Regulation Of Oxidative Stress Response Research Articles

Regulation of oxidative stress response and antioxidant modification in Corynebacterium glutamicum.

As an efficient and safe industrial bacterium, Corynebacterium glutamicum has extensive application in amino acid production. However, it often faces oxidative stress induced by reactive oxygen species (ROS), leading to diminished production efficiency. To enhance the robustness of C. glutamicum, numerous studies have focused on elucidating its regulatory mechanisms under various stress conditions such as heat, acid, and sulfur stress. However, a comprehensive review of its defense mechanisms against oxidative stress is needed. This review offers an in-depth overview of the mechanisms C. glutamicum employs to manage oxidative stress. It covers both enzymatic and non-enzymatic systems, including antioxidant enzymes, regulatory protein families, sigma factors involved in transcription, and physiological redox reduction pathways. This review provides insights for advancing research on the antioxidant mechanisms of C. glutamicum and sheds light on its potential applications in industrial production.

Neem Leaf Extract Exhibits Anti-Aging and Antioxidant Effects from Yeast to Human Cells.

Neem leaves have long been used in traditional medicine for promoting longevity. However, the precise mechanisms underlying their anti-aging effects remain elusive. In this study, we investigated the impact of neem leaf extract (NLE) extracted from a 50% ethanol solution on the chronological lifespan of Saccharomyces cerevisiae, revealing an extension in lifespan, heightened oxidative stress resistance, and a reduction in reactive oxygen species. To discern the active compounds in NLE, LC/MS and the GNPS platform were employed. The majority of identified active compounds were found to be flavonoids. Subsequently, compound-target pharmacological networks were constructed using the STP and STITCH platforms for both S. cerevisiae and Homo sapiens. GOMF and KEGG enrichment analyses of the predicted targets revealed that "oxidoreductase activity" was among the top enriched terms in both yeast and human cells. These suggested a potential regulation of oxidative stress response (OSR) by NLE. RNA-seq analysis of NLE-treated yeast corroborated the anti-oxidative effect, with "oxidoreductase activity" and "oxidation-reduction process" ranking high in enriched GO terms. Notably, CTT1, encoding catalase, emerged as the most significantly up-regulated gene within the "oxidoreductase activity" cluster. In a ctt1 null mutant, the enhanced oxidative stress resistance and extended lifespan induced by NLE were nullified. For human cells, NLE pretreatment demonstrated a decrease in reactive oxygen species levels and senescence-associated β-galactosidase activity in HeLa cells, indicative of anti-aging and anti-oxidative effects. This study unveils the anti-aging and anti-oxidative properties of NLE while delving into their mechanisms, providing novel insights for pharmacological interventions in aging using phytochemicals.

SdrR, a LysR-Type Regulator, responds to the mycobacterial antioxidant defense.

Protection against oxidative stress is a vital defense mechanism for Mycobacterium tuberculosis within the host. However, few transcription factors that control bacterial antioxidant defense are known. Here, we present evidence that SdrR, encoded by the MSMEG_5712 (Ms5712) gene, functions as an oxidative stress response regulator in Mycobacterium smegmatis. SdrR recognizes an 11-bp motif sequence in the operon's upstream regulatory region and negatively regulates the expression of short-chain dehydrogenases/reductases (SDR). Overexpressing sdrR inhibited SDR expression, which rendered the strain oxidative more stress-sensitive. Conversely, sdrR knockout alleviates SDR repression, which increases its oxidative stress tolerance. Thus, SdrR responds to oxidative stress by negatively regulating sdr expression. Therefore, this study elucidated an underlying regulatory mechanism behind mycobacterial oxidative stress adaptation.

