British Journal of Pharmacology最新文献

Background and purpose: Novel heart failure (HF) pharmacotherapies, including angiotensin receptor-neprilysin inhibitor (ARNI) and sodium-glucose cotransporter 2 inhibitors (SGLT2is), may confer cardiovascular benefits by attenuating myocardial fibrosis. However, direct evidence from human failing myocardial samples is limited. Thus, we assessed the associations of ARNI and SGLT2i therapies with fibrosis on myocardial samples from advanced HF patients undergoing heart transplantation (HTX).

Experimental approach: Ninety-three patients receiving stable combined HF pharmacotherapy (uninterrupted use of β-blockers, mineralocorticoid receptor antagonists, angiotensin-converting enzyme inhibitors or ARNI, and optionally SGLT2is) without the need for pharmacological or mechanical circulatory support for at least three months pre-HTX, were enrolled. Standardized regions of the left ventricular anterior wall from explanted hearts were analysed. The primary outcome was the percentage of interstitial myocardial fibrosis area measured via histology. The expression of pro-fibrotic and pathological remodelling markers was quantified with qRT-PCR.

Key results: Both ARNI and SGLT2i therapies were associated with reduced interstitial collagen accumulation in adjusted analyses. None of the tested clinical parameters (sex, age, serum creatinine, presence of hypertension or diabetes) interacted with the effects of ARNI or SGLT2i on fibrosis. Additionally, no interaction was observed between ARNI and SGLT2i use regarding collagen content. Among the examined genes, ARNI was linked to a decreased beta-to-alpha myosin heavy chain expression ratio, while in SGLT2i-treated patients, reduced mRNA levels of matrix metalloproteinase 9 (MMP9) and tissue inhibitor of metalloproteinases 2 (TIMP2) were observed in adjusted models.

Conclusions and implications: ARNI and SGLT2i therapies might exert anti-fibrotic effects in advanced HF.

Background and purpose: Silicosis is a major cause of occupational disease-related morbidity and mortality worldwide, yet effective pharmacological treatments remain limited. Nerandomilast, a novel inhibitor of phosphodiesterase-4B (PDE4B), has demonstrated anti-fibrotic potential in idiopathic pulmonary fibrosis (IPF); however, its efficacy and mechanisms in silicosis have not been investigated.

Experimental approach: The therapeutic effects (and their underlying mechanisms) of PDE4B inhibition was evaluated in in models of silicosis, both in vivo (male C57BL/6N mice) and in vitro (THP-1 macrophages and MRC-5 cells). Single-cell RNA sequencing, using lung tissue, was first performed to identify PDE4B as a key regulatory target in the pathogenesis of silicosis. Based on this finding, the therapeutic effects and underlying mechanisms of PDE4B inhibition were assessed using nerandomilast. Pulmonary function tests, inflammatory marker analyses and fibrosis evaluations were conducted to determine treatment efficacy. In addition, bulk RNA sequencing and transcriptomic analyses were performed to explore the molecular pathways modulated by nerandomilast.

Key results: PDE4B inhibition effectively prevented and attenuated silica-induced lung inflammation in the mouse model by suppressing both canonical and non-canonical NLRP3 inflammasome pathways in lung macrophages. Furthermore, PDE4B inhibition down-regulated TGF-β/Smad signalling in lung fibroblasts of silicosis, leading to a significant reduction in fibrosis-related gene expression.

Conclusions and implications: These findings suggest that nerandomilast, a PDE4B inhibitor, may be a promising treatment for silicosis, which currently lacks effective therapies.

Background and purpose: Neutrophil extracellular traps (NETs) play a key role in the local and systemic pathologies induced by Echis carinatus and Naja naja venoms. This study aimed to identify the toxin/s responsible for N. naja venom-induced NET formation.

