ART duration and immunometabolic state determine efficacy of DC-based treatment restoring functional HIV- specific CD8+ T cells in Chronic PLWH.

Dysfunction of CD8+ T cells in people living with HIV-1 (PLWH) receiving anti-retroviral therapy (ART) has restricted the efficacy of dendritic cell (DC)-based immunotherapies against HIV-1. Heterogeneous immune exhaustion and metabolic states of CD8+ T cells might differentially associate with dysfunction. However, specific parameters associated to functional restoration of CD8+ T cells after DC treatment have not been investigated in detail. Here, we studied the association of ART duration with memory subsets, exhaustion and metabolic profiles of CD8+ T cells from PLWH and improvement of polyfunctional and effector HIV-1 specific responses after stimulation with Gag-adjuvant-primed DC. HIV-1-specific CD8+ T cell responses from a larger proportion PLWH on ART for more than 10 years (LT-ARTp) improved polyfunctionality and capacity to eliminate autologous p24+ infected CD4+ T cells in vitro. In contrast, CD8+ T cells from PLWH on ART for less than a decade (ST-ARTp) were less responsive to DC treatment and functional improvement was limited in this group. This was associated with lower frequencies of central memory CD8+ T cells, increased co-expression of PD1 and TIGIT and reduced mitochondrial respiration and glycolytic induction upon TCR activation. In contrast, CD8+ T cells from LT-ARTp showed increased frequencies of TIM3+PD1- cells and preserved induction of glycolysis. Treatment of dysfunctional CD8+ T cells from ST-ARTp with combined anti-PD1 and anti-TIGIT antibodies plus a glycolysis promoting drug restored their ability to eliminate infected CD4+ T cells. Together, our study identifies specific immunometabolic parameters for different PLWH subgroups potentially useful for future personalized DC-based HIV-1 vaccines.

Placental Pathology in Maternal Ornithine Transcarbamylase Deficiency

Introduction Ornithine transcarbamylase (OTC) deficiency is the most common urea cycle disorder, inherited in an X-linked manner. Males are severely affected. Female phenotypes vary from asymptomatic to severe, and symptoms may be triggered by high metabolic states like childbirth. Literature on OTC deficiency in pregnancy and placental pathology is limited. Methods Pathology records were searched at a single referral center from 2000–2020 and identified three placental cases from two mothers heterozygous for OTC deficiency. Placental pathology and maternal and neonatal history were reviewed in detail. Results The placenta from one symptomatic mother carrying an affected male fetus showed widespread high-grade fetal vascular malperfusion (FVM) lesions of varying age. These lesions were not seen in the two placentas from the asymptomatic mother. Discussion In cases of symptomatic maternal OTC deficiency, our findings highlight the need for placental examination. Since thrombotic events in the placenta have the potential to associate with fetal and neonatal endothelial damage, a high index of suspicion for neonatal thrombosis may be warranted.

Transcriptomes and metabolism define mouse and human MAIT cell heterogeneity

Mucosal-associated invariant T (MAIT) cells are a subpopulation of T lymphocytes that respond to microbial metabolites. We performed single-cell RNA sequencing and metabolic analyses of MAIT cell subsets in thymus and peripheral tissues from mice and humans to define the heterogeneity and developmental pathway of these innate-like lymphocytes. We show that the predominant mouse subset, which produces IL-17 (MAIT17), and the subset that produces IFNγ (MAIT1), have greatly different transcriptomes and metabolic states in the thymus and periphery. A splenic MAIT subset has a transcriptome similar to circulating lymphocytes, and in mice these also are found in recent thymic emigrants, suggesting partially mature cells emigrate from the thymus. Human MAIT cells are predominantly MAIT1 cells, but have a different metabolism from their mouse counterparts with increased fatty acid uptake and storage. Although mouse and human subsets are similar in thymus, in the periphery they diverge, likely reflecting environmental influences.

