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Fibrosis is a pathological reparative process that can occur in most organs and is responsible for nearly half of deaths in the developed world. Despite considerable research, few therapies have proven effective and been approved clinically for treatment of fibrosis. Artemisinin compounds are best known as antimalarial therapeutics, but they also demonstrate antiparasitic, antibacterial, anticancer, and anti-fibrotic effects. Here we summarize literature describing anti-fibrotic effects of artemisinin compounds in in vivo and in vitro models of tissue fibrosis, and we describe the likely mechanisms by which artemisinin compounds appear to inhibit cellular and tissue processes that lead to fibrosis. To consider alternative routes of administration of artemisinin for treatment of internal organ fibrosis, we also discuss the potential for more direct oral delivery of Artemisia plant material to enhance bioavailability and efficacy of artemisinin compared to administration of purified artemisinin drugs at comparable doses. It is our hope that greater understanding of the broad anti-fibrotic effects of artemisinin drugs will enable and promote their use as therapeutics for treatment of fibrotic diseases., Graphical abstract Artemisinin drugs, isolated from Artemisia, effectively prevent or treat multiple types of tissue fibrosis when administered to several preclinical animal models of varied etiologies.Image 1

Visual Abstract, Highlights • Leptin protects against progression to heart failure after myocardial infarction. • This beneficial effect requires activation of the brain melanocortin system. • Stimulation of brain MC4R recapitulates the cardiac protective effects of leptin. • Leptin-MC4R activation improves cardiac substrate oxidation and mitochondrial function. • It also improves Ca2+ coupling and contractile function in viable cardiomyocytes after MI., Summary Heart failure has a high mortality rate, and current therapies offer limited benefits. The authors demonstrate that activation of the central nervous system leptin-melanocortin pathway confers remarkable protection against progressive heart failure following severe myocardial infarction. The beneficial cardiac-protective actions of leptin require activation of brain melanocortin-4 receptors and elicit improvements in cardiac substrate oxidation, cardiomyocyte contractility, Ca2+ coupling, and mitochondrial efficiency. These findings highlight a potentially novel therapeutic approach for myocardial infarction and heart failure.

Tumor cells often switch from mitochondrial oxidative metabolism to glycolytic metabolism even under aerobic conditions. Tumor cell glycolysis is accompanied by several nonenzymatic activities among which induction of drug resistance has important therapeutic implications. In this article, we review the main aspects of glycolysis-induced drug resistance. We discuss the classes of antitumor drugs that are affected and the components of the glycolytic pathway (transporters, enzymes, metabolites) that are involved in the induction of drug resistance. Glycolysis-associated drug resistance occurs in response to stimuli, either cell-autonomous (e.g., oncoproteins) or deriving from the tumor microenvironment (e.g., hypoxia or pseudohypoxia, mechanical cues, etc.). Several mechanisms mediate the induction of drug resistance in response to glycolytic metabolism: inhibition of apoptosis, induction of epithelial-mesenchymal transition, induction of autophagy, inhibition of drug influx and increase of drug efflux. We suggest that drug resistance in response to glycolysis comes into play in presence of qualitative (e.g., expression of embryonic enzyme isoforms, post-translational enzyme modifications) or quantitative (e.g., overexpression of enzymes or overproduction of metabolites) alterations of glycolytic metabolism. We also discern similarities between changes occurring in tumor cells in response to stimuli inducing glycolysis-associated drug resistance and those occurring in cells of the innate immune system in response to danger signals and that have been referred to as danger-associated metabolic modifications. Eventually, we briefly address that also mitochondrial oxidative metabolism may induce drug resistance and discuss the therapeutic implications deriving from the fact that the main energy-generating metabolic pathways may be both at the origin of antitumor drug resistance.

