Introduction. Multiparameter flow cytometry (FC) has demonstrated superior sensitivity over conventional cytology (CC) in detecting occult central system disease (OCNSD) in adult acute lymphoblastic leukemia (ALL) patients (pts), a condition associated with a more unfavorable outcome. This has led to intensified CNS-directed therapy (CNSDT) in OCNSD pts, though its impact on relapse risk remains uncertain.
Aim. To prospectively assess disease-free survival (DFS) of OCNSD pts via a multicenter, observational Campus ALL study.
Methods. We recorded data on all consecutive cerebrospinal fluid (CSF) samples from adult ALL pts (since 1 June 2020) analyzed by CC and FC. Data on demographics, ALL type, biology, CSF features, intrathecal (IT) prophylaxis/therapy, FC event count, treatments, complete remission (CR), relapse (RE), and mortality were also collected. Positivity by CC was defined by standard criteria. FC used 8–10 antibodies, with at least 10 clonal and abnormal events considered positive. CSF results identified 3 groups: manifest CNS disease (MCNSD, CC+FC+), OCNSD (CC–FC+), and CNS neg (CC–FC–).
Results. A total of 134 pts (median age 48, range 19–81; M/F 52%/48%) were enrolled across 14 Italian centers. Treatments followed the GIMEMA/NILG or Hyper-CVAD protocols. CNSDT intensification consisted of biweekly IT therapy until CSF blast clearance, followed by two weekly doses. A CR was achieved in 82% of pts (111/134), with a RE in 38/110 (34.5%) including one isolated CNS RE. MCNSD, OCNSD and CNS neg were found in 14 (10%), 10 (8%), and 110 (82%) pts, respectively. CNSDT was intensified in all MCNSD and in 9/10 OCNSD pts. In the OCNSD, the median of the total acquired, and the leukemic events were 4548 (60-79697) and 61 (13-2100), respectively. No significant differences were observed among groups for gender, B/T lineage, cytogenetic/genetic features. Compared with CNS neg group, OCNSD pts had more frequent WBCc >30x109/L (p=.016). The 3-year DFS in CNS neg, OCNSD and MCNSD pts did not differ: 62% (95% CI, 51.1-75), 47% (95% CI, 21.5-100) and 35% (95% CI, 12.3-100), respectively (p<0.42) (Figure).
Conclusions. Our data confirm that OCNSD is uncommon. Apart from high WBC, no other predictive feature was identified, highlighting the need of routine FC in all pts. Although preliminary, our findings suggest that intensified CNSDT may improve prognosis in OCNSD pts, which could be particularly relevant as systemic chemotherapy strategies are being de-escalated.
Recombinant adeno-associated virus (rAAV) vectors stand out as highly promising for in vivo gene transfer, particularly in targeting the skeletal muscle for treating muscular genetic diseases or secreting therapeutic factors. Despite the simplicity and efficacy of the established intramuscular (IM) route, it has been often associated with an immune-induced rapid loss of transgene expression, in particular in large animal models, and generally considered irreversible as a consequence of a cytotoxic elimination of transduced cells. Here, we report in a non-human primate model that transgene expression loss after IM delivery of an rAAV1 expressing an immunogenic protein is only transient, with the re-expression of the transgene lasting up to 5 years post-injection. We show that the recovery of transgene expression is due to persisting viral genomes in the injected muscles despite the detection of peripheral anti-transgene cellular immunity. Persisting genomes were observed in the presence of infiltrated mononuclear CD8 and CD4 T lymphocytes, among which we were able to detect FoxP3+ regulatory cells. This is to our knowledge the first report of a transient immune-mediated loss of gene expression in a large animal model after rAAV delivery that should shed new light on the issue of rAAV vector immunogenicity.
Paulina Czechowicz, Magdalena Gebert, Sylwia Bartoszewska
et al.
Abstract Regulation of endoplasmic reticulum (ER) homeostasis plays a critical role in maintaining cell survival. When ER stress occurs, a network of three pathways called the unfolded protein response (UPR) is activated to reestablish homeostasis. While it is known that there is cross-talk between these pathways, how this complex network is regulated is not entirely clear. Using human cancer and non-cancer cell lines, two different genome-wide approaches, and two different ER stress models, we searched for miRNAs that were decreased during the UPR and surprisingly found only one, miR-1244, that was found under all these conditions. We also verified that ER-stress related downregulation of miR-1244 expression occurred with 5 different ER stressors and was confirmed in another human cell line (HeLa S3). These analyses demonstrated that the outcome of this reduction during ER stress supported both IRE1 signaling and elevated BIP expression. Further analysis using inhibitors specific for IRE1, ATF6, and PERK also revealed that this miRNA is impacted by all three pathways of the UPR. This is the first example of a complex mechanism by which this miRNA serves as a regulatory check point for all 3 pathways that is switched off during UPR activation. In summary, the results indicate that ER stress reduction of miR-1244 expression contributes to the pro-survival arm of UPR.