Experimental approach: N. naja venom was subjected to molecular sieving to identify the toxins responsible for NET formation. The venom was fractionated, and the third fraction, cytotoxin fraction (CTXF), was found to induce vital NET formation. Various in vivo and in vitro experiments were conducted to comprehend the effects and mechanisms involved, including histochemical and immunohistochemical studies. To draw appropriate conclusions, CTXF was comparatively characterised along with purified cardiotoxin (Np-CTX), neurotoxin (Nn-NTX), and phospholipase A₂ (VRV-PL-XII).

Key results: CTXF is a non-enzymatic cytotoxin fraction. Peptide analysis revealed that CTXF is a mixture of four cytotoxins/cardiotoxins of molecular mass ranging from 6668 to 6778 Da, and they belong to the 3FTx family. In vivo studies demonstrated that CTXF damaged cardiac muscle extensively, where CTXF and H3Cit specifically localised in the cardiac tissue. CTXF and Np-CTX induced the formation of NETs, whereas Nn-NTX and VRV-PL-XII did not. NET formation was associated with increased intracellular Ca²⁺ flux via the store-operated Ca²⁺ entry mechanism, accompanied by alterations in actin dynamics.

Conclusions and implications: This study identified N. naja venom cytotoxins/cardiotoxins of the 3FTx family as key inducers of NETs. The NET-forming mechanism involves a cytosolic Ca²⁺ spike, resulting in cytoskeletal rearrangements, highlighting potential therapeutic targets for mitigating venom-induced pathologies.

Background and purpose: Oxidative stress induces a rerouting of metabolic flux from glycolysis to the pentose phosphate pathway. One proposed mechanism involves negative feedback via tonic inhibition of glucose-6-phosphate dehydrogenase by NADPH. However, recent evidence shows that NADPH levels do not decrease 5-s after hydrogen peroxide treatment. This finding is inconsistent with the canonical model wherein a feedback inhibition loop is modulated by NADPH depletion. This inconsistency prompted us to test the involvement of feedback inhibition at high temporal resolution.

Experimental approach: We employed genetically encoded fluorescent indicators for hydrogen peroxide (HyPerRed) and NADPH (iNap1) expressed in human embryonic kidney HEK293 cells. These tools enabled simultaneous real-time, single-cell monitoring of NADPH and hydrogen peroxide.

Key results: Glucose sustained NADPH levels under acute oxidative stress in the first seconds following hydrogen peroxide exposure. This result contradicts the reported feedback inhibition, which is considered one of the fundamental mechanisms to explain the acute rerouting of glycolysis to the pentose phosphate pathway. Furthermore, pharmacological inhibition of G6PDH suggests that the pentose phosphate pathway is the primary source of cytosolic NADPH under oxidative stress. Monitoring NADPH levels following G6PDH inhibition allowed assessment of the NADPH consumption flux, a parameter that rises markedly under oxidative stress.

Conclusion and implications: Our results support an anticipatory phenomenon that maintains NADPH levels under acute hydrogen peroxide exposure, thereby discarding the proposed feedback inhibition loop. This work offers a new perspective on the regulatory nuances of a metabolic pathway involved in ageing, cancer and many other pathological conditions.

Background and purpose: Chronic pain is a widespread and debilitating condition with limited treatment options. While the ventral tegmental area (VTA) and posterior hypothalamic nucleus (PH) have been independently implicated in pain modulation, the specific neural circuitry linking them remains unclear. This study aimed to investigate the role of the PH → VTA glutamatergic projection in chronic pain regulation.

Experimental approach: Using a mouse model of neuropathic pain induced by chronic constriction injury (CCI), we combined viral tracing, immunofluorescence, in vitro electrophysiology, optogenetics, chemogenetics and fibre photometry. Behavioural assays were used to assess mechanical allodynia and thermal hyperalgesia following circuit manipulation.

Key results: We identified a glutamatergic projection from PH to VTA dopamine (DA) neurons that modulates pain behaviour. Optogenetic activation of the PH → VTA circuit in naïve mice induced pain-like hypersensitivity. Conversely, inhibition of this pathway reduced pain-related behaviours in CCI mice. Mechanistically, PH activation increased glutamatergic input onto VTA DA neurons, enhancing dopamine release in the nucleus accumbens (NAc). Fibre photometry confirmed increased activity in this circuit during nociceptive states.