Metabolic dysfunction is a major driver of tumorigenesis. The serine/threonine kinase mechanistic target of rapamycin (mTOR) constitutes a key central regulator of metabolic pathways promoting cancer cell proliferation and survival. mTOR activity is regulated by metabolic sensors as well as by numerous factors comprising the phosphatase and tensin homolog/PI3K/AKT canonical pathway, which are often mutated in cancer. However, some cancers displaying constitutively active mTOR do not carry alterations within this canonical pathway, suggesting alternative modes of mTOR regulation. Since DEPTOR, an endogenous inhibitor of mTOR, was previously found to modulate both mTOR complexes 1 and 2, we investigated the different post-translational modification that could affect its inhibitory function. We found that tyrosine (Tyr) 289 phosphorylation of DEPTOR impairs its interaction with mTOR, leading to increased mTOR activation. Using proximity biotinylation assays, we identified SYK (spleen tyrosine kinase) as a kinase involved in DEPTOR Tyr 289 phosphorylation in an ephrin (erythropoietin-producing hepatocellular carcinoma) receptor–dependent manner. Altogether, our work reveals that phosphorylation of Tyr 289 of DEPTOR represents a novel molecular switch involved in the regulation of both mTOR complex 1 and mTOR complex 2.

Author(s): Dewan, Anna K; Gupta, Sameer; Bach, Daniel Q; Jadeja, Saagar; Granter, Scott; LaCasce, Ann; Joyce, Robin; Davids, Matthew S; Huang, Victor; LeBoeuf, Nicole R

The mechanistic target of rapamycin (mTOR) pathway integrates metabolic cues into cell fate decisions. A particularly fateful event during the adaptive immune response is the engagement of a T cell receptor by its cognate antigen presented by an antigen-presenting cell (APC). Here, the induction of adequate T cell activation and lineage specification is critical to mount protective immunity; at the same time, inadequate activation, which could lead to autoimmunity, must be avoided. mTOR forms highly conserved protein complexes 1 and 2 that shape lineage specification by integrating signals originating from TCR engagement, co-stimulatory or co-inhibitory receptors and cytokines and availability of nutrients. If one considers autoimmunity as the result of aberrant lineage specification in response to such signals, the importance of this pathway becomes evident; this provides the conceptual basis for mTOR inhibition in the treatment of systemic autoimmunity, such as systemic lupus erythematosus (SLE). Clinical trials in SLE patients have provided preliminary evidence that mTOR blockade by sirolimus (rapamycin) can reverse pro-inflammatory lineage skewing, including the expansion of Th17 and double-negative T cells and plasma cells and the contraction of regulatory T cells. Moreover, sirolimus has shown promising efficacy in the treatment of refractory idiopathic multicentric Castleman disease, newly characterized by systemic autoimmunity due to mTOR overactivation. Alternatively, mTOR blockade enhances responsiveness to vaccination and reduces infections by influenza virus in healthy elderly subjects. Such seemingly contradictory findings highlight the importance to further evaluate the clinical effects of mTOR manipulation, including its potential role in treatment of COVID-19 infection. mTOR blockade may extend healthy lifespan by abrogating inflammation induced by viral infections and autoimmunity. This review provides a mechanistic assessment of mTOR pathway activation in lineage specification within the adaptive and innate immune systems and its role in health and autoimmunity. We then discuss some of the recent experimental and clinical discoveries implicating mTOR in viral pathogensis and aging.

Cellular growth and proliferation are primarily dictated by the mechanistic target of rapamycin complex 1 (mTORC1), which balances nutrient availability against the cell's anabolic needs. Central to the activity of mTORC1 is the RagA-RagC GTPase heterodimer, which under favorable conditions recruits the complex to the lysosomal surface to promote its activity. The RagA-RagC heterodimer has a unique architecture in that both subunits are active GTPases. To promote mTORC1 activity, the RagA subunit is loaded with GTP and the RagC subunit is loaded with GDP, while the opposite nucleotide-loading configuration inhibits this signaling pathway. Despite its unique molecular architecture, how the Rag GTPase heterodimer maintains the oppositely loaded nucleotide state remains elusive. Here, we applied structure-function analysis approach to the crystal structures of the Rag GTPase heterodimer and identified a key hydrogen bond that stabilizes the GDP-loaded state of the Rag GTPases. This hydrogen bond is mediated by the backbone carbonyl of Asn30 in the nucleotide-binding domain of RagA or Lys84 of RagC and the hydroxyl group on the side chain of Thr210 in the C-terminal roadblock domain of RagA or Ser266 of RagC, respectively. Eliminating this interdomain hydrogen bond abolishes the ability of the Rag GTPase to maintain its functional state, resulting in a distorted response to amino acid signals. Our results reveal that this long-distance interdomain interaction within the Rag GTPase is required for the maintenance and regulation of the mTORC1 nutrient-sensing pathway.

