Prospective genetic germline evaluation in a consecutive group of adult patients aged <60 years with myelodysplastic syndromes.

Relevance of germline (GL) predisposition in myelodysplastic syndromes (MDSs) was stressed in both 2022 WHO and International Consensus classifications, but its incidence is probably underestimated, especially in young adult patients. We selected a cohort of 31 consecutive de novo MDS patients with unusual young age (<60 years). We performed exome sequencing (ES) on DNA extracted from noninvasive sources (peripheral blood and saliva), filtering for a panel of 344 genes specifically tailored for detecting GL variants related to clonal and nonclonal cytopenia. We observed at least one high- or low-confidence GL MDS variant in 7/31 (22.6%) and 9/31 (29.0%) of cases, respectively. Four of 31 patients (12.9%) confirmed having established MDS/AML predisposing disorders. We found heterozygous variants in genes involved in DNA repair/cancer predisposition (ATM, ATR, FANCM, PARN, BRCA1, BRCA2, CHEK2, MSH2) in 9/31 (29.0%) cases and variants affecting ribosome biogenesis (SBDS), hematopoietic stem cell (GATA2), and megakaryocyte (ANKRD26) differentiation in single cases. Two cases had variants in RBBP6, a gene previously described exclusively in familial myeloproliferative neoplasms. Lastly, four cases had variants in genes related to inherited anemias (CUBN and PIEZO1 genes). Our results showed that "young" MDS patients aged 40-60 years carried reported and unreported GL variants with an unexpectedly high proportion, and these events co-occurred with somatic mutations recurrent in myeloid neoplasms. We explored the "no man's land" of the young adult MDS cases adopting a practical and scalable diagnostic tool, capable to detect GL variants avoiding invasive methods.

Chlorpromazine overcomes temozolomide resistance in glioblastoma by inhibiting Cx43 and essential DNA repair pathways.

BACKGROUND: In the fight against GBM, drug repurposing emerges as a viable and time-saving approach to explore new treatment options. Chlorpromazine, an old antipsychotic medication, has recently arisen as a promising candidate for repositioning in GBM therapy in addition to temozolomide, the first-line standard of care. We previously demonstrated the antitumor efficacy of chlorpromazine and its synergistic effects with temozolomide in suppressing GBM cell malignant features in vitro. This prompted us to accomplish a Phase II clinical trial to evaluate the efficacy and safety of adding chlorpromazine to temozolomide in GBM patients with unmethylated MGMT gene promoter. In this in vitro study, we investigate the potential role of chlorpromazine in overcoming temozolomide resistance. METHODS: In our experimental set, we analyzed Connexin-43 expression at both the transcriptional and protein levels in control- and chlorpromazine-treated GBM cells. DNA damage and subsequent repair were assessed by immunofluorescence of γ-H2AX and Reverse-Phase Protein microArrays in chlorpromazine treated GBM cell lines. To elucidate the relationship between DNA repair systems and chemoresistance, we analyzed a signature of DNA repair genes in GBM cells after treatment with chlorpromazine, temozolomide and Connexin-43 downregulation. RESULTS: Chlorpromazine treatment significantly downregulated connexin-43 expression in GBM cells, consequently compromising connexin-dependent cellular resilience, and ultimately contributing to cell death. In line with this, we observed concordant post-translational modifications of molecular determinants involved in DNA damage and repair pathways. Our evaluation of DNA repair genes revealed that temozolomide elicited an increase, while chlorpromazine, as well as connexin-43 silencing, a decrease in DNA repair gene expression in GBM cells. CONCLUSIONS: Chlorpromazine potentiates the cytotoxic effects of the alkylating agent temozolomide through a mechanism involving downregulation of Cx43 expression and disruption of the cell cycle arrest essential for DNA repair processes. This finding suggests that chlorpromazine may be a potential therapeutic strategy to overcome TMZ resistance in GBM cells by inhibiting their DNA repair mechanisms.

Cardiac rupture as a life-threatening outcome of Takotsubo syndrome: A systematic review.