Saba Gharibi, Bahram Moghimi, Mohammad Bagher Mahmoudi
et al.
Multiple sclerosis (MS) is a chronic autoimmune disease with an unknown etiology. The purpose of this research was to assess miR-223, miR-146a, and miR-193a in acute and chronic phases of experimental autoimmune encephalomyelitis (EAE) mice to consider the possible role of these genes in the pathogenesis of MS. EAE induction was given by myelin oligodendrocyte glycoprotein peptide on female C57BL/6 mice. Clinical scores and other criteria were followed daily until day 21 for the acute group and day 77 for the chronic group. At the end of the course, inflammation and demyelination of the central nervous system (CNS) were assessed by histological analysis. MicroRNA expression levels were assessed by real-time PCR. EAE development attenuated in the chronic group, and histological analysis showed less infiltration and demyelination in the chronic group compared to the acute group. The upper expression of miR-223 is demonstrated in the acute phase of EAE. Moreover, the expression levels of miR-146a and miR-193a decreased in the chronic phase of EAE. MiR-223 showed a highly coordinated elevation in the acute phase both in vivo and in vitro. MiR-146a shares a pathway with miR-223 through effecting IL-6 expression. Further studies are needed to reveal their impact on EAE and possible applications as drug targets and biomarkers.
Heart failure (HF) is an emerging epidemic with a high mortality rate. Apart from conventional treatment methods, such as surgery or use of vasodilation drugs, metabolic therapy has been suggested as a new therapeutic strategy. The heart relies on fatty acid oxidation and glucose (pyruvate) oxidation for ATP-mediated contractility; the former meets most of the energy requirement, but the latter is more efficient. Inhibition of fatty acid oxidation leads to the induction of pyruvate oxidation and provides cardioprotection to failing energy-starved hearts. One of the non-canonical types of sex hormone receptors, progesterone receptor membrane component 1 (<i>Pgrmc1</i>), is a non-genomic progesterone receptor associated with reproduction and fertility. Recent studies revealed that <i>Pgrmc1</i> regulates glucose and fatty acid synthesis. Notably, <i>Pgrmc1</i> has also been associated with diabetic cardiomyopathy, as it reduces lipid-mediated toxicity and delays cardiac injury. However, the mechanism by which <i>Pgrmc1</i> influences the energy-starved failing heart remains unknown. In this study, we found that loss of <i>Pgrmc1</i> inhibited glycolysis and increased fatty acid/pyruvate oxidation, which is directly associated with ATP production, in starved hearts. Loss of <i>Pgrmc1</i> during starvation activated the phosphorylation of AMP-activated protein kinase, which induced cardiac ATP production. <i>Pgrmc1</i> loss increased the cellular respiration of cardiomyocytes under low-glucose conditions. In isoproterenol-induced cardiac injury, <i>Pgrmc1</i> knockout resulted in less fibrosis and low heart failure marker expression. In summary, our results revealed that <i>Pgrmc1</i> ablation in energy-deficit conditions increases fatty acid/pyruvate oxidation to protect against cardiac damage via energy starvation. Moreover, <i>Pgrmc1</i> may be a regulator of cardiac metabolism that switches the dominance of glucose-fatty acid usage according to nutritional status and nutrient availability in the heart.
Krzysztof M. Tokarz, Wojciech Makowski, Barbara Tokarz
et al.
The aim of this study was to examine the effect of the modified light spectrum of glass containing red luminophore on the performance of the photosynthetic apparatus of two types of lettuce cultivated in soil in a greenhouse. Butterhead and iceberg lettuce were cultivated in two types of greenhouses: (1) covered with transparent glass (control) and (2) covered with glass containing red luminophore (red). After 4 weeks of culture, structural and functional changes in the photosynthetic apparatus were examined. The presented study indicated that the red luminophore used changed the sunlight spectrum, providing an adequate blue:red light ratio, while decreasing the red:far-red radiation ratio. In such light conditions, changes in the efficiency parameters of the photosynthetic apparatus, modifications in the chloroplast ultrastructure, and altered proportions of structural proteins forming the photosynthetic apparatus were observed. These changes led to a decrease of CO<sub>2</sub> carboxylation efficiency in both examined lettuce types.
