生物学最新文献

Stem cell-based therapies are emerging as a promising treatment for diabetes by differentiating these cells into insulin-producing cells (IPCs). However, using growth factors for differentiation has always been challenging. Physical differentiation of stem cells presents a promising approach to reduce reliance on chemical growth factors. One method of physical cell differentiation is cell imprinting. This study aimed to physically induce the differentiation of rat adipose-derived mesenchymal stem cells (rADSCs) into β-like cells using the cell-imprinting technique. For this purpose, RIN-5F cells were used to transfer their geometry and cell-specific topographies to a polydimethylsiloxane (PDMS) substrate. After cell imprinting, the rADSCs were seeded on the substrate, and their differentiation into β-like cells was evaluated after 14 and 21 days by assessing insulin production using dithizone staining and ELISA, as well as real-time PCR and immunocytochemistry (ICC) for expression analysis of the genes effective in cell differentiation into β-like cells, including PDX1, NKX6.1, NGN3, and insulin. The results of dithizone staining and ELISA confirmed insulin secretion by differentiated cells compared to stem cells (p ≤ 0.05). Real-time PCR and ICC results showed that after 21 days, the differentiated cells expressed key β-cell genes significantly more than stem cells (p ≤ 0.05).

Background: Fatty acid oxidation (FAO) is implicated in lung diseases, but its role in bronchial asthma is not fully understood. We investigated its effect on airway epithelial barrier integrity.

Methods: Using a house dust mite (HDM)-induced murine asthma model and HDM, IL-4, IL-13, or TNF-α stimulated human primary bronchial epithelial cells (BECs) and bronchial epithelial (Beas-2b) cells, we modulated FAO with L-carnitine (agonist) and Etomoxir (inhibitor). BECs and Beas-2b cells were infected with lentivirus-mediated CPT1A shRNA prior to stimulation. Barrier function, mitochondrial oxidative stress, inflammation, and metabolism were assessed.

Results: FAO level in lungs negatively correlated with increased inflammation and tissue injury in HDM-induced asthmatic mice (all p < 0.05), while positively regulating tight junction protein expression. In BECs and Beas-2b cells, Etomoxir treatment and CPT1A knockdown exacerbated the impairment of FAO caused by various stimulants (all p < 0.05). Furthermore, FAO negatively regulated HDM/cytokine-induced epithelial barrier damage, hyperactive inflammatory response, and mitochondrial dysfunction in Beas-2b cells (all p < 0.05). In contrast, treatment with L-carnitine significantly alleviated these pathophysiological features in both in vivo and in vitro models.

Conclusion: FAO plays a protective role in the occurrence and development of asthma by maintaining airway epithelial cell homeostasis and barrier function.

Infectious prions (PrP^Sc) are largely resistant to proteolytic digestion, including proteinase K (PK) digestion. While nucleic acid extracts are generally considered non-infectious from a classical microbiology context (i.e. free of intact bacteria and viruses), we investigated whether standard DNA purification methods co-purify PrP^Sc, posing an unrecognized biosafety risk. Commercial DNA extraction kits can eliminate conventional pathogens but are likely ineffective against PrP^Sc due to resistance to kit reagents and enzymatic degradation. Two laboratories, the University of Minnesota Center for Prion Research and Outreach (MNPRO) and the Canadian Food Inspection Agency (CFIA), independently tested filter-based and magnetic bead-based DNA extraction kits using tissues from chronic wasting disease (CWD)-positive and -negative white-tailed deer (WTD; Odocoileus virginianus), as well as prion-infected and control Syrian hamster (Mesocricetus auratus) brains. CFIA used two filter-based kits (one automated, one manual), while MNPRO tested two manual kits (filter- and magnetic bead-based). PrP^Sc seeding activity was measured in extracted DNA and source tissues using real-time quaking-induced conversion (RT-QuIC). MNPRO found substantial to almost perfect agreement between RT-QuIC seeding activity of DNA eluates from both extraction methods and that of the source WTD tissue homogenate. CFIA optimized RT-QuIC to a 30-hour runtime, achieving 74% sensitivity and 94% specificity in 88 archived WTD DNA samples. Both laboratories concluded that commercial DNA extraction kits do not eliminate PrP^Sc, enabling carry-over into DNA eluates. Until infectivity is resolved by animal bioassay, DNA from PrP^Sc-positive tissues should be handled under biosafety protocols appropriate for the originating prion disease, with decontamination and containment procedures.