BACKGROUND: Takotsubo syndrome (TS) is a reversible cause of heart failure; however, a minority of patients can develop serious complications, including cardiac rupture (CR). OBJECTIVES: Analyze case reports of CR related to TS, detailing patient characteristics to uncover risk factors and prognosis for this severe complication. METHODS: We conducted a systematic search of MEDLINE and Embase databases to identify case reports of patients with TS complicated by CR, from inception to October 2023. RESULTS: We included 44 subjects (40 females; 4 males) with a median age of 75 (IQR 71-82) years, of White/Caucasian (61%) or East Asian/Japanese (39%) ethnicity. An emotional trigger was present in 15 (34%) subjects and an apical ballooning pattern was observed in all cases (100%). ST-segment elevation was reported in 39 (93%) of 42 cases, with the anterior myocardial segments (37 [88%]) being the most compromised, followed by lateral (26 [62%]) and inferior (14 [33%]) segments. The median time to cardiac rupture was 48 (5-120) hours since admission, with the left ventricular free wall (25 [57%]) being the most frequent site of perforation. Surgery was attempted in 16 (36%) cases, and 28 (64%) patients did not survive. CONCLUSIONS: CR related to TS is a rare complication associated with high mortality and affecting elderly females, specially from White/Caucasian or East Asian/Japanese descent, presenting with anterior or lateral ST-segment elevation, and an apical ballooning pattern. Although data is limited and additional prospective studies are needed, the awareness of this life-threatening complication is crucial to early identify high-risk patients. CONDENSED ABSTRACT: Cardiac rupture is a rare complication of Takotsubo syndrome. We conducted a systematic review of cases complicated by cardiac rupture, and we identified 44 subjects (40 females and 4 males) with a median age of 75 (IQR 71-82) years, of White/Caucasian (61%) or East Asian/Japanese (39%) ethnicity, all with an apical ballooning pattern (100%). The median time to cardiac rupture was 48 (5-120) hours since admission, with the left ventricular free wall (25 [57%]) being the most frequent site of perforation. Surgery treatment was attempted in 16 (36%) cases, and 28 (64%) patients did not survive.

Retinoic Acid-Mediated Control of Energy Metabolism Is Essential for Lung Branching Morphogenesis.

Lung branching morphogenesis relies on intricate epithelial-mesenchymal interactions and signaling networks. Still, the interplay between signaling and energy metabolism in shaping embryonic lung development remains unexplored. Retinoic acid (RA) signaling influences lung proximal-distal patterning and branching morphogenesis, but its role as a metabolic modulator is unknown. Hence, this study investigates how RA signaling affects the metabolic profile of lung branching. We performed ex vivo lung explant culture of embryonic chicken lungs treated with DMSO, 1 µM RA, or 10 µM BMS493. Extracellular metabolite consumption/production was evaluated by using 1H-NMR spectroscopy. Mitochondrial respiration and biogenesis were also analyzed. Proliferation was assessed using an EdU-based assay. The expression of crucial metabolic/signaling components was examined through Western blot, qPCR, and in situ hybridization. RA signaling stimulation redirects glucose towards pyruvate and succinate production rather than to alanine or lactate. Inhibition of RA signaling reduces lung branching, resulting in a cystic-like phenotype while promoting mitochondrial function. Here, RA signaling emerges as a regulator of tissue proliferation and lactate dehydrogenase expression. Furthermore, RA governs fatty acid metabolism through an AMPK-dependent mechanism. These findings underscore RA's pivotal role in shaping lung metabolism during branching morphogenesis, contributing to our understanding of lung development and cystic-related lung disorders.

Immune-monitoring of myelodysplastic neoplasms: Recommendations from the i4MDS consortium.