Nazira El-Hage, Matthew J. Haney, Yuling Zhao
et al.
Over the recent decades, the use of extracellular vesicles (EVs) has attracted considerable attention. Herein, we report the development of a novel EV-based drug delivery system for the transport of the lysosomal enzyme tripeptidyl peptidase-1 (TPP1) to treat Batten disease (BD). Endogenous loading of macrophage-derived EVs was achieved through transfection of parent cells with TPP1-encoding <i>p</i>DNA. More than 20% ID/g was detected in the brain following a single intrathecal injection of EVs in a mouse model of BD, ceroid lipofuscinosis neuronal type 2 (CLN2) mice. Furthermore, the cumulative effect of EVs repetitive administrations in the brain was demonstrated. TPP1-loaded EVs (EV-TPP1) produced potent therapeutic effects, resulting in efficient elimination of lipofuscin aggregates in lysosomes, decreased inflammation, and improved neuronal survival in CLN2 mice. In terms of mechanism, EV-TPP1 treatments caused significant activation of the autophagy pathway, including altered expression of the autophagy-related proteins LC3 and P62, in the CLN2 mouse brain. We hypothesized that along with TPP1 delivery to the brain, EV-based formulations can enhance host cellular homeostasis, causing degradation of lipofuscin aggregates through the autophagy–lysosomal pathway. Overall, continued research into new and effective therapies for BD is crucial for improving the lives of those affected by this condition.
Pistachio genetic variety includes a wide range of female variations and male genotypes, and Iran is regarded as one of the critical sites for this diversity in the world. The genus Pistacia consists of eleven species that only have edible nuts and are commercially important. Four important species of pistachios include P. vera, P. khinjuk Stocks, P. eurycarpa Yalt. (P. atlantica subsp. Kurdica Zoh.), and P. atlantica Dsef are found in Iran. Genetic diversity is one aspect of biological diversity that is extremely important for conservation strategies, especially in rare and narrowly endemic species. In Iran, there is no knowledge concerning the genomic organization of the population, genetic diversity, or phenotypic variations of the species. Pistacia khinjuk has eight distinct regional populations, all of which were studied for genetic variation and demographic organization because of the species' therapeutic value. For this reason, we employed six inter-retrotransposon amplified polymorphism (IRAP) indicators and 15 mixed IRAP indicators to highlight genomic variation in this plant both within and across populations in this study. It was discovered that 73% of overall genomic variability was related to within-population variety and 27% was attributable to inter-population genomic divergence using the AMOVA test among the examined populations (PhiPT = 0.49, P = 0.010). It was discovered by the Mantel analysis that there was a substantial positive association between genomic isolation and geographic distance among the tested populations. STRUCTURE analyses and population assignment tests revealed some degree of gene flow among these populations. There was consistency between the PCoA plots of communities and the UPGMA grouping of molecular information. Based on (IRAP) indicators, these findings demonstrated that regional communities of the plant Pistacia khinjuk are well distinct.
Elizabeth R. Stirling, Steven M. Bronson, Jessica D. Mackert
et al.
Expression of immune checkpoint proteins restrict immunosurveillance in the tumor microenvironment; thus, FDA-approved checkpoint inhibitor drugs, specifically PD-1/PD-L1 and CTLA-4 inhibitors, promote a cytotoxic antitumor immune response. Aside from inflammatory signaling, immune checkpoint proteins invoke metabolic reprogramming that affects immune cell function, autonomous cancer cell bioenergetics, and patient response. Therefore, this review will focus on the metabolic alterations in immune and cancer cells regulated by currently approved immune checkpoint target proteins and the effect of costimulatory receptor signaling on immunometabolism. Additionally, we explore how diet and the microbiome impact immune checkpoint blockade therapy response. The metabolic reprogramming caused by targeting these proteins is essential in understanding immune-related adverse events and therapeutic resistance. This can provide valuable information for potential biomarkers or combination therapy strategies targeting metabolic pathways with immune checkpoint blockade to enhance patient response.