Loss of macrophage TSC1 exacerbates sterile inflammatory liver injury through inhibiting the AKT/MST1/NRF2 signaling pathway

Tuberous sclerosis complex 1 (TSC1) plays important roles in regulating innate immunity. However, the precise role of TSC1 in macrophages in the regulation of oxidative stress response and hepatic inflammation in liver ischemia/reperfusion injury (I/R) remains unknown. In a mouse model of liver I/R injury, deletion of myeloid-specific TSC1 inhibited AKT and MST1 phosphorylation, and decreased NRF2 accumulation, whereas activated TLR4/NF-κB pathway, leading to increased hepatic inflammation. Adoptive transfer of AKT- or MST1-overexpressing macrophages, or Keap1 disruption in myeloid-specific TSC1-knockout mice promoted NRF2 activation but reduced TLR4 activity and mitigated I/R-induced liver inflammation. Mechanistically, TSC1 in macrophages promoted AKT and MST1 phosphorylation, and protected NRF2 from Keap1-mediated ubiquitination. Furthermore, overexpression AKT or MST1 in TSC1-knockout macrophages upregulated NRF2 expression, downregulated TLR4/NF-κB, resulting in reduced inflammatory factors, ROS and inflammatory cytokine-mediated hepatocyte apoptosis. Strikingly, TSC1 induction in NRF2-deficient macrophages failed to reverse the TLR4/NF-κB activity and production of pro-inflammatory factors. Conclusions: Macrophage TSC1 promoted the activation of the AKT/MST1 signaling pathway, increased NRF2 levels via reducing Keap1-mediated ubiquitination, and modulated oxidative stress-driven inflammatory responses in liver I/R injury. Our findings underscore the critical role of macrophage TSC1 as a novel regulator of innate immunity and imply the therapeutic potential for the treatment of sterile liver inflammation in transplant recipients.Schematic illustration of macrophage TSC1-mediated AKT/MST1/NRF2 signaling pathway in I/R-triggered liver inflammation. Macrophage TSC1 can be activated in I/R-stressed livers. TSC1 activation promotes phosphorylation of AKT and MST1, which in turn increases NRF2 expression and inhibits ROS production and TLR4/NF-κB activation, resulting in reduced hepatocellular apoptosis in I/R-triggered liver injury.

ATM inhibition blocks glucose metabolism and amplifies the sensitivity of resistant lung cancer cell lines to oncogene driver inhibitors

BackgroundATM is a multifunctional serine/threonine kinase that in addition to its well-established role in DNA repair mechanisms is involved in a number of signaling pathways including regulation of oxidative stress response and metabolic diversion of glucose through the pentose phosphate pathway. Oncogene-driven tumorigenesis often implies the metabolic switch from oxidative phosphorylation to glycolysis which provides metabolic intermediates to sustain cell proliferation. The aim of our study is to elucidate the role of ATM in the regulation of glucose metabolism in oncogene-driven cancer cells and to test whether ATM may be a suitable target for anticancer therapy.MethodsTwo oncogene-driven NSCLC cell lines, namely H1975 and H1993 cells, were treated with ATM inhibitor, KU55933, alone or in combination with oncogene driver inhibitors, WZ4002 or crizotinib. Key glycolytic enzymes, mitochondrial complex subunits (OXPHOS), cyclin D1, and apoptotic markers were analyzed by Western blotting. Drug-induced toxicity was assessed by MTS assay using stand-alone or combined treatment with KU55933 and driver inhibitors. Glucose consumption, pyruvate, citrate, and succinate levels were also analyzed in response to KU55933 treatment. Both cell lines were transfected with ATM-targeted siRNA or non-targeting siRNA and then exposed to treatment with driver inhibitors.ResultsATM inhibition deregulates and inhibits glucose metabolism by reducing HKII, p-PKM2Tyr105, p-PKM2Ser37, E1α subunit of pyruvate dehydrogenase complex, and all subunits of mitochondrial complexes except ATP synthase. Accordingly, glucose uptake and pyruvate concentrations were reduced in response to ATM inhibition, whereas citrate and succinate levels were increased in both cell lines indicating the supply of alternative metabolic substrates. Silencing of ATM resulted in similar changes in glycolytic cascade and OXPHOS levels. Furthermore, the driver inhibitors amplified the effects of ATM downregulation on glucose metabolism, and the combined treatment with ATM inhibitors enhanced the cytotoxic effect of driver inhibitors alone by increasing the apoptotic response.ConclusionsInhibition of ATM reduced both glycolytic enzymes and OXPHOS levels in oncogene-driven cancer cells and enhanced apoptosis induced by driver inhibitors thus highlighting the possibility to use ATM and the driver inhibitors in combined regimens of anticancer therapy in vivo.