Objectives: Reperfusion, an essential therapeutic strategy for salvaging ischemic myocardium in ischemic heart disease, paradoxically exacerbates myocardial injury. Ferroptosis is a pivotal mechanism underlying myocardial ischemia-reperfusion injury (MIRI). Nrf2 can regulate ferroptosis, which could undergo SUMOylation at lysine 110 (K110) and was subsequently de-SUMOylated by Senp1. This study aimed to determine whether Nrf2 de-SUMOylation could mitigate MIRI by inhibiting myocardial ferroptosis.

Methods: Nrf2 K110R mice, mimicking Nrf2 de-SUMOylation, were generated. Mice cardiac morphology and function were observed by hematoxylin-eosin staining (HE) and echocardiography under normal and MIRI conditions. Ferroptosis inhibitor liproxstatin-1 (Lip-1) was used to demonstrate ferroptosis participation in Nrf2 de-SUMOylation regulated MIRI. In vitro, SUMO1/sentrin-specific protease 1 Senp1 KO H9C2 cells were subjected to RSL₃-induced ferroptosis to explore underlying mechanism.

Results: Nrf2 K110R mice showed normal cardiac morphology and function at baseline. However, de-SUMOylation of Nrf2 alleviated myocardial ferroptosis, resulting in a reduction of MIRI severity in MIRI mice. The administration of Lip-1 attenuated the differences in MIRI between Nrf2 wild-type and K110R mice. Mechanistically, Nrf2 de-SUMOylation was associated with a reduction in Transferrin receptor (Tfr) expression level, thereby mitigating ferroptosis in cardiomyocytes.

Conclusion: This study highlighted the role of Nrf2 SUMOylation in promoting ferroptosis during MIRI and identified Nrf2 de-SUMOylation as a potential therapeutic target for MIRI.

Preeclampsia (PE) is a severe pregnancy complication with unclear molecular mechanisms. Our research investigated the effect of UNC5C-AS1 on human umbilical vein endothelial cell (HUVEC) function in PE. UNC5C-AS1 was downregulated in PE placentas. Upregulating UNC5C-AS1 promoted HUVEC migration, invasion, tube formation, and the expression of vascular permeability factors, while UNC5C-AS1 silencing exhibited an opposite effect. UNC5C-AS1 directly targeted the miR148a3p/EMP1 axis. MiR-148a-3p was up-regulated and EMP1 was downregulated in PE. The regulatory effects of UNC5C-AS1 overexpression on HUVEC functions were reversed by miR-148a-3p mimics, and this reversal was subsequently rescued by EMP1 upregulation. UNC5C-AS1 overexpression ameliorated tissue damage in the PE mouse model. UNC5C-AS1 alleviated the PE-associated injury and modulated HUVEC function by targeting miR-148a-3p/EMP1 axis.

Torovirus (ToV), while resembling coronavirus (CoV), belongs to a distinct family Tobaniviridae in the order Nidovirales. Porcine ToV (PToV) is widespread in pig populations across many countries, yet its potential pathogenicity in pigs remains poorly understood. The viral 3C-like protease (3CLP) plays a crucial role in processing viral polyproteins and manipulating the host antiviral immune response by targeting cellular proteins through its catalytic activity. In this study, we focused on PToV 3CLP due to its unique catalytic dyad characteristics and substrate recognition properties, which are distinct from those of CoV 3CLPs. We revealed that PToV 3CLP induces pyroptosis in porcine small intestinal IPEC-J2 cells and further demonstrated that porcine gasdermin D (pGSDMD) is a cleavage substrate for PToV 3CLP associated with this process. The catalytic residues, histidine 53 and serine 160, essential for the protease activity of PToV 3CLP, were required for the cleavage of pGSDMD at two distinct sites, glutamine 193 (Q193) and glutamine 277 (Q277). One of fragments produced by PToV 3CLP cleavage, pGSDMD_1-277, mimicked the activity of the N-terminal domain of pGSDMD (pGSDMD_1-279) in forming pores and ultimately triggering pyroptosis. Intriguingly, these results contrast with the inhibitory effect of CoV 3CLPs on pyroptosis, previously reported to target pGSDMD at the Q193 site. The study provides additional evidence of the distinct nature of 3CLP between ToV and CoV, which may partly explain the divergent clinical manifestations and pathogenesis observed in pigs infected by these nidoviruses.