Abstract Background The coronavirus disease 2019 (COVID-19) outbreak caused by severe acute respiratory syndrome coronavirus 2 (SARS-Cov-2) has become an ongoing pandemic. Understanding the respiratory immune microenvironment which is composed of multiple cell types, together with cell communication based on ligand–receptor interactions is important for developing vaccines, probing COVID-19 pathogenesis, and improving pandemic control measures. Methods A total of 102 consecutive hospitalized patients with confirmed COVID-19 were enrolled in this study. Clinical information, routine laboratory tests, and flow cytometry analysis data with different conditions were collected and assessed for predictive value in COVID-19 patients. Next, we analyzed public single-cell RNA-sequencing (scRNA-seq) data from bronchoalveolar lavage fluid, which offers the closest available view of immune cell heterogeneity as encountered in patients with varying severity of COVID-19. A weighting algorithm was used to calculate ligand–receptor interactions, revealing the communication potentially associated with outcomes across cell types. Finally, serum cytokines including IL6, IL1β, IL10, CXCL10, TNFα, GALECTIN-1, and IGF1 derived from patients were measured. Results Of the 102 COVID-19 patients, 42 cases (41.2%) were categorized as severe. Multivariate logistic regression analysis demonstrated that AST, D-dimer, BUN, and WBC were considered as independent risk factors for the severity of COVID-19. T cell numbers including total T cells, CD4+ and CD8+ T cells in the severe disease group were significantly lower than those in the moderate disease group. The risk model containing the above mentioned inflammatory damage parameters, and the counts of T cells, with AUROCs ranged from 0.78 to 0.87. To investigate the molecular mechanism at the cellular level, we analyzed the published scRNA-seq data and found that macrophages displayed specific functional diversity after SARS-Cov-2 infection, and the metabolic pathway activities in the identified macrophage subtypes were influenced by hypoxia status. Importantly, we described ligand–receptor interactions that are related to COVID-19 serverity involving macrophages and T cell subsets by communication analysis. Conclusions Our study showed that macrophages driving ligand–receptor crosstalk contributed to the reduction and exhaustion of CD8+ T cells. The identified crucial cytokine panel, including IL6, IL1β, IL10, CXCL10, IGF1, and GALECTIN-1, may offer the selective targets to improve the efficacy of COVID-19 therapy. Trial registration: This is a retrospective observational study without a trial registration number. Video Abstract
Valproic acid (VPA) is an antiepileptic drug found to induce mitochondrial dysfunction and autophagy in cancer cell lines. We treated the SH-SY5Y cell line with various concentrations of VPA (1, 5, and 10 mM). The treatment decreased cell viability, ATP production, and mitochondrial membrane potential and increased reactive oxygen species production. In addition, the mitochondrial DNA copy number increased after VPA treatment in a dose-dependent manner. Western blotting showed that the levels of mitochondrial biogenesis-related proteins (PGC-1α, TFAM, and COX4) increased, though estrogen-related receptor expression decreased after VPA treatment. Further, VPA treatment increased the total and acetylated FOXO3a protein levels. Although SIRT1 expression was decreased, SIRT3 expression was increased, which regulated FOXO3 acetylation in the mitochondria. Furthermore, VPA treatment induced autophagy via increased LC3-II levels and decreased p62 expression and mTOR phosphorylation. We suggest that VPA treatment induces mitochondrial biogenesis and autophagy via changes in FOXO3a expression and posttranslational modification in the SH-SY5Y cell line.
During past decades, survival rates in cancer patients have drastically improved due to the successful development of novel, promising chemical compounds and therapeutic schedules [...]
Abstract Aging is a natural and progressive process characterized by an increased frequency of age-related diseases such as cancer. But its mechanism is unclear. TNFAIP8L2 (Tipe2) is an important negative regulator for homeostasis through inhibiting TLR and TCR signaling. Our work reveals that Tipe2 might have dual function by regulating senescence. One side, the overexpression of Tipe2 in CRC cells could induce typical senescent phenotype, especially exposure to oxidative stress. Tipe2 inhibits telomerase activity by regulating c-Myc and c-Est-2 binding to the hTERT promotor. Interestingly, Tipe2 KO mice treated with D-Gal showed a less serious inverse of CD4:CD8 ratio, a lower percentage of Treg compared to WT. Besides, Tipe2 KO mice were more tolerant to the initiation of AOM/DSS-induced CRC, accompanied by a lower level of Treg within IEL. Therefore, specific antibodies against CD25 effectively ameliorate tumorigenesis. These data suggest strongly that the overexpressed Tipe2 suppresses tumor cells proliferation and survival, but endogenous Tipe2 promotes the initiation of tumorigenesis when exposure to dangerous environment such as AOM/DSS-related inflammation.