The mechanistic target of rapamycin (mTOR) is often referred to as a master regulator of the cellular metabolism that can integrate the growth factor and nutrient signaling. Fasting suppresses hepatic mTORC1 activity via the activity of the tuberous sclerosis complex (TSC), a negative regulator of mTORC1, to suppress anabolic metabolism. The loss of TSC1 in the liver locks the liver in a constitutively anabolic state even during fasting, which was suggested to regulate peroxisome proliferator-activated receptor alpha (PPARα) signaling and ketogenesis, but the molecular determinants of this regulation are unknown. Here, we examined if the activation of the mTORC1 complex in mice by the liver-specific deletion of TSC1 (TSC1L−/−) is sufficient to suppress PPARα signaling and therefore ketogenesis in the fasted state. We found that the activation of mTORC1 in the fasted state is not sufficient to repress PPARα-responsive genes or ketogenesis. Furthermore, we examined whether the activation of the anabolic program mediated by mTORC1 complex activation in the fasted state could suppress the robust catabolic programming and enhanced PPARα transcriptional response of mice with a liver-specific defect in mitochondrial long-chain fatty acid oxidation using carnitine palmitoyltransferase 2 (Cpt2L−/−) mice. We generated Cpt2L−/−; Tsc1L−/− double-KO mice and showed that the activation of mTORC1 by deletion of TSC1 could not suppress the catabolic PPARα-mediated phenotype of Cpt2L−/− mice. These data demonstrate that the activation of mTORC1 by the deletion of TSC1 is not sufficient to suppress a PPARα transcriptional program or ketogenesis after fasting.

Highlight • NET possesses pro-tumorigenesis effect on human breast epithelial MCF10A cells. • NET upregulates PGRMC1/EGFR and suppresses PTEN expression. • NET stimulates the PI3K/Akt/mTOR signal pathway. • Overexpression of miR-181a abrogates the effects of NET., Background Research suggests that hormone replacement therapy may increase the risk of breast cancer, and progestins such as norethisterone (NET) play a key role in this phenomenon. We have demonstrated that microRNA-181a (miR-181a) suppresses NET-promoted breast cancer cell survival. Nonetheless, the effects of NET and miR-181a on the tumorigenesis of human breast epithelial cells have not yet been elaborated. Methods Assays of cell viability, proliferation, migration, apoptosis, and colony formation were performed to investigate the pro-tumorigenesis effect of NET and the effects of miR-181a on human breast epithelial MCF10A cells. The expressions of cell-proliferation-related genes and apoptotic factors were analyzed by quantitative RT-PCR and Western blot in MCF10A cells treated with NET and miR-181a. Results NET significantly increased MCF10A cell viability, proliferation, migration, and colony formation, but reduced cellular apoptosis. In addition, NET increased the expression of progesterone receptor membrane component 1 (PGRMC1), EGFR, B-cell lymphoma 2, cyclin D1, and proliferating cell nuclear antigen, but decreased the expression of pro-apoptosis factors, such as Bax, caspase-7, and caspase-9. Overexpression of miR-181a strongly inhibited the effects of NET on MCF10A cells and abrogated NET-stimulated PGRMC1, EGFR, and mTOR expression. Conclusions Activation of the PGRMC1/EGFR–PI3K/Akt/mTOR signaling pathway is the primary mechanism underlying the pro-tumorigenesis effects of NET on human breast epithelial MCF10A cells. Additionally, miR-181a can suppress the effects of NET on these cells. These data suggest a therapeutic potential for miR-181a in reducing or preventing the risk of breast cancer in hormone replacement therapy using NET.