Advancements in comprehending myelodysplastic neoplasms (MDS) have unfolded significantly in recent years, elucidating a myriad of cellular and molecular underpinnings integral to disease progression. While molecular inclusions into prognostic models have substantively advanced risk stratification, recent revelations have emphasized the pivotal role of immune dysregulation within the bone marrow milieu during MDS evolution. Nonetheless, immunotherapy for MDS has not experienced breakthroughs seen in other malignancies, partly attributable to the absence of an immune classification that could stratify patients toward optimally targeted immunotherapeutic approaches. A pivotal obstacle to establishing "immune classes" among MDS patients is the absence of validated accepted immune panels suitable for routine application in clinical laboratories. In response, we formed International Integrative Innovative Immunology for MDS (i4MDS), a consortium of multidisciplinary experts, and created the following recommendations for standardized methodologies to monitor immune responses in MDS. A central goal of i4MDS is the development of an immune score that could be incorporated into current clinical risk stratification models. This position paper first consolidates current knowledge on MDS immunology. Subsequently, in collaboration with clinical and laboratory specialists, we introduce flow cytometry panels and cytokine assays, meticulously devised for clinical laboratories, aiming to monitor the immune status of MDS patients, evaluating both immune fitness and identifying potential immune "risk factors." By amalgamating this immunological characterization data and molecular data, we aim to enhance patient stratification, identify predictive markers for treatment responsiveness, and accelerate the development of systems immunology tools and innovative immunotherapies.

Exposure to toxicologically relevant atrazine concentrations impair the glycolytic function of mouse Sertoli cells through the downregulation of lactate dehydrogenase.

Atrazine (ATZ), a widely used herbicide with potent endocrine-disrupting properties, has been implicated in hormonal disturbances and fertility issues. Sertoli cells (SCs) play a crucial role in providing mechanical and nutritional support of spermatogenesis. Herein, we aimed to study the effects of environmentally relevant ATZ concentrations on the nutritional support of spermatogenesis provided by SCs. For that, mouse SCs (TM4) were exposed to increasing ATZ concentrations (in µg/L: 0.3, 3, 30, 300, or 3000). After 24 h, cellular proliferation and metabolic activity were assessed. Mitochondrial activity and endogenous reactive oxygen species (ROS) production were evaluated using JC-1 and CM-H2DCFDA probes, respectively. We also analyzed protein levels of lactate dehydrogenase (LDH) using Western Blot and live cells glycolytic function through Seahorse XF Glycolysis Stress Test Kit. ATZ exposure decreased the activity of oxidoreductases in SCs, suggesting a decreased metabolic activity. Although ATZ is reported to induce oxidative stress, we did not observe alterations in mitochondrial membrane potential and ROS production across all tested concentrations. When we evaluated the glycolytic function of SCs, we observed that ATZ significantly impaired glycolysis and the glycolytic capacity at all tested concentrations. These results were supported by the decreased expression of LDH in SCs. Overall, our findings suggest that ATZ impairs the glycolytic function of SCs through LDH downregulation. Since lactate is the preferential energetic substrate for germ cells, exposure to ATZ may detrimentally impact the nutritional support crucial for spermatogenesis, hinting for a relationship between ATZ exposure and male infertility.

Management and outcome of fetal abdominal cysts in first trimester: systematic review of the literature.

OBJECTIVES: The finding of an abdominal cyst during pregnancy has an estimated prevalence of 1 in 1000 pregnancies, mostly in second and third trimester. The detection of a fetal abdominal cyst during the first trimester scan is a rare event, whose natural history and prognosis are often unknown and unpredictable as these anomalies can be related to various underlying conditions and originate from different structures. The aim of this study is to evaluate the outcome of fetal abdominal cysts detected in the first trimester in order to understand their possible clinical significance and to offer the proper management according to the available data. METHODS: We present a case report of a first trimester fetal abdominal cyst detected with subsequent diagnosis of congenital multiple arthrogryposis and we performed a systematic review of the literature to identify the incidence and the outcomes of similar cases. The systematic literature review was conducted in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement 25 and registered with PROSPERO (CRD42023491729). RESULTS: A total of 60 cases of first trimester abdominal cysts were included. Of these, 35% were associated with concurrent or late onset structural anomalies, as in our case report, and 65% were isolated. In pregnancies with isolated fetal abdominal cysts, 56% had a completely normal outcome. CONCLUSIONS: The finding of an abdominal cyst during the first trimester of pregnancy is in most cases an isolated event with a moderate to good prognosis but it could also be an early sign of other associated abnormalities, including arthrogryposis. Increased ultrasound surveillance and additional genetic testing to rule out possible associated anomalies are pivotal to assess the risk of adverse pregnancy outcomes and to provide appropriate counselling to the patient. This article is protected by copyright. All rights reserved.