Hydration lubrication has emerged as a new paradigm for lubrication in aqueous and biological media, accounting especially for the extremely low friction (friction coefficients down to 0.001) of articular cartilage lubrication in joints. Among the ensemble of molecules acting in the joint, phosphatidylcholine (PC) lipids have been proposed as the key molecules forming, in a complex with other molecules including hyaluronic acid (HA), a robust layer on the outer surface of the cartilage. HA, ubiquitous in synovial joints, is not in itself a good boundary lubricant, but binds the PC lipids at the cartilage surface; these, in turn, massively reduce the friction via hydration lubrication at their exposed, highly hydrated phosphocholine headgroups. An important unresolved issue in this scenario is why the free HA molecules in the synovial fluid do not suppress the lubricity by adsorbing simultaneously to the opposing lipid layers, i.e., forming an adhesive, dissipative bridge between them, as they slide past each other during joint articulation. To address this question, we directly examined the friction between two hydrogenated soy PC (HSPC) lipid layers (in the form of liposomes) immersed in HA solution or two palmitoyl–oleoyl PC (POPC) lipid layers across HA–POPC solution using a surface force balance (SFB). The results show, clearly and surprisingly, that HA addition does not affect the outstanding lubrication provided by the PC lipid layers. A possible mechanism indicated by our data that may account for this is that multiple lipid layers form on each cartilage surface, so that the slip plane may move from the midplane between the opposing surfaces, which is bridged by the HA, to an HA-free interface within a multilayer, where hydration lubrication is freely active. Another possibility suggested by our model experiments is that lipids in synovial fluid may complex with HA, thereby inhibiting the HA molecules from adhering to the lipids on the cartilage surfaces.
Mako Ueda, Hiroshi Tsubamoto, Mina Kashima-Morii
et al.
Aim. To reveal current problems and challenges faced by our gynecologic services department in managing patients with hereditary cancers. Methods. We collected clinical data of patients with hereditary cancers, identified via genetic testing (or clinically diagnosed in cases of Cowden syndrome or Peutz–Jeghers syndrome), and treated in our gynecological department from 2012 to 2018. Results. Fifteen patients had hereditary breast and ovarian cancer (HBOC), 6 had Lynch syndrome, 2 had Cowden syndrome, and 2 had Peutz–Jeghers syndrome. Five patients diagnosed with HBOC were younger than 40 years at diagnosis. Risk-reducing salpingo-oophorectomy (RRSO) was performed on 1 patient with a BRCA1 mutation at age 38 years. Seven patients overall underwent RRSO, and none had malignancies on pathological examinations. Peritoneal washing cytology (PWC) was suspicious for malignancy in one patient; however, subsequent PWC at 6 months after RRSO was negative. A patient with endometrial cancer and Lynch syndrome and a patient with atypical endometrial hyperplasia (AEH) and Cowden syndrome strongly desired fertility preservation. They achieved remission after medroxyprogesterone acetate treatment and multiple dilations and curettages, respectively. One patient with Lynch syndrome developed AEH after 11 years of surveillance. Laparotomy revealed adjacent low-grade and high-grade serous ovarian cancer with positive ascites cytology. She had no recurrence during 7-year follow-up after laparotomy. Conclusion. Managing patients with hereditary cancer, positive or false-positive ascites cytology discovered during RRSO, and desired preservation of fertility is highly challenging.
Daisuke Minami, Yuki Takigawa, Hiroe Kayatani
et al.
A 46‐year‐old male with a superior mediastinal mass presented with a one‐month history of hoarseness and chest pain and was referred to our hospital. Although endobronchial, ultrasound‐guided, transbronchial needle aspiration (EBUS‐TBNA) was initially performed, we could not obtain an adequate specimen because of his severe cough and an inadequate EBUS view. During the same endoscopic session, we performed endoscopic, ultrasound‐guided, bronchoscopic fine‐needle aspiration (EUS‐B‐FNA) via a transoesophageal approach using the BF‐UC290F (Olympus, Tokyo, Japan), a third‐generation EBUS‐TBNA endoscope. The BF‐UC290F enabled smooth access through the oesophagus and a clear EBUS view of the mass, attributable, respectively, to the compact distal tip and the powerful angulation. Rapid on‐site cytology revealed that an adequate specimen had been obtained, and we terminated the procedure without inducing a severe cough. Histologically, the mass was a squamous cell carcinoma. EUS‐B‐FNA employing the BF‐UC290F was useful to diagnose the superior mediastinal mass.