Plastrum testudinis Ameliorates Oxidative Stress in Nucleus Pulposus Cells via Downregulating the TNF-α Signaling Pathway.

BackgroundPlastrum testudinis (PT), a widely used traditional Chinese medicine, exerts protective effects against bone diseases such as intervertebral disc degeneration (IDD). Despite its effectiveness, the molecular mechanisms underlying the effects of PT on IDD remain unclear. Methods In this study, we used a comprehensive strategy combining bioinformatic analysis with experimental verification to investigate the possible molecular mechanisms of PT against IDD. We retrieved targets for PT and IDD, and then used their overlapped targets for protein-protein interaction (PPI) analysis. In addition, we used Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses to investigate the anti-IDD mechanisms of PT. Moreover, in vivo and in vitro experiment validations including hematoxylin-eosin (HE) and safranine O-green staining, senescence-associated β-galactosidase (SA-β-gal) assay, cell immunofluorescence staining, intracellular ROS measurement and Western blot analysis were performed to verify bioinformatics findings. Results We identified 342 and 872 PT- and IDD-related targets (32 overlapping targets). GO enrichment analysis yielded 450 terms related to oxidative stress and inflammatory response regulation. KEGG analysis identified 48 signaling pathways, 10 of which were significant; the TNF-α signaling pathway had the highest p-value, and prostaglandin G/H synthase 2 (PTGS2), endothelin-1 (EDN1), TNF-α, JUN and FOS were enriched in this pathway. Histopathological results and safranin O/green staining demonstrated that PT attenuated IDD, and SA-β-gal assay showed that PT ameliorated nucleus pulposus cell (NPC) senescence. An ROS probe was adopted to confirm the protective effect of PT against oxidative stress. Western blot analyses confirmed that PT downregulated the protein expression of PTGS2, EDN1, TNF-α, JUN and FOS in the TNF-α signaling pathway as well as cellular senescence marker p16, proinflammatory cytokine interleukin-6 (IL6), while PT upregulated the expression of NPC-specific markers including COL2A1 and ACAN in a concentration-dependent manner. Conclusions To the best of our knowledge, this study is the first to report that PT alleviates IDD by downregulating the protein expression of PTGS2, EDN1, TNF-α, JUN and FOS in the TNF-α signaling pathway and upregulating that of COL2A1 and ACAN, thus suppressing inflammatory responses and oxidative stress in NPCs.

Relationship between long non-coding RNA MALAT1 and HOTAIR expression with sperm parameters, DNA and malondialdehyde levels in male infertility

BackgroundSperm quality is a complex index used to evaluate the fertility potential of men. The long non-coding RNA (lncRNA) MALAT1 participate in sperm development and HOTAIR have critical roles in the regulation of oxidative stress responses. This study aimed to evaluate the relationship of lncRNA MALAT1 and HOTAIR expression with sperm parameters, DNA fragmentation and malondialdehyde (MDA)levels in sperm fertility. MethodsIn this experimental study, semen samples (n = 30 fertile, n = 30 infertile) men were collected and evaluated for sperm parameters by computer-aided sperm analysis(CASA). Sperm DNA integrity quality was assessed by the Acridine orange(AO) test. MDA levels were determined by the Thiobarbituric acid reaction method. The expression of MALAT1 and HOTAIR was detected by RT-PCR. ResultsWe observed a decreased level of MALAT1and HOTAIR expression in the infertile men (p < 0.001). The relative expression level of MALAT1and HOTAIR showed a positive correlation with motility and morphology (p < 0.001). Subsequently, we found the DNA damage and MDA levels was negatively correlated with expression level of genes of sperm (p < 0.001). ConclusionIn this study the low expression of MALATI and HOTAIR resulted in the high level of MDA, DNA damage, and reduced motility of sperm. This study suggests the therapeutic opportunities in respect to MALATI and HOTAIR expression in the sperm function.