Laparoscopic heller myotomy versus peroral endoscopic myotomy for the treatment of achalasia.

PURPOSE OF REVIEW: To compare different therapeutic modalities and determine their role in the treatment of esophageal achalasia. RECENT FINDINGS: The last 3 decades have seen a significant improvement in the diagnosis and treatment of esophageal achalasia. Conventional manometry has been replaced by high-resolution manometry, which has determined a more precise classification of achalasia in three subtypes, with important treatment implications. Therapy, while still palliative, has evolved tremendously. While pneumatic dilatation was for a long time the main choice of treatment, this approach slowly changed at the beginning of the nineties when minimally invasive surgery was adopted, initially thoracoscopically and then laparoscopically with the addition of partial fundoplication. And in 2010, the first report of a new endoscopic technique - peroral endoscopic myotomy (POEM) - was published, revamping the interest in the endoscopic treatment of achalasia. SUMMARY: This review focuses particularly on the comparison of POEM and laparoscopic Heller myotomy (LHM) with partial fundoplication as primary treatment modality for esophageal achalasia. Based on the available data, we believe that LHM with partial fundoplication should be the primary treatment modality in most patients. POEM should be selected when surgical expertise is not available, for type III achalasia, for the treatment of recurrent symptoms, and for patients who had prior abdominal operations that would make LHM challenging and unsafe.

The evolution of the treatment of esophageal achalasia. Chronicle of a 35-year journey.

Over the last few decades, significant improvement has been made in both the evaluation and treatment of esophageal achalasia. The Chicago classification, today in version 4.0, is now the standard for diagnosis of achalasia, providing a classification into 3 subtypes with important therapeutic and prognostic implications. Therapy, which was at first mostly limited to pneumatic dilatation, today includes minimally invasive surgery and peroral endoscopic myotomy, allowing for a more tailored approach to patients and better treatment of recurrent symptoms. This review chronicles my personal experience with achalasia over the last 35 years, describing the progress made in the treatment of patients with achalasia.

The connection of motor improvement after deep brain stimulation in Parkinson's disease and microstructural integrity of the substantia nigra and subthalamic nucleus.

BACKGROUND: Nigrostriatal microstructural integrity has been suggested as a biomarker for levodopa response in Parkinson's disease (PD), which is a strong predictor for motor response to deep brain stimulation (DBS) of the subthalamic nucleus (STN). This study aimed to explore the impact of microstructural integrity of the substantia nigra (SN), STN, and putamen on motor response to STN-DBS using diffusion microstructure imaging. METHODS: Data was collected from 23 PD patients (mean age 63 ± 7, 6 females) who underwent STN-DBS, had preoperative 3 T diffusion magnetic resonance imaging including multishell diffusion-weighted MRI with b-values of 1000 and 2000 s/mm2 and records of motor improvement available. RESULTS: The association between a poorer DBS-response and increased free interstitial fluid showed notable effect sizes (rho > |0.4|) in SN and STN, but not in putamen. However, this did not reach significance after Bonferroni correction and controlling for sex and age. CONCLUSION: Microstructural integrity of SN and STN are potential biomarkers for the prediction of therapy efficacy following STN-DBS, but further studies are required to confirm these associations.

Relevance of real-time analyzers to determine mitochondrial quality in endothelial cells and oxidative stress in preeclampsia.