Long-chain fatty acid oxidation is frequently impaired in primary and systemic metabolic diseases affecting the heart; thus, therapeutically increasing reliance on normally minor energetic substrates, such as ketones and medium-chain fatty acids, could benefit cardiac health. However, the molecular fundamentals of this therapy are not fully known. Here, we explored the ability of octanoate, an eight-carbon medium-chain fatty acid known as an unregulated mitochondrial energetic substrate, to ameliorate cardiac hypertrophy in long-chain fatty acid oxidation-deficient hearts because of carnitine palmitoyltransferase 2 deletion (Cpt2M−/−). CPT2 converts acylcarnitines to acyl-CoAs in the mitochondrial matrix for oxidative bioenergetic metabolism. In Cpt2M−/− mice, high octanoate-ketogenic diet failed to alleviate myocardial hypertrophy, dysfunction, and acylcarnitine accumulation suggesting that this alternative substrate is not sufficiently compensatory for energy provision. Aligning this outcome, we identified a major metabolic distinction between muscles and liver, wherein heart and skeletal muscle mitochondria were unable to oxidize free octanoate, but liver was able to oxidize free octanoate. Liver mitochondria, but not heart or muscle, highly expressed medium-chain acyl-CoA synthetases, potentially enabling octanoate activation for oxidation and circumventing acylcarnitine shuttling. Conversely, octanoylcarnitine was oxidized by liver, skeletal muscle, and heart, with rates in heart 4-fold greater than liver and, in muscles, was not dependent upon CPT2. Together, these data suggest that dietary octanoate cannot rescue CPT2-deficient cardiac disease. These data also suggest the existence of tissue-specific mechanisms for octanoate oxidative metabolism, with liver being independent of free carnitine availability, whereas cardiac and skeletal muscles depend on carnitine but not on CPT2.

Graphical abstract, Necrostatin-1 (Nec-1) is a RIP1-targeted inhibitor of necroptosis, a form of programmed cell death discovered and investigated in recent years. There are already many studies demonstrating the essential role of necroptosis in various diseases, including inflammatory diseases, cardiovascular diseases and neurological diseases. However, the potential of Nec-1 in diseases has not received much attention. Nec-1 is able to inhibit necroptosis signaling pathway and thus ameliorate necroptotic cell death in disease development. Recent research findings indicate that Nec-1 could be applied in several types of diseases to alleviate disease development or improve prognosis. Moreover, we predict that Nec-1 has the potential to protect against the complications of coronavirus disease 2019 (COVID-19). This review summarized the effect of Nec-1 in disease models and the underlying molecular mechanism, providing research evidence for its future application.

Numerous observations indicate that red blood cells (RBCs) affect T-cell activation and proliferation. We have studied effects of packed RBCs (PRBCs) on T-cell receptor (TCR) signaling and the molecular mechanisms whereby (P)RBCs modulate T-cell activation. In line with previous reports, PRBCs attenuated the expression of T-cell activation markers CD25 and CD69 upon costimulation via CD3/CD28. In addition, T-cell proliferation and cytokine expression were markedly reduced when T-cells were stimulated in the presence of PRBCs. Inhibitory activity of PRBCs required direct cell–cell contact and intact PRBCs. The production of activation-induced cellular reactive oxygen species, which act as second messengers in T-cells, was completely abrogated to levels of unstimulated T-cells in the presence of PRBCs. Phosphorylation of the TCR-related zeta chain and thus proximal TCR signal transduction was unaffected by PRBCs, ruling out mechanisms based on secreted factors and steric interaction restrictions. In large part, downstream signaling events requiring reactive oxygen species for full functionality were affected, as confirmed by an untargeted MS-based phosphoproteomics approach. PRBCs inhibited T-cell activation more efficiently than treatment with 1 mM of the antioxidant N-acetyl cysteine. Taken together, our data imply that inflammation-related radical reactions are modulated by PRBCs. These immunomodulating effects may be responsible for clinical observations associated with transfusion of PRBCs.