Conclusions and implications: Our results define a functional PH^Glu → VTA^DA → NAc circuit that contributes to chronic pain processing. Hyperactivity in this pathway facilitates nociceptive behaviours, while its inhibition exerts analgesic effects. Modulating this circuit may offer new therapeutic strategies for treating chronic pain.

Background and purpose: Ventricular arrhythmias are a leading cause of death among patients with cardiovascular diseases and are associated with elevated levels of catecholamines. Mitochondrial Ca²⁺ transport is essential for initiating an adrenergic response. However, continuous stimulation might lead to mitochondrial Ca²⁺ overload and dysfunction within cardiac tissue. This study investigates the role of mitochondrial Ca²⁺ in lethal arrhythmogenesis and the effects of its modulation.

Experimental approach: Male C57BL/6 mice were administered either Ru360 (oxo-bridged dinuclear ruthenium ammine complex) a potent and selective mitochondrial Ca²⁺ transport inhibitor, or normal saline via intravenous injection. A baseline electrocardiogram (ECG) was recorded, followed by subcutaneous administration of isoprenaline. The ECG was monitored for an additional 20 min, after which cardiomyocytes and mitochondria were isolated for further characterization studies.

Key results: Isoprenaline administration led to ventricular tachycardia and fibrillation, but Ru360 pretreatment successfully prevented these arrhythmias. Mitochondria from isoprenaline-treated hearts showed higher Ca²⁺ content, indicating overload that compromised mitochondrial function and membrane integrity, evidenced by decreased respiratory control, reduced Ca²⁺ retention capacity and diminished membrane potential. Isoprenaline also increased oxidative stress, illustrated by elevated peroxide production, electron leak and acute oxidative modifications, and erratic cellular Ca²⁺ dynamics. This mitochondrial dysfunction correlated with a decreased respirasome activity, but not a difference in respirasome abundance quantified by complexome profiling, which was prevented by Ru360 pretreatment.

Conclusion: Mitochondrial Ca²⁺ overload significantly contributes to arrhythmias by disrupting respirasome function and increasing oxidative stress, impairing cellular Ca²⁺ dynamics. Modulating mitochondrial Ca²⁺ transport might be a promising strategy for developing innovative antiarrhythmic therapies.

Background and purpose: Drug repurposing (DR) presents a compelling alternative to traditional drug discovery, offering lower risk and cost by applying approved drugs to new indications. Computational methods play a vital role in early-stage drug repurposing and the development of robust computational workflows can accelerate antibiotic discovery.

Experimental approach: We established a novel computational workflow comprising three key steps: target screening (based on CEG 2.0 database), drug screening (utilizing DrugBank database, antiBac-Pred database, molecular docking and molecular dynamics [MD] simulation) and in vitro antibacterial experiments.

Key results: Our workflow identified numerous commercially available drugs predicted to target the bacterial chaperone GroEL. Antibacterial assays revealed that both daprodustat and ezetimibe exhibited efficacy against Staphylococcus aureus and Escherichia coli ΔtolC. Notably, the efflux pump inhibitor PAβN enhanced the antibacterial efficacy of daprodustat against both S. aureus and E. coli, while potentiating the antibacterial potency of ezetimibe specifically against S. aureus. MD simulations confirmed stable binding of both drugs to S. aureus or E. coli GroEL, aligning with the antibacterial results.

Conclusion and implications: This study validated our computational workflow for repurposing non-antibacterial drugs as antibacterial agents, demonstrating that cost-effective, computer-aided drug repurposing is a feasible strategy for identifying new therapeutic approaches to diseases, such as cancer, diabetes and COVID-19. Furthermore, the synergistic effect of daprodustat combined with an efflux pump inhibitor (e.g. PAβN) represents a promising therapeutic approach against both Gram-positive and Gram-negative bacterial pathogens.