Single-cell resolution unravels spatial alterations in metabolism, transcriptome and epigenome of ageing liver

Epigenetic ageing clocks have revealed that tissues within an organism can age with different velocity. However, it has not been explored whether cells of one type experience different ageing trajectories within a tissue depending on their location. Here, we employed lipidomics, spatial transcriptomics and single-cell ATAC-seq in conjunction with available single-cell RNA-seq data to address how cells in the murine liver are affected by age-related changes of the microenvironment. Integration of the datasets revealed zonation-specific and age-related changes in metabolic states, the epigenome and transcriptome. Particularly periportal hepatocytes were characterized by decreased mitochondrial function and strong alterations in the epigenetic landscape, while pericentral hepatocytes,despite accumulation of large lipid droplets, did not show apparent functional differences. In general, chromatin alterations did not correlate well with transcriptional changes, hinting at post-transcriptional processes that shape gene expression during ageing. Together, we provide evidence that changing microenvironments within a tissue exert strong influences on their resident cells that can shape epigenetic, metabolic and phenotypic outputs.

Metabolic properties of murine kidney mitochondria

We show that mitochondria from the kidney of mice (MKM), rat brain (RBM), and heart (RHM) oxidize long-chain fatty acids at high rates in all metabolic states only in the presence of any other mitochondrial metabolites: succinate, glutamate, or pyruvate. All supporting substrates increased several folds the respiration rates in State 4 and State 3. The stimulations of the State 3 respiration with palmitoyl-carnitine + malate oxidation (100%) were: with succinate in MKM 340%, RBM 370%, and RHM 340%; with glutamate - MKM 200%, RBM 270%, and RHM 270%; and with pyruvate - MKM 150%, RBM 260%, and RHM 280%. The increases in O2 consumption in State 4 were due to increased leakage of electrons to produce superoxide radicals (O2•). Earlier, we have shown that the brain and heart mitochondria possess a strong intrinsic inhibition of succinate oxidation to prevent the excessive O2• production at diminished functional loads. We show that kidney mitochondria lack the intrinsic inhibition of SDH. The new methodology to study β-oxidation of LCFAs opens the opportunity to study energy metabolism under normal and pathological conditions, particularly in the organs that utilize LCFAs as the main energy source.

Leptin: A Metabolic Signal for the Differentiation of Pituitary Cells

Pituitary cell function is impacted by metabolic states and therefore must receive signals that inform them about nutritional status or adiposity. A primary signal from adipocytes is leptin, which recent studies have shown regulates most pituitary cell types. Subsets of all pituitary cell types express leptin receptors and leptin has been shown to exert transcriptional control through classical JAK/STAT pathways. Recent studies show that leptin also signals through post-transcriptional pathways that involve the translational regulatory protein Musashi. Mechanistically, post-transcriptional control would permit rapid cellular regulation of critical pre-existing pituitary transcripts as energy states change. The chapter will review evidence for transcriptional and/or post-transcriptional regulation of leptin targets (including Gnrhr, activin, Fshb, Gh, Ghrhr, and Pou11f1) and the consequences of the loss of leptin signaling to gonadotrope and somatotrope functions.

Mitochondrial DNA metabolism is coupled with 20S proteasome function via regulation of deoxyribonucleotide homeostasis in Saccharomyces cerevisiae

The synthesis of mitochondrial DNA (mtDNA) is not coupled with cell cycle. Previous studies have shown that the size of deoxyribonucleoside triphosphate (dNTP) pools plays an important role in regulating mtDNA replication and amplification. In yeast, dNTPs are synthesized by the cytosolic ribonucleotide reductase (RNR). It is currently poorly understood as to how RNR activity is regulated in non-dividing or quiescent cells to finely tune mtDNA metabolism to cope with different metabolic states. Here, we show that defect in the 20S proteasome drastically destabilizes mtDNA. The mtDNA instability phenotype in 20S proteasome mutants is suppressed by overexpression of RNR3 or by the deletion of SML1, encoding a minor catalytic subunit and an intrinsic inhibitor of RNR respectively. We found that Sml1 is stabilized in the 20S proteasomal mutants, suggesting that 20S affects mtDNA stability by stabilizing Sml1. Interestingly, defect in the regulatory 19S proteasomal function has only subtle effect on mtDNA stability, supporting a role of the 20S proteasome in dNTP homeostasis independent of 19S. Finally, we found that when cells are transitioned from glycolytic to oxidative growth, Sml1 level is reduced in a 20S-dependent manner. In summary, our study establishes a link between cellular proteostasis and mtDNA metabolism through the regulation of dNTP homeostasis. We propose that increased degradation of Sml1 by the 20S proteasome under respiratory conditions provides a mechanism to stimulate dNTP synthesis and promote mtDNA amplification.