Mantle cell lymphoma (MCL) is an especially aggressive and highly heterogeneous mature B-cell lymphoma. Heat shock protein 90 (HSP90) is considered an attractive therapeutic target in a variety of cancers, including MCL, but no HSP90 inhibitors have succeeded in the clinical trials to date. Exploring fine mechanisms of HSP90 inhibition in cancer cells may shed light on novel therapeutic strategies. Here, we found that HSP90 knockdown and continuous inhibition with ganetespib inhibited growth of MCL cells in vitro and in vivo. To our surprise, transient exposure over 12 h was almost as efficient as continuous exposure, and treatment with ganetespib for 12 h efficiently inhibited growth and induced G1 cell cycle arrest and apoptosis of MCL cells. Transcriptome analysis complemented by functional studies was performed to define critical MCL signaling pathways that are exceptionally sensitive to HSP90 inhibition and vital to cell fate. Six genes (cell division cycle 6, cell division cycle 45, minichromosome maintenance 4, minichromosome maintenance 7, RecQ-mediated genome instability 2, and DNA primase polypeptide 1) involved in DNA replication and repair were identified as consistently downregulated in three MCL cell lines after transient ganetespib treatment. E2F1, an important transcription factor essential for cell cycle progression, was identified as a ganetespib target mediating transcriptional downregulation of these six genes, and its stability was also demonstrated to be maintained by HSP90. This study identifies E2F1 as a novel client protein of HSP90 that is very sensitive and worthy of targeting and also finds that HSP90 inhibitors may be useful in combination therapies for MCL.

Objective Nonalcoholic hepatic steatosis, also known as fatty liver, is a uniform response of the liver to hyperlipidic-hypercaloric diet intake. However, the post-ingestive signals and mechanistic processes driving hepatic steatosis are not well understood. Emerging data demonstrate that protein kinase C beta (PKCβ), a lipid-sensitive kinase, plays a critical role in energy metabolism and adaptation to environmental and nutritional stimuli. Despite its powerful effect on glucose and lipid metabolism, knowledge of the physiological roles of hepatic PKCβ in energy homeostasis is limited. Methods The floxed-PKCβ and hepatocyte-specific PKCβ-deficient mouse models were generated to study the in vivo role of hepatocyte PKCβ on diet-induced hepatic steatosis, lipid metabolism, and mitochondrial function. Results We report that hepatocyte-specific PKCβ deficiency protects mice from development of hepatic steatosis induced by high-fat diet, without affecting body weight gain. This protection is associated with attenuation of SREBP-1c transactivation and improved hepatic mitochondrial respiratory chain. Lipidomic analysis identified significant increases in the critical mitochondrial inner membrane lipid, cardiolipin, in PKCβ-deficient livers compared to control. Moreover, hepatocyte PKCβ deficiency had no significant effect on either hepatic or whole-body insulin sensitivity supporting dissociation between hepatic steatosis and insulin resistance. Conclusions The above data indicate that hepatocyte PKCβ is a key focus of dietary lipid perception and is essential for efficient storage of dietary lipids in liver largely through coordinating energy utilization and lipogenesis during post-prandial period. These results highlight the importance of hepatic PKCβ as a drug target for obesity-associated nonalcoholic hepatic steatosis., Highlights • Hepatocyte-specific PKCβ deficiency protects from diet-induced hepatic steatosis but not either obesity or insulin resistance. • The underlying mechanism involves modulation of mitochondrial function and lipogenesis. • Hepatocyte PKCβ does not regulate hepatic insulin signaling. • Our study provides a link between dietary lipid intake, hepatic protein kinase Cβ, modulation of hepatic lipid metabolism, and development of hepatic steatosis.