ChIP-seq identifies McSLC35E2 as a novel target gene of McNrf2 in Mytilus coruscus, highlighting its role in the regulation of oxidative stress response in marine mollusks.

NF-E2-related factor 2 (Nrf2) plays a crucial role in the oxidative regulatory process, which could trigger hundreds of antioxidant elements to confront xenobiotics. In the previous study, we identified Nrf2 from the marine mussel Mytilus coruscus, and the findings demonstrated that McNrf2 effectively protected the mussels against oxidative stress induced by benzopyrene (Bap). In order to delve deeper into the underlying mechanism, we utilized Chromatin Immunoprecipitation followed by sequencing (ChIP-seq) technology to systematically identify potential novel target genes of McNrf2. A total of 3,465 potential target genes were screened, of which 219 owned binding sites located within the promoter region. During subsequent experimental verification, it was found that McSLC35E2, a candidate target gene of McNrf2, exhibited negative regulation by McNrf2, as confirmed through dual luciferase and qRT-PCR detection. Further, the enzyme activity tests demonstrated that McNrf2 could counteract Bap induced oxidative stress by inhibiting McSLC35E2. The current study provides valuable insights into the application of ChIP-seq technology in the research of marine mollusks, advancing our understanding of the key role of Nrf2 in antioxidant defense mechanisms, and highlighting the significance of SLC35E2 in the highly sophisticated regulation of oxidative stress response in marine invertebrates.

Tolfenpyrad displays Francisella-targeted antibiotic activity that requires an oxidative stress response regulator for sensitivity.

Francisella species are highly pathogenic bacteria that pose a threat to global health security. These bacteria can be made resistant to antibiotics through facile methods, and we lack a safe and protective vaccine. Given their history of development as bioweapons, new treatment options must be developed to bolster public health preparedness. Here, we report that tolfenpyrad, a pesticide that is currently in use worldwide, effectively inhibits the growth of Francisella. This drug has an extensive history of use and a plethora of safety and toxicity data, making it a good candidate for development as an antibiotic. We identified mutations in Francisella novicida that confer resistance to tolfenpyrad and characterized a transcriptional regulator that is required for sensitivity to both tolfenpyrad and reactive oxygen species.

PeAP1-mediated oxidative stress response plays an important role in the growth and pathogenicity of Penicillium expansum.

Penicillium expansum is the causal agent of post-harvest blue mold in various fruits and serves as a model for understanding fungal pathogenicity and mycotoxin production. The relevance of oxidative stress response in the growth and virulence of P. expansum has been largely unexplored. Here, we identify the transcriptional factor PeAP1 as a regulator of oxidative stress response in P. expansum. Gene expression and protein abundance of PeAP1, as well as its nuclear localization, are specifically induced by H2O2. Deletion of PeAP1 results in increased sensitivity to H2O2, and PeAP1 mutants exhibit a variety of defects in hyphal growth and virulence. PeAP1 prevents the accumulation of both intracellular H2O2 during vegetative growth and host-derived H2O2 during biotrophic growth. Application of an antioxidant glutathione and a NADPH oxidase inhibitor, diphenylene iodonium, to the PeAP1 mutant partially restored fungal growth and virulence. RNA sequencing analysis revealed 144 H2O2-induced PeAP1 target genes, including four antioxidant-related genes, PeGST1, PePrx1, PePrx2, and PeTRX2, that were also demonstrated to be involved in oxidative stress response and/or virulence. Collectively, our results demonstrate the global regulatory role of PeAP1 in response to oxidative stress and provide insights into the critical role of the PeAP1-mediated oxidative stress response to regulate growth and virulence of P. expansum. IMPORTANCE Reactive oxygen species are the core of host plant defense and also play a vital role in the successful invasion of host plants by pathogenic fungi. Despite its importance, the relevance of oxidative stress response in fungal growth and virulence is poorly understood in P. expansum. In this study, we reveal that the transcription factor PeAP1 acts as a central regulator of oxidative stress response in P. expansum and that there is a major link between PeAP1-mediated oxidative stress response and fungal growth and virulence. To explore the underlying mechanisms, we performed comparative transcriptomic studies and identified a number of H2O2-induced PeAP1 target genes, including four novel ones, PePrx1, PePrx2, PeGST1, and PeTRX2, whose functions were linked to PeAP1 and pathogenicity. These findings provide novel insights into the regulation mechanism of PeAP1 on growth and virulence, which might offer promising targets for control of blue mold and patulin contamination.

