sma)-Cre-ER^T2 mice, and these mice were administered carbon tetrachloride (CCl₄) or underwent bile duct ligation to induce liver fibrosis. Tβ4 was selectively suppressed in the activated HSCs of DTG mouse liver, and this reduction attenuated liver injury, including fibrosis, in both fibrotic models by repressing Hedgehog (Hh) signaling. In addition, the re-expression of Tβ4 by an adeno-associated virus reversed the effect of HSC-specific Tβ4 deletion and led to liver fibrosis with Hh activation in CCl₄-exposed mice treated with tamoxifen. In conclusion, our results demonstrate that Tβ4 is a crucial regulator of HSC activation, suggesting it as a novel therapeutic target for curing liver fibrosis. Full article

Thymosin α1 (Tα1) is an immunostimulatory peptide that is commonly used as an immune enhancer in viral infectious diseases such as hepatitis B, hepatitis C, and acquired immune deficiency syndrome (AIDS). Tα1 can influence the functions of immune cells, such as T cells, B cells, macrophages, and natural killer cells, by interacting with various Toll-like receptors (TLRs). Generally, Tα1 can bind to TLR3/4/9 and activate downstream IRF3 and NF-κB signal pathways, thus promoting the proliferation and activation of target immune cells. Moreover, TLR2 and TLR7 are also associated with Tα1. TLR2/NF-κB, TLR2/p38MAPK, or TLR7/MyD88 signaling pathways are activated by Tα1 to promote the production of various cytokines, thereby enhancing the innate and adaptive immune responses. At present, there are many reports on the clinical application and pharmacological research of Tα1, but there is no systematic review to analyze its exact clinical efficacy in these viral infectious diseases via its modulation of immune function. This review offers an overview and discussion of the characteristics of Tα1, its immunomodulatory properties, the molecular mechanisms underlying its therapeutic effects, and its clinical applications in antiviral therapy. Full article

The primary cilium (PC) is a microtubule-based antenna-like organelle projecting from the surface of the cell membrane. We previously reported that PC formation could be regulated by nephronophthisis 3 (NPHP3) expression followed by its interaction with thymosin β4. Here, we investigated whether cancer cell viability is regulated by NPHP3-mediated PC formation. The total and viable cell number were reduced by incubating cells under serum deprivation (SD) without fetal bovine serum (-FBS). PC frequency was increased by SD which enhanced NPHP3 expression and hypoxia inducible factor (HIF)-1α. The role of HIF-1α on NPHP3 expression and PC formation was confirmed by the binding of HIF-1α to the NPHP3 promoter and siRNA-based inhibition of HIF-1α (siHIF-1α), respectively. HIF-1α-stabilizing dimethyloxallyl glycine (DMOG) and hypoxic conditions increased NPHP3 expression and PC formation. In addition, as SD elevated the reactive oxygen species (ROS), PC frequency and NPHP3 expression were inhibited by a treatment with N-acetylcysteine (NAC), a ROS scavenger. PC formation was increased by H₂O₂ treatment, which was inhibited by siHIF-1α. The inhibition of ERK with P98059 decreased the frequency of PC formation and NPHP3 expression. Cell viability was reduced by a treatment with ciliobrevin A (CilioA) to inhibit PC formation, which was re-affirmed by using PC-deficient IFT88^−/− cells. Taken together, the results imply that PC formation in cancer cells could be controlled by NPHP3 expression through ROS-induced HIF-1α and ERK activation under SD conditions. It suggests that cancer cell viability under SD conditions could be maintained by NPHP3 expression to regulate PC formation. Full article

Due to its essential role in cellular processes, actin is a common target for bacterial toxins. One such toxin, TccC3, is an effector domain of the ABC-toxin produced by entomopathogenic bacteria of Photorhabdus spp. Unlike other actin-targeting toxins, TccC3 uniquely ADP-ribosylates actin at Thr-148, resulting in the formation of actin aggregates and inhibition of phagocytosis. It has been shown that the fully modified F-actin is resistant to depolymerization by cofilin and gelsolin, but their effects on partially modified actin were not explored. We found that only F-actin unprotected by tropomyosin is the physiological TccC3 substrate. Yet, ADP-ribosylated G-actin can be produced upon cofilin-accelerated F-actin depolymerization, which was only mildly inhibited in partially modified actin. The affinity of TccC3-ADP-ribosylated G-actin for profilin and thymosin-β4 was weakened moderately but sufficiently to potentiate spontaneous polymerization in their presence. Interestingly, the Arp2/3-mediated nucleation was also potentiated by T148-ADP-ribosylation. Notably, even partially modified actin showed reduced bundling by plastins and α-actinin. In agreement with the role of these and other tandem calponin-homology domain actin organizers in the assembly of the cortical actin network, TccC3 induced intense membrane blebbing in cultured cells. Overall, our data suggest that TccC3 imposes a complex action on the cytoskeleton by affecting F-actin nucleation, recycling, and interaction with actin-binding proteins involved in the integration of actin filaments with each other and cellular elements. Full article