Graphical abstract, Introduction Due to its high specificity and sensitivity, liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) is the gold standard method for immunosuppressant quantification in therapeutic drug monitoring. In this context, dried blood spots (DBS) have become a promising strategy as a sample collection procedure. Although the advantages of DBS over venipuncture are well known, this approach has limitations that strongly influence the acceptance of analytical results. Among them, the most important is hematocrit (Ht). The easiest way of overcoming this problem is by analyzing complete spots. In this strategy, called dried matrix on paper discs (DMPD), blood is volumetrically applied on pre-punched discs. Objectives To validate an LC-MS/MS method for the quantification of tacrolimus, sirolimus, everolimus and cyclosporin A using DMPD. Methods The procedure was validated according to international guidelines using a commercial kit. The following performance parameters were evaluated: selectivity, carryover, linearity, accuracy, precision, lower limit of quantitation, relative recovery, commutability and stability. In addition, a method comparison study was performed to evaluate the clinical influence of Ht on the results. Results All performance parameters were within acceptance criteria and, hence, it was determined that the validated method is fit for the intended purpose. Likewise, calculated bias values on medical decision levels showed that there was no clinical influence of Ht on the results. Conclusion Unlike other similar methodologies that have been published, here, a simple method has been fully validated. This is the first LC-MS/MS methodology adapting a commercial kit to use DMPD as a sampling strategy.

Background & Aims Metabolic imbalance and inflammation are common features of chronic liver diseases. Molecular factors controlling these mechanisms represent potential therapeutic targets. CD73 is the major enzyme that dephosphorylates extracellular adenosine monophosphate (AMP) to form the anti-inflammatory adenosine. CD73 is expressed on pericentral hepatocytes, which are important for long-term liver homeostasis. We aimed to determine if CD73 has nonredundant hepatoprotective functions. Methods Liver-specific CD73 knockout (CD73-LKO) mice were generated by targeting the Nt5e gene in hepatocytes. The CD73-LKO mice and hepatocytes were characterized using multiple approaches. Results Deletion of hepatocyte Nt5e resulted in an approximately 70% reduction in total liver CD73 protein (P < .0001). Male and female CD73-LKO mice developed normally during the first 21 weeks without significant liver phenotypes. Between 21 and 42 weeks, the CD73-LKO mice developed spontaneous-onset liver disease, with significant severity in male mice. Middle-aged male CD73-LKO mice showed hepatocyte swelling and ballooning (P < .05), inflammation (P < .01), and variable steatosis. Female CD73-LKO mice had lower serum albumin levels (P < .05) and increased inflammatory genes (P < .01), but did not show the spectrum of histopathologic changes in male mice, potentially owing to compensatory induction of adenosine receptors. Serum analysis and proteomic profiling of hepatocytes from male CD73-LKO mice showed significant metabolic imbalance, with increased blood urea nitrogen (P < .0001) and impairments in major metabolic pathways, including oxidative phosphorylation and AMP-activated protein kinase (AMPK) signaling. There was significant hypophosphorylation of AMPK substrates in CD73-LKO livers (P < .0001), while in isolated hepatocytes treated with AMP, soluble CD73 induced AMPK activation (P < .001). Conclusions Hepatocyte CD73 supports long-term metabolic liver homeostasis through AMPK in a sex-dependent manner. These findings have implications for human liver diseases marked by CD73 dysregulation., Graphical abstract

Background & Aims Cancer-associated fibroblasts (CAFs) from pancreatic adenocarcinoma (PDA) present high protein synthesis rates. CAFs express the G-protein–coupled somatostatin receptor sst1. The sst1 agonist SOM230 blocks CAF protumoral features in vitro and in immunocompromised mice. We have explored here the therapeutic potential of SOM230, and underlying mechanisms, in immunocompetent models of murine PDA mimicking the heavy fibrotic and immunosuppressive stroma observed in patient tumors. Methods Large-scale mass spectrometry analyses were performed on media conditioned from 9 patient PDA-derived CAF primary cultures. Spontaneous transgenic and experimental (orthotopic co-graft of tumor cells plus CAFs) PDA-bearing mice were longitudinally ultrasound-monitored for tumor and metastatic progression. Histopathology and flow cytometry analyses were performed on primary tumors and metastases. Stromal signatures were functionally validated through bioinformatics using several published, and 1 original, PDA database. Results Proteomics on the CAF secretome showed that SOM230 controls stromal activities including inflammatory responses. Among the identified secreted proteins, we validated that colony-stimulating factor 1 (CSF-1) (a macrophage growth factor) was reduced by SOM230 in the tumor and plasma of PDA-harboring mice, alongside intratumor stromal normalization (reduced CAF and macrophage activities), and dramatic metastasis reduction. In transgenic mice, these SOM230 benefits alleviate the chemotherapy-induced (gemcitabine) immunosuppressive stroma reshaping. Mechanistically, SOM230 acts in vivo on CAFs through sst1 to disrupt prometastatic CAF production of CSF-1 and cross-talk with macrophages. We found that in patients, stromal CSF-1 was associated with aggressive PDA forms. Conclusions We propose SOM230 as an antimetastatic therapy in PDA for its capacity to remodel the fibrotic and immunosuppressive myeloid stroma. This pharmacotherapy should benefit PDA patients treated with chemotherapies., Graphical abstract