Oxidative stress and mitochondrial dysfunction are important elements for the pathophysiology of preeclampsia (PE), a multisystemic hypertensive syndrome of pregnancy, characterized by endothelial dysfunction and responsible for a large part of maternal and fetal morbidity and mortality worldwide. Researchers have dedicated their efforts to unraveling the intricate ways in which certain molecules influence both energy metabolism and oxidative stress. Exploring established methodologies from existing literature, shows that these investigations predominantly focus on the placenta, identified as a pivotal source that drives the changes observed in the disease. In this review, we discuss the role of oxidative stress in pathophysiology of PE, as well as metabolic/endothelial dysfunction. We further discuss the use of seahorse analyzers to study real-time bioenergetics of endothelial cells. Although the benefits are clear, few studies have presented results using this method to assess mitochondrial metabolism in these cells. We performed a search on MEDLINE/PubMed using the terms "Seahorse assay and endothelial dysfunction in HUVEC" as well as "Seahorse assay and preeclampsia". From our research, we selected 16 original peer-review papers for discussion. Notably, the first search retrieved studies involving Human Umbilical Vein Endothelial Cells (HUVECs) but none investigating bioenergetics in PE while the second search retrieved studies exploring the technique in PE but none of the studies used HUVECs. Additional studies are required to investigate real-time mitochondrial bioenergetics in PE. Clearly, there is a need for more complete studies to examine the nuances of mitochondrial bioenergetics, focusing on the contributions of HUVECs in the context of PE.

Signatures of metabolic diseases on spermatogenesis and testicular metabolism.

Diets leading to caloric overload are linked to metabolic disorders and reproductive function impairment. Metabolic and hormonal abnormalities stand out as defining features of metabolic disorders, and substantially affect the functionality of the testis. Metabolic disorders induce testicular metabolic dysfunction, chronic inflammation and oxidative stress. The disruption of gastrointestinal, pancreatic, adipose tissue and testicular hormonal regulation induced by metabolic disorders can also contribute to a state of compromised fertility. In this Review, we will delve into the effects of high-fat diets and metabolic disorders on testicular metabolism and spermatogenesis, which are crucial elements for male reproductive function. Moreover, metabolic disorders have been shown to influence the epigenome of male gametes and might have a potential role in transmitting phenotype traits across generations. However, the existing evidence strongly underscores the unmet need to understand the mechanisms responsible for transgenerational paternal inheritance of male reproductive function impairment related to metabolic disorders. This knowledge could be useful for developing targeted interventions to prevent, counteract, and most of all break the perpetuation chain of male reproductive dysfunction associated with metabolic disorders across generations.

Hit-to-Lead Identification and Validation of a Triaromatic Pleuromutilin Antibiotic Candidate.

Herein, we report the hit-to-lead identification of a drug-like pleuromutilin conjugate 16, based on a triaromatic hit reported in 2020. The lead arose as the clear candidate from a hit-optimization campaign in which Gram-positive antibacterial activity, solubility, and P-gp affinity were optimized. Conjugate 16 was extensively evaluated for its in vitro ADMET performance which, apart from solubility, was overall on par with lefamulin. This evaluation included Caco-2 cell permeability, plasma protein binding, hERG inhibition, cytotoxicity, metabolism in microsomes and CYP3A4, resistance induction, and time-kill kinetics. Intravenous pharmacokinetics of 16 proved satisfactory in both mice and pigs; however, oral bioavailability was limited likely due to insufficient solubility. The in vivo efficacy was evaluated in mice, systemically infected with Staphylococcus aureus, where 16 showed rapid reduction in blood bacteriaemia. Through our comprehensive studies, lead 16 has emerged as a highly promising and safe antibiotic candidate for the treatment of Gram-positive bacterial infections.

Modern insights into the pathophysiology and treatment of pseudoachalasia.