Progression of the PI3K/Akt signaling pathway in chronic obstructive pulmonary disease.

Chronic Obstructive Pulmonary Disease (COPD) is a chronic respiratory disease characterized by a slow progression and caused by the inhalation of harmful particulate matter. Cigarette smoke and air pollutants are the primary contributing factors. Currently, the pathogenesis of COPD remains incompletely understood. The PI3K/Akt signaling pathway has recently emerged as a critical regulator of inflammation and oxidative stress response in COPD, playing a pivotal role in the disease's progression and treatment. This paper reviews the association between the PI3K/Akt pathway and COPD, examines effective PI3K/Akt inhibitors and novel anti-COPD agents, aiming to identify new therapeutic targets for clinical intervention in this disease.

Chronic memantine disrupts spatial memory and up-regulates Htr1a gene expression in the hippocampus of GPR39 (zinc-sensing receptor) KO male mice

GPR39 is a receptor involved in zincergic neurotransmission, and its role in regulating psychological functions is an active area of research. The purported roles of GPR39 at the cellular level include regulation of inflammatory and oxidative stress response, and modulation of GABAergic and endocannabinoid neurotransmission. GPR39 knock-out (KO) mice exhibit episodic-like and spatial memory (ELM and SM, respectively) deficits throughout their lifetime, and are similar in that respect to senescent wild-type (WT) conspecifics. Since a role for zinc has been postulated in neurodegenerative disorders, in this study we investigated the possibility of a pharmacological rescue of both types of declarative memory with memantine - a noncompetitive NMDAR antagonist used for slowing down dementia; or, a putative GPR39 agonist – TC-G 1008.First, we tested adult WT and GPR39KO male mice under acute 5 mg/kg memantine or vehicle treatment in an object recognition task designed to simultaneously probe the “what?”, “where?” and “when?” components of ELM. Next, we investigated the impact of chronic memantine or TC-G 1008 on ELM and SM (Morris water maze, MWM) in both WT and GPR39KO mice. Following chronic experiments, we assessed with qRT-PCR hippocampal gene expression of targets previously associated with GPR39.We report: no effects of acute memantine on ELM; a tendency to improve the “where?” component of ELM in both WT and GPR39 KO mice following 12 days of memantine; and, a disruption of SM in GPR39KO mice after 24 days of memantine treatment. The latter result was associated with upregulation of Htr1a hippocampal expression.

LLDPE/TiO&lt;sub&gt;2&lt;/sub&gt;-ZnO Nanocomposite Films induces Transitory Oxidative Stress Response on Human Fibroblast and Blood Cell Lines Models

Healthcare-associated infections (HAIs) are a major safety concern globally that contribute to mortality rates amongst patients especially associated with indwelling or implanted medical devices. The advanced metal-oxide nanocomposites (MNPs) embedded in polymer matrix present an outstanding antibacterial profile, especially for MDR strains owing to reactive oxygen species (ROS) and free radicals’ mode of action. To date, there is still a lack of knowledge on the implication of external reactive species from MNPs-based polymers to humans. This study investigates the bio-interaction of TiO2-ZnO nanocomposite films embedded in linear low-density polyethylene (LLDPE/ TiO2-ZnO) on human fibroblast and blood cell lines model at molecular genes and protein level. The initial analysis of the in vitro bio-interaction responses on fibroblast and blood cell line models showed signs of cell membrane integrity disturbance, which might be due to free radicals’ activities, such as the release of intracellular ROS and Zn ions (Zn2+) during the initial cellular adaptation process on the TiO2–ZnO polymer nanocomposite film. Further findings found that cell–polymer nanocomposite film interaction could possibly trigger transitory oxidative stress response and cellular redox regulation via NF-kβ interactions. However, further comprehensive studies are needed to support this study, especially involving animal models.