The present investigation aimed to explore the intact proteome of tissues of pediatric brain tumors of different WHO grades and localizations, including medulloblastoma, pilocytic astrocytoma, and glioblastoma, in comparison with the available data on ependymoma, to contribute to the understanding of the molecular mechanisms underlying the onset and progression of these pathologies. Tissues have been homogenized in acidic water–acetonitrile solutions containing proteases inhibitors and analyzed by LC–high resolution MS for proteomic characterization and label-free relative quantitation. Tandem MS spectra have been analyzed by either manual inspection or software elaboration, followed by experimental/theoretical MS fragmentation data comparison by bioinformatic tools. Statistically significant differences in protein/peptide levels between the different tumor histotypes have been evaluated by ANOVA test and Tukey’s post-hoc test, considering a p-value > 0.05 as significant. Together with intact protein and peptide chains, in the range of molecular mass of 1.3–22.8 kDa, several naturally occurring fragments from major proteins, peptides, and proteoforms have been also identified, some exhibiting proper biological activities. Protein and peptide sequencing allowed for the identification of different post-translational modifications, with acetylations, oxidations, citrullinations, deamidations, and C-terminal truncations being the most frequently characterized. C-terminal truncations, lacking from two to four amino acid residues, particularly characterizing the β-thymosin peptides and ubiquitin, showed a different modulation in the diverse tumors studied. With respect to the other tumors, medulloblastoma, the most frequent malignant brain tumor of the pediatric age, was characterized by higher levels of thymosin β4 and β10 peptides, the latter and its des-IS form particularly marking this histotype. The distribution pattern of the C-terminal truncated forms was also different in glioblastoma, particularly underlying gender differences, according to the definition of male and female glioblastoma as biologically distinct diseases. Glioblastoma was also distinguished for the peculiar identification of the truncated form of the α-hemoglobin chain, lacking the C-terminal arginine, and exhibiting oxygen-binding and vasoconstrictive properties different from the intact form. The proteomic characterization of the undigested proteome, following the top-down approach, was challenging to originally investigate the post-translational events that differently characterize pediatric brain tumors. This study provides a contribution to elucidate the molecular profiles of the solid tumors most frequently affecting the pediatric age, and which are characterized by different grades of aggressiveness and localization. Full article

Human interferon α2 (IFNα2) and thymosin α1 (Tα1) are therapeutic proteins used for the treatment of viral infections and different types of cancer. Both IFNα2 and Tα1 show a synergic effect in their activities when used in combination. Furthermore, the therapeutic fusion proteins produced through the genetic fusion of two genes can exhibit several therapeutic functions in one molecule. In this study, we determined the anticancer and antiviral effect of human interferon α2–thymosin α1 fusion protein (IFNα2–Tα1) produced in our laboratory for the first time. The cytotoxic and genotoxic effect of IFNα2–Tα1 was evaluated in HepG2 and MDA-MB-231 cells. The in vitro assays confirmed that IFNα2–Tα1 inhibited the growth of cells more effectively than IFNα2 alone and showed an elevated genotoxic effect. The expression of proapoptotic genes was also significantly enhanced in IFNα2–Tα1-treated cells compared to IFNα2-treated cells. Furthermore, the HCV RNA level was significantly reduced in IFNα2–Tα1-treated HCV-infected Huh7 cells compared to IFNα2-treated cells. The quantitative PCR analysis showed that the expression of various genes, the products of which inhibit HCV replication, was significantly enhanced in IFNα2–Tα1-treated cells compared to IFNα2-treated cells. Our findings demonstrate that IFNα2–Tα1 is more effective than single IFNα2 as an anticancer and antiviral agent. Full article