Mitochondrial TFAM as a Signaling Regulator between Cellular Organelles: A Perspective on Metabolic Diseases

Tissues actively involved in energy metabolism are more likely to face metabolic challenges from bioenergetic substrates and are susceptible to mitochondrial dysfunction, leading to metabolic diseases. The mitochondria receive signals regarding the metabolic states in cells and transmit them to the nucleus or endoplasmic reticulum (ER) using calcium (Ca2+) for appropriate responses. Overflux of Ca2+ in the mitochondria or dysregulation of the signaling to the nucleus and ER could increase the incidence of metabolic diseases including insulin resistance and type 2 diabetes mellitus. Mitochondrial transcription factor A (Tfam) may regulate Ca2+ flux via changing the mitochondrial membrane potential and signals to other organelles such as the nucleus and ER. Since Tfam is involved in metabolic function in the mitochondria, here, we discuss the contribution of Tfam in coordinating mitochondria-ER activities for Ca2+ flux and describe the mechanisms by which Tfam affects mitochondrial Ca2+ flux in response to metabolic challenges.

Reversion of antibiotic resistance in multidrug-resistant pathogens using non-antibiotic pharmaceutical benzydamine

Antimicrobial resistance has been a growing concern that gradually undermines our tradition treatment regimens. The fact that few antibacterial drugs with new scaffolds or targets have been approved in the past two decades aggravates this crisis. Repurposing drugs as potent antibiotic adjuvants offers a cost-effective strategy to mitigate the development of resistance and tackle the increasing infections by multidrug-resistant (MDR) bacteria. Herein, we found that benzydamine, a widely used non‐steroidal anti‐inflammatory drug in clinic, remarkably potentiated broad-spectrum antibiotic-tetracyclines activity against a panel of clinically important pathogens, including MRSA, VRE, MCRPEC and tet(X)-positive Gram-negative bacteria. Mechanistic studies showed that benzydamine dissipated membrane potential (▵Ψ) in both Gram-positive and Gram-negative bacteria, which in turn upregulated the transmembrane proton gradient (▵pH) and promoted the uptake of tetracyclines. Additionally, benzydamine exacerbated the oxidative stress by triggering the production of ROS and suppressing GAD system-mediated oxidative defensive. This mode of action explains the great bactericidal activity of the doxycycline-benzydamine combination against different metabolic states of bacteria involve persister cells. As a proof-of-concept, the in vivo efficacy of this drug combination was evidenced in multiple animal infection models. These findings indicate that benzydamine is a potential tetracyclines adjuvant to address life-threatening infections by MDR bacteria.

Recognition of acetylated histone by Yaf9 regulates metabolic cycling of transcription initiation and chromatin regulatory factors

How transcription programs rapidly adjust to changing metabolic and cellular cues remains poorly defined. Here, we reveal a function for the Yaf9 component of the SWR1-C and NuA4 chromatin regulatory complexes in maintaining timely transcription of metabolic genes across the yeast metabolic cycle (YMC). By reading histone acetylation during the oxidative and respiratory phase of the YMC, Yaf9 recruits SWR1-C and NuA4 complexes to deposit H2A.Z and acetylate H4, respectively. Increased H2A.Z and H4 acetylation during the oxidative phase promotes transcriptional initiation and chromatin machinery occupancy and is associated with reduced RNA polymerase II levels at genes—a pattern reversed during transition from oxidative to reductive metabolism. Prevention of Yaf9-H3 acetyl reading disrupted this pattern of transcriptional and chromatin regulator recruitment and impaired the timely transcription of metabolic genes. Together, these findings reveal that Yaf9 contributes to a dynamic chromatin and transcription initiation factor signature that is necessary for the proper regulation of metabolic gene transcription during the YMC. They also suggest that unique regulatory mechanisms of transcription exist at distinct metabolic states.