RGS4 impacts carbohydrate and siderophore metabolism in Trichoderma reesei

BackgroundAdaptation to complex, rapidly changing environments is crucial for evolutionary success of fungi. The heterotrimeric G-protein pathway belongs to the most important signaling cascades applied for this task. In Trichoderma reesei, enzyme production, growth and secondary metabolism are among the physiological traits influenced by the G-protein pathway in a light dependent manner.ResultsHere, we investigated the function of the SNX/H-type regulator of G-protein signaling (RGS) protein RGS4 of T. reesei. We show that RGS4 is involved in regulation of cellulase production, growth, asexual development and oxidative stress response in darkness as well as in osmotic stress response in the presence of sodium chloride, particularly in light. Transcriptome analysis revealed regulation of several ribosomal genes, six genes mutated in RutC30 as well as several genes encoding transcription factors and transporters. Importantly, RGS4 positively regulates the siderophore cluster responsible for fusarinine C biosynthesis in light. The respective deletion mutant shows altered growth on nutrient sources related to siderophore production such as ornithine or proline in a BIOLOG phenotype microarray assay. Additionally, growth on storage carbohydrates as well as several intermediates of the D-galactose and D-arabinose catabolic pathway is decreased, predominantly in light.ConclusionsWe conclude that RGS4 mainly operates in light and targets plant cell wall degradation, siderophore production and storage compound metabolism in T. reesei.

ATO Increases ROS Production and Apoptosis of Cells by Enhancing Calpain-Mediated Degradation of the Cancer Survival Protein TG2.

Transglutaminase 2 (TG2) is a critical cancer cell survival factor that activates several signalling pathways to foster drug resistance, cancer stem cell survival, metastasis, inflammation, epithelial-mesenchymal transition, and angiogenesis. All-trans retinoic acid (ATRA) and chemotherapy have been the standard treatments for acute promyelocytic leukaemia (APL), but clinical studies have shown that arsenic trioxide (ATO), alone or in combination with ATRA, can improve outcomes. ATO exerts cytotoxic effects in a variety of ways by inducing oxidative stress, genotoxicity, altered signal transduction, and/or epigenetic modification. In the present study, we showed that ATO increased ROS production and apoptosis ratios in ATRA-differentiated NB4 leukaemia cells, and that these responses were enhanced when TG2 was deleted. The combined ATRA + ATO treatment also increased the amount of nuclear factor erythroid 2-related factor 2 (NRF2) transcription factor, an adaptive regulator of the cellular oxidative stress response, and calpain proteolytic activity, resulting in TG2 degradation and the reduced survival of WT leukaemia cells. We further showed that the induced TG2 protein expression was degraded in the MCF-7 epithelial cell line and primary peripheral blood mononuclear cells upon ATO treatment, thereby sensitising these cell types to apoptotic signals.

Protective effect of cafestol against doxorubicin-induced cardiotoxicity in rats by activating the Nrf2 pathway.

Doxorubicin (DOX) is an efficient antineoplastic agent with a broad antitumor spectrum; however, doxorubicin-associated cardiotoxic adverse effect through oxidative damage and apoptosis limits its clinical application. Cafestol (Caf) is a naturally occurring diterpene in unfiltered coffee with unique antioxidant, antimutagenic, and anti-inflammatory activities by activating the Nrf2 pathway. The present study aimed to investigate the potential chemoprotective effect of cafestol on DOX-induced cardiotoxicity in rats. Wistar albino rats of both sexes were administered cafestol (5mg/kg/day) for 14 consecutive days by oral gavage alone or with doxorubicin which was injected as a single dose (15mg/kg intraperitoneally at day 14) to induce toxicity. The result showed that Caf significantly improved cardiac injury induced by doxorubicin, decreased serum levels of CK-MB, LDH, ALP, and ALT, and improved histopathological changes. In addition, cafestol significantly inhibited DOX-induced cardiac oxidative stress as seen in the reduced level of MDA and increased GSH, SOD, CAT, and Gpx-1 cardiac tissue levels; cafestol significantly enhanced Nrf2 gene and protein expression and promoted the expression of downstream antioxidant genes HO-1 and NQO-1 and downregulated Keap1 and NF-κB genes' expression; in addition, Caf significantly reduced inflammatory mediators, TNF-α, and IL-1β levels and inhibited cardiac apoptosis by modulating Bax and Casp 3 tissue levels and reduced TUNEL-positive cardiomyocytes. In conclusion, the present study confirmed that cafestol improved the cardiotoxic effects induced by doxorubicin through the regulation of apoptosis and oxidative stress response through the Nrf2 pathway; this study suggests that cafestol may serve as a potential adjuvant in chemotherapy to alleviate DOX-induced toxicities.