BACKGROUND: Secondary achalasia or pseudoachalasia is a clinical presentation undistinguishable from achalasia in terms of symptoms, manometric, and radiographic findings, but associated with different and identifiable underlying causes. METHODS: A literature review was conducted on the PubMed database restricting results to the English language. Key terms used were "achalasia-like" with 63 results, "secondary achalasia" with 69 results, and "pseudoachalasia" with 141 results. References of the retrieved papers were also manually reviewed. RESULTS: Etiology, diagnosis, and treatment were reviewed. CONCLUSIONS: Pseudoachalasia is a rare disease. Most available evidence regarding this condition is based on case reports or small retrospective series. There are different causes but all culminating in outflow obstruction. Clinical presentation and image and functional tests overlap with primary achalasia or are inaccurate, thus the identification of secondary achalasia can be delayed. Inadequate diagnosis leads to futile therapies and could worsen prognosis, especially in neoplastic disease. Routine screening is not justifiable; good clinical judgment still remains the best tool. Therapy should be aimed at etiology. Even though Heller's myotomy brings the best results in non-malignant cases, good clinical judgment still remains the best tool as well.

Effects of diabetes-induced hyperglycemia on epigenetic modifications and DNA packaging and methylation during spermatogenesis; A narrative review.

The impact of diabetes on various organs failure including testis has been highlighted during the last decades. If on one hand diabetes-induced hyperglycemia has a key role in induced damages; on the other hand, glucose deprivation plays a key role in inducing male infertility. Indeed, glucose metabolism during spermatogenesis has been highlighted due to post-meiotic germ cells drastic dependence on glucose-derived metabolites, especially lactate. In fact, hyperglycemia-induced spermatogenesis arrest has been demonstrated in various studies. Moreover, various sperm maturation processes related to sperm function such as motility are directly depending on glucose metabolism in Sertoli cells. It has been demonstrated that diabetes-induced hyperglycemia adversely impacts sperm morphology, motility and DNA integrity, leading to infertility. However, fertility quality is another important factor to be considered. Diabetes-induced hyperglycemia is not only impacting sperm functions, but also affecting sperm epigenome. DNA packing process and epigenetics modifications occur during spermatogenesis process, determining next generation genetic quality transmitted through sperm. Critical damages may occur due to under- or downregulation of key proteins during spermatogenesis. Consequently, unpacked DNA is more exposed to oxidative stress, leading to intensive DNA damages. Moreover, epigenetic dysregulation occurred during spermatogenesis may impact embryo quality and be transmitted to next generations, increasing offspring genetic issues. Herein we discuss the mechanisms by which diabetes-induced hyperglycemia can affect epigenetic modifications and DNA packaging and methylation during spermatogenesis thus promoting long-lasting effects to the next generation.

Comparative phylogenomics and phylotranscriptomics provide insights into the genetic complexity of nitrogen-fixing root-nodule symbiosis.

Plant root-nodule symbiosis (RNS) with mutualistic nitrogen-fixing bacteria is restricted to a single clade of angiosperms, the Nitrogen-Fixing Nodulation Clade (NFNC), and is best understood in the legume family. Nodulating species share many commonalities, explained either by divergence from a common ancestor over 100 million years ago or by convergence following independent origins over that same time period. Regardless, comparative analyses of diverse nodulation syndromes can provide insights into constraints on nodulation-what must be acquired or cannot be lost for a functional symbiosis-and the latitude for variation in the symbiosis. However, much remains to be learned about nodulation, especially outside of legumes. Here, we employed a large-scale phylogenomic analysis across 88 species, complemented by 151 RNA-seq libraries, to elucidate the evolution of RNS. Our phylogenomic analyses further emphasize the uniqueness of the transcription factor NIN as a master regulator of nodulation and identify key mutations that affect its function across the NFNC. Comparative transcriptomic assessment revealed nodule-specific upregulated genes across diverse nodulating plants, while also identifying nodule-specific and nitrogen-response genes. Approximately 70% of symbiosis-related genes are highly conserved in the four representative species, whereas defense-related and host-range restriction genes tend to be lineage specific. Our study also identified over 900 000 conserved non-coding elements (CNEs), over 300 000 of which are unique to sampled NFNC species. NFNC-specific CNEs are enriched with the active H3K9ac mark and are correlated with accessible chromatin regions, thus representing a pool of candidate regulatory elements for genes involved in RNS. Collectively, our results provide novel insights into the evolution of nodulation and lay a foundation for engineering of RNS traits in agriculturally important crops.