The amount of bonds between constituting parts of a protein aggregate were determined in wild type (WT) and A53T α-synuclein (αS) oligomers, amyloids and in the complex of thymosin-β₄–cytoplasmic domain of stabilin-2 (Tβ₄-stabilin CTD). A53T αS aggregates have more extensive βsheet contents reflected by constant regions at low potential barriers in difference (to monomers) melting diagrams (MDs). Energies of the intermolecular interactions and of secondary structures bonds, formed during polymerization, fall into the 5.41 kJ mol⁻¹ ≤ E_a ≤ 5.77 kJ mol⁻¹ range for αS aggregates. Monomers lose more mobile hydration water while forming amyloids than oligomers. Part of the strong mobile hydration water–protein bonds break off and these bonding sites of the protein form intermolecular bonds in the aggregates. The new bonds connect the constituting proteins into aggregates. Amyloid–oligomer difference MD showed an overall more homogeneous solvent accessible surface of A53T αS amyloids. From the comparison of the nominal sum of the MDs of the constituting proteins to the measured MD of the Tβ₄-stabilin CTD complex, the number of intermolecular bonds connecting constituent proteins into complex is 20(1) H₂O/complex. The energies of these bonds are in the 5.40(3) kJ mol⁻¹ ≤ E_a ≤ 5.70(5) kJ mol⁻¹ range. Full article

Thymosin α1 (Tα1) is an immunostimulatory peptide for the treatment of hepatitis B virus (HBV) and hepatitis C virus (HCV) infections and used as an immune enhancer, which also offers prospects in the context of COVID-19 infections and cancer. Manufacturing of this N-terminally acetylated 28-residue peptide is demanding, and its short plasma half-life limits in vivo efficacy and requires frequent dosing. Here, we combined the PASylation technology with enzymatic in situ N-acetylation by RimJ to produce a long-acting version of Tα1 in Escherichia coli at high yield. ESI-MS analysis of the purified fusion protein indicated the expected composition without any signs of proteolysis. SEC analysis revealed a 10-fold expanded hydrodynamic volume resulting from the fusion with a conformationally disordered Pro/Ala/Ser (PAS) polypeptide of 600 residues. This size effect led to a plasma half-life in rats extended by more than a factor 8 compared to the original synthetic peptide due to retarded kidney filtration. Our study provides the basis for therapeutic development of a next generation thymosin α1 with prolonged circulation. Generally, the strategy of producing an N-terminally protected PASylated peptide solves three major problems of peptide drugs: (i) instability in the expression host, (ii) rapid degradation by serum exopeptidases, and (iii) low bioactivity because of fast renal clearance. Full article

β-thymosin is known for having 43 amino acids, being water-soluble, having a light molecular weight and ubiquitous polypeptide. The biological activities of β-thymosin are diverse and include the promotion of wound healing, reduction of inflammation, differentiation of T cells and inhibition of apoptosis. Our previous studies showed that oyster β-thymosin originated from the mantle of the Pacific oyster, Crassostrea gigas and had antimicrobial activity. In this study, we investigated the anti-inflammatory effects of oyster β-thymosin in lipopolysaccharide (LPS)-induced RAW264.7 macrophage cells using human β-thymosin as a control. Oyster β-thymosin inhibited the nitric oxide (NO) production as much as human β-thymosin in LPS-induced RAW264.7 cells. It also showed that oyster β-thymosin suppressed the expression of prostaglandin E₂ (PGE₂), inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). Moreover, oyster β-thymosin reduced inflammatory cytokines such as tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6). Oyster β-thymosin also suppressed the nuclear translocation of phosphorylated nuclear factor-κB (NF-κB) and degradation of inhibitory κB (IκB) in LPS-induced RAW264.7 cells. These results suggest that oyster β-thymosin, which is derived from the mantle of the Pacific oyster, has as much anti-inflammatory effects as human β-thymosin. Additionally, oyster β-thymosin suppressed NO production, PGE₂ production and inflammatory cytokines expression via NF-κB in LPS-induced RAW264.7 cells. Full article

Thymosin α1 (Tα1), is a peptidic hormone, whose immune regulatory properties have been demonstrated both in vitro and in vivo and approved in different countries for treatment of several viral infections and cancers. Tα1 assumes a conformation in negative membranes upon insertion into the phosphatidylserine exposure as found in several pathologies and in apoptosis. These findings are in agreement with the pleiotropy of Tα1, which targets both normal and tumor cells, interacting with multiple cellular components, and have generated renewed interest in the topic. Hyaluronan (HA) occurs ubiquitously in the extracellular matrix and on cell surfaces and has been related to a variety of diseases, and developmental and physiological processes. Proteins binding HA, among them CD44 and the Receptor for HA-mediated motility (RHAMM) receptors, mediate its biological effects. NMR spectroscopy indicated preliminarily that an interaction of Tα1 with HA occurs specifically around lysine residues of the sequence LKEKK of Tα1 and is suggestive of a possible interference of Tα1 in the binding of HA with CD44 and RHAMM. Further studies are needed to deepen these observations because Tα1 is known to potentiate the T-cell immunity and anti-tumor effect. The binding inhibitory activity of Tα1 on HA-CD44 or HA-RHAMM interactions can suppress both T-cell reactivity and tumor progression. Full article