Mechanism of Liuwei Dihuang Pills in treatment of mice with diminished ovarian reserve based on proteomics

This study aims to investigate the efficacy and possible mechanism of Liuwei Dihuang Pills in the treatment of diminished ovarian reserve(DOR) by using proteomic techniques. Firstly, cyclophosphamide(60 mg·kg~(-1)) combined with busulfan(6 mg·kg~(-1)) was injected intraperitoneally to establish the mouse model of DOR. After drug injection, the mice were continuously observed and the success of modeling was evaluated by the disturbance of the estrous cycle. After successful modeling, the mice were administrated with the suspension of Liuwei Dihuang Pills by gavage for 28 days. At the end of the gavage, four female mice were selected and caged together with males at a ratio of 2∶1 for the determination of the pregnancy rate. Blood and ovary samples were collected from the remaining mice on the next day after the end of gavage. Hematoxylin-eosin(HE) staining and transmission electron microscopy(TEM) were then employed to observe the morphological and ultrastructural changes in the ovaries. The serum levels of hormones and oxidation indicators were measured by enzyme-linked immunosorbent assay. Quantitative proteomics techniques were used to compare the ovarian protein expression before and after modeling and before and after the intervention with Liuwei Dihuang Pills. The results showed that Liuwei Dihuang Pills regulated the estrous cycle of DOR mice, elevated the serum levels of hormones and anti-oxidation indicators, promoted follicle development, protected the mitochondrial morphology of ovarian granulosa cells, and increased the litter size and survival of DOR mice. Furthermore, Liuwei Dihuang Pills negatively regulated the expression of 12 differentially expressed proteins associated with DOR, which were mainly involved in lipid catabolism, inflammatory response, immune regulation, and coenzyme biosynthesis. These differentially expressed proteins were significantly enriched in sphingolipid metabolism, arachidonic acid metabolism, ribosomes, ferroptosis, and cGMP-PKG signaling pathway. In summary, the occurrence of DOR and the treatment of DOR with Liuwei Dihuang Pills are associated with multiple biological pathways, mainly including oxidative stress response, inflammatory response, and immune regulation. &quot;Mitochondria-oxidative stress-apoptosis&quot; is the key to the treatment of DOR by Liuwei Dihuang Pills. YY1 and CYP4F3 may be the key upstream targets that trigger mitochondrial dysfunction and ROS accumulation, and the metabolism of arachidonic acid is the main signaling pathway of drug action.

The age-related changes in lymph node stromal cells underlie defects in peripheral T cell maintenance and immune function decline in old mice.