Fowl adenoviruses (FAdVs) are widely distributed in poultry populations around the world, and many diseases are associated with FAdV infection in chickens. This study documented the first characterization of coinfection with fowl adenovirus serotypes 1 and 4 (FAdV-1 and -4) associated with hydropericardium hepatitis syndrome (HHS) in Chinese layer flocks, revealing a novel viral cooperation mechanism. Two novel strains (CH/SX/201805-1 and -4) were identified and isolated, with whole-genome sequencing showing CH/SX/201805-1 clustering with FAdV-1 (99.7% identity to FAdV-A-61/11z), whereas CH/SX/201805-4 displayed characteristic ORF19/27/29 deletions mirroring emergent Chinese FAdV-4 variants. Experimental coinfection in SPF chickens resulted in 87.5% mortality, which was 16.7% greater than that resulting from infection alone, with exacerbated pathology. In vitro coinfection experiments demonstrated concurrent viral replication within same LMH cells, a previously unreported phenomenon, where FAdV-1 increased FAdV-4 replication efficiency 21-fold (p < 0.001). Transcriptomic profiling revealed heat shock protein A2 (HSPA2) as the most differentially expressed gene, which was upregulated 2.8-fold during coinfection compared with infection with FAdV-4 alone. Functional validation through HSPA2 knockdown reduced FAdV-4 replication, establishing that FAdV-1 potentiates FAdV-4 through HSPA2-mediated host modulation. These findings provide the first evidence of HSPA2-dependent interserotype synergy in FAdV and can be used to develop a cellular model for FAdV coinfection studies. These insights redefine the understanding of FAdV pathogenesis and create new avenues for targeted intervention strategies against emerging FAdV coinfections.

Objectives: Tramadol, a clinically approved analgesic widely used for managing postoperative pain, has recently been shown to possess anticancer properties in several tumor models, especially in breast cancer. In this study, we explored the intricate molecular mechanisms by which tramadol induces cytotoxicity in breast cancer cell lines.

Methods: Two invasive ductal carcinoma lines MCF-7 and MDA-MB-231 were used to verify the molecular cytotoxicity of tramadol using cell viability analysis, flow cytometry analysis, real-time polymerase chain reaction, western blotting, Seahorse biogenetic, and transmission electron microscopy analyses.

Results: Our findings demonstrate that tramadol induces the normoxic stabilization and nuclear translocation of hypoxia-inducible factor- 1 alpha (HIF-1α) to activate hypoxia responsive genes. Concurrently, tramadol triggers endoplasmic reticulum (ER) stress and activates the p-eIF2α/ATF4/CHOP signaling axis, leading to the generation of reactive oxygen species, impaired autophagy, mitochondrial dysfunction, including mitochondrial membrane depolarization and the decline of ATP production, cytoplasmic vacuolization, and lipid droplet accumulation which is characteristics of paraptosis-like cell death. Notably, the knockout of HIF-1α or ATF4 significantly reduced tramadol-induced cytotoxicity, highlighting their crucial roles in mediating these cellular responses.

Conclusion: Tramadol induced breast cancer cell death via paraptosis which highlights its therapeutic potential in targeting resistant cancer subtypes such as triple-negative breast cancer.

Critical metabolic enzymes and pathways specific to bacterial adaptation in different host microenvironments directly contribute to bacterial pathogenicity. In this study, a virulent strain of the important zoonotic pathogen Streptococcus suis was found to show enhanced growth under anaerobic conditions compared to aerobic conditions. Transcriptomic analysis found a significant suppression of many central metabolic genes during anaerobic growth of S. suis. The transcriptomic data were used to reconstruct a genome-scale metabolic network to assess the distribution of metabolic fluxes in S. suis under different conditions. Significant activation of the arginine deiminase (ADI) and branched-chain amino acid (BCAA) biosynthesis pathways was identified. Gene deletion mutants of arcB and ilvC participating in these two pathways, respectively, were constructed. Compared to the wild-type strain, the ΔarcB mutant showed more significant growth deficiency under anaerobic conditions than under aerobic conditions. Accumulation of ATP and NH₃, the metabolites of the ADI pathway, was significantly higher when S. suis was cultured under anaerobic conditions, and this effect was attenuated in the ΔarcB mutant. The knockout of IlvC of the BCAA pathway disrupted the normal growth of S. suis in valine- and isoleucine-limited medium under anaerobic conditions. Both ΔarcB and ΔilvC showed attenuation in mice with decreased lethality, bacterial loads in tissues, and cytokine levels in serum, with the hypoxia-induced gene up-regulated in tissues. Therefore, ADI and BCAA pathways are critical for S. suis survival in response to hypoxia and infection in vivo, with ArcB and IlvC being promising drug targets.