Stroke is considered a leading cause of mortality and neurological disability, which puts a huge burden on individuals and the community. To date, effective therapy for stroke has been limited by its complex pathological mechanisms. Autophagy refers to an intracellular degrading process with the involvement of lysosomes. Autophagy plays a critical role in maintaining the homeostasis and survival of cells by eliminating damaged or non-essential cellular constituents. Increasing evidence support that autophagy protects neuronal cells from ischemic injury. However, under certain circumstances, autophagy activation induces cell death and aggravates ischemic brain injury. Diverse naturally derived compounds have been found to modulate autophagy and exert neuroprotection against stroke. In the present work, we have reviewed recent advances in naturally derived compounds that regulate autophagy and discussed their potential application in stroke treatment., Graphical abstract This review summarizes the current knowledge on natural compounds, focusing on the potential implication of stroke, acting as modulators of autophagy.Image 1

ABHD5 is an essential coactivator of ATGL, the rate-limiting triglyceride (TG) lipase in many cell types. Importantly, ABHD5 also functions as a tumor suppressor, and ABHD5 mRNA expression levels correlate with patient survival for several cancers. Nevertheless, the mechanisms involved in ABHD5-dependent tumor suppression are not known. We found that overexpression of ABHD5 induces cell cycle arrest at the G1 phase and causes growth retardation in a panel of prostate cancer cells. Transcriptomic profiling and biochemical analysis revealed that genetic or pharmacological activation of lipolysis by ABHD5 potently inhibits mTORC1 signaling, leading to a significant downregulation of protein synthesis. Mechanistically, we found that ABHD5 elevates intracellular AMP content, which activates AMPK, leading to inhibition of mTORC1. Interestingly, ABHD5-dependent suppression of mTORC1 was abrogated by pharmacological inhibition of DGAT1 or DGAT2, isoenzymes that re-esterify fatty acids in a process that consumes ATP. Collectively, this study maps out a novel molecular pathway crucial for limiting cancer cell proliferation, in which ABHD5-mediated lipolysis creates an energy-consuming futile cycle between TG hydrolysis and resynthesis, leading to inhibition of mTORC1 and cancer cell growth arrest.