Abstract Older adults are more vulnerable to infection and cancer, and poorly respond to vaccination. The structural stromal cells in the lymph nodes (LN) are essential regulators of T cell homeostasis and immune response. However, how does aging of LN stromal cells contribute to T cell maintenance defects and functional immunity is incompletely understood. Here we used single cell RNA-Seq of purified stromal cells from adult (3mo) and old (21mo) peripheral LN of C57BL/6 mice to characterize the age-related transcriptional differences in major stromal cell subsets. Our data revealed the enrichment of CCL19 hiand CCL19 lowwhile diminution of Nr4a1 +and Inmt +FRC in old LN. The enriched FRC in old LN exhibited a gene signature of cellular and oxidative stress response, mitochondrial defects, and negative regulation of immunity. The ex vivo functional characterization revealed that old LN stromal cells exhibit elevated mitochondrial reactive oxygen species coupled with increased mitochondrial mass and reduced membrane potential, indicative of mitochondrial defects during the senescence. Interestingly, these defects are observed in older but not mid-age LN. Furthermore, in vitro co-culture of LN stromal cells and purified naive T (T N) cells showed that old LN stromal cells poorly support CD8 T Ncell survival. Finally, boosting the function of old LN stromal cells using agonistic anti-LTbR or mitigating oxidative stress using N-Acetyl Cysteine reversed this effect, via stabilization of BCL2 in T Ncells. Thus, these results suggest that age-related adverse changes in LN stromal cells contribute to defects in maintenance of T Ncells, and highlight that these defects could be reversed to restore T Ncell homeostasis and functional immunity. This work is supported by grants from NIH (P01 AG052359 and R37 AG020719) to Dr. Janko Nikolich-Zugich.

BosR and PlzA reciprocally regulate RpoS function to sustain Borrelia burgdorferi in ticks and mammals.

The RNA polymerase alternative σ factor RpoS in Borrelia burgdorferi (Bb), the Lyme disease pathogen, is responsible for programmatic-positive and -negative gene regulation essential for the spirochete's dual-host enzootic cycle. RpoS is expressed during tick-to-mammal transmission and throughout mammalian infection. Although the mammalian-phase RpoS regulon is well described, its counterpart during the transmission blood meal is unknown. Here, we used Bb-specific transcript enrichment by tick-borne disease capture sequencing (TBDCapSeq) to compare the transcriptomes of WT and ΔrpoS Bb in engorged nymphs and following mammalian host-adaptation within dialysis membrane chambers. TBDCapSeq revealed dramatic changes in the contours of the RpoS regulon within ticks and mammals and further confirmed that RpoS-mediated repression is specific to the mammalian-phase of Bb's enzootic cycle. We also provide evidence that RpoS-dependent gene regulation, including repression of tick-phase genes, is required for persistence in mice. Comparative transcriptomics of engineered Bb strains revealed that the Borrelia oxidative stress response regulator (BosR), a noncanonical Fur family member, and the cyclic diguanosine monophosphate (c-di-GMP) effector PlzA reciprocally regulate the function of RNA polymerase complexed with RpoS. BosR is required for RpoS-mediated transcription activation and repression in addition to its well-defined role promoting transcription of rpoS by the RNA polymerase alternative σ factor RpoN. During transmission, ligand-bound PlzA antagonizes RpoS-mediated repression, presumably acting through BosR.

The APC/C Activator Cdh1p Plays a Role in Mitochondrial Metabolic Remodelling in Yeast

Cdh1p is one of the two substrate adaptor proteins of the anaphase promoting complex/cyclosome (APC/C), a ubiquitin ligase that regulates proteolysis during cell cycle. In this work, using a proteomic approach, we found 135 mitochondrial proteins whose abundance was significantly altered in the cdh1Δ mutant, with 43 up-regulated proteins and 92 down-regulated proteins. The group of significantly up-regulated proteins included subunits of the mitochondrial respiratory chain, enzymes from the tricarboxylic acid cycle and regulators of mitochondrial organization, suggesting a metabolic remodelling towards an increase in mitochondrial respiration. In accordance, mitochondrial oxygen consumption and Cytochrome c oxidase activity increased in Cdh1p-deficient cells. These effects seem to be mediated by the transcriptional activator Yap1p, a major regulator of the yeast oxidative stress response. YAP1 deletion suppressed the increased Cyc1p levels and mitochondrial respiration in cdh1Δ cells. In agreement, Yap1p is transcriptionally more active in cdh1Δ cells and responsible for the higher oxidative stress tolerance of cdh1Δ mutant cells. Overall, our results unveil a new role for APC/C-Cdh1p in the regulation of the mitochondrial metabolic remodelling through Yap1p activity.