Objective Laron syndrome (LS) is a rare, autosomal recessive disorder in humans caused by loss-of-function mutations of the growth hormone receptor (GHR) gene. To establish a large animal model for LS, pigs with GHR knockout (KO) mutations were generated and characterized. Methods CRISPR/Cas9 technology was applied to mutate exon 3 of the GHR gene in porcine zygotes. Two heterozygous founder sows with a 1-bp or 7-bp insertion in GHR exon 3 were obtained, and their heterozygous F1 offspring were intercrossed to produce GHR-KO, heterozygous GHR mutant, and wild-type pigs. Since the latter two groups were not significantly different in any parameter investigated, they were pooled as the GHR expressing control group. The characterization program included body and organ growth, body composition, endocrine and clinical-chemical parameters, as well as signaling studies in liver tissue. Results GHR-KO pigs lacked GHR and had markedly reduced serum insulin-like growth factor 1 (IGF1) levels and reduced IGF-binding protein 3 (IGFBP3) activity but increased IGFBP2 levels. Serum GH concentrations were significantly elevated compared with control pigs. GHR-KO pigs had a normal birth weight. Growth retardation became significant at the age of five weeks. At the age of six months, the body weight of GHR-KO pigs was reduced by 60% compared with controls. Most organ weights of GHR-KO pigs were reduced proportionally to body weight. However, the weights of liver, kidneys, and heart were disproportionately reduced, while the relative brain weight was almost doubled. GHR-KO pigs had a markedly increased percentage of total body fat relative to body weight and displayed transient juvenile hypoglycemia along with decreased serum triglyceride and cholesterol levels. Analysis of insulin receptor related signaling in the liver of adult fasted pigs revealed increased phosphorylation of IRS1 and PI3K. In agreement with the loss of GHR, phosphorylation of STAT5 was significantly reduced. In contrast, phosphorylation of JAK2 was significantly increased, possibly due to the increased serum leptin levels and increased hepatic leptin receptor expression and activation in GHR-KO pigs. In addition, increased mTOR phosphorylation was observed in GHR-KO liver samples, and phosphorylation studies of downstream substrates suggested the activation of mainly mTOR complex 2. Conclusion GHR-KO pigs resemble the pathophysiology of LS and are an interesting model for mechanistic studies and treatment trials., Highlights • GHR-deficient pigs reveal postnatal growth retardation, disproportionate organ growth and an increased total body fat content. • GHR-deficient pigs show markedly reduced serum IGF1 and IGFBP3 levels, and transient juvenile hypoglycemia. • Increased expression and phosphorylation of IRS1 in liver of adult GHR-deficient pigs suggest increased insulin sensitivity. • Increased phosphorylation of JAK2 in liver of GHR-deficient pigs may be explained by higher serum leptin levels and activation of hepatic LEPR.

Preliminary analysis from the Vax-On study did not find a correlation between cancer treatment type and antibody response to COVID-19 vaccination. We carried out a secondary subgroup analysis to verify the effects of comprehensive cancer treatment classification on vaccine immunogenicity.The Vax-On study prospectively enrolled patients who started a two-dose messenger RNA-BNT162b2 vaccine schedule from 9 March 2021 to 12 April 2021 (timepoint-1). Those on active treatment within the previous 28 days accounted for the exposed cases. Patients who had discontinued such treatment by at least 28 days or received intravesical therapy represented the control cases. Quantification of immunoglobulin G (IgG) antibodies against the receptor binding domain of the S1 subunit of the SARS-CoV-2 spike protein was carried out before the second dose (timepoint-2) and 8 weeks thereafter (timepoint-3). Seroconversion response was defined at ≥50 arbitrary units/ml IgG titer. Classification of antineoplastic agents was based on their pharmacodynamic properties.Three hundred and sixty-six patients were enrolled (86 and 260 as control and exposed cases, respectively). Univariate analysis revealed a significantly lower IgG titer after both doses of vaccine in subgroups treated with tyrosine kinase inhibitors (TKIs), multiple cytotoxic agents, alkylating agents, and topoisomerase inhibitors. At timepoint-3, seroconversion response was significantly impaired in the topoisomerase inhibitors and mechanistic target of rapamycin (mTOR) inhibitors subgroups. After multivariate testing, treatment with alkylating agents and TKIs was significantly associated with a reduced change in IgG titer at timepoint-2. Treatment with mTOR inhibitors resulted in a similar interaction at each timepoint. Cyclin-dependent kinase 4/6 inhibitor treatment was independently correlated with an incremental variation in IgG titer at timepoint-3. Specific subgroups (TKIs, antimetabolites, alkylating agents, and multiple-agent chemotherapy) predicted lack of seroconversion at timepoint-2, but their effect was not retained at timepoint-3. Eastern Cooperative Oncology Group performance status 2, immunosuppressive corticosteroid dosing, and granulocyte colony-stimulating factor use were independently linked to lower IgG titer after either dose of vaccine.Drugs interfering with DNA synthesis, multiple-agent cytotoxic chemotherapy, TKIs, mTOR and cyclin-dependent kinase 4/6 inhibitors differentially modulate humoral response to messenger RNA-BNT162b2 vaccine.