RESUMEN

Glioblastoma (GBM) is the most common malignant primary brain tumor, resulting in poor survival despite aggressive therapies. GBM is characterized by a highly heterogeneous and immunosuppressive tumor microenvironment (TME) made up predominantly of infiltrating peripheral immune cells. One significant immune cell type that contributes to glioma immune evasion is a population of immunosuppressive cells, termed myeloid-derived suppressor cells (MDSCs). Previous studies suggest that a subset of myeloid cells, expressing monocytic (M)-MDSC markers and dual expression of chemokine receptors CCR2 and CX3CR1, utilize CCR2 to infiltrate the TME. This study evaluated the mechanism of CCR2+/CX3CR1+ M-MDSC differentiation and T cell suppressive function in murine glioma models. We determined that bone marrow-derived CCR2+/CX3CR1+ cells adopt an immune suppressive cell phenotype when cultured with glioma-derived factors. Glioma-secreted CSF1R ligands M-CSF and IL-34 were identified as key drivers of M-MDSC differentiation while adenosine and iNOS pathways were implicated in the M-MDSC suppression of T cells. Mining a human GBM spatial RNAseq database revealed a variety of different pathways that M-MDSCs utilize to exert their suppressive function that is driven by complex niches within the microenvironment. These data provide a more comprehensive understanding of the mechanism of M-MDSCs in glioblastoma.

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BACKGROUND: G6PD (glucose-6-phosphate-dehydrogenase) is a key enzyme in the glycolytic pathway and has been implicated in the pathogenesis of cancer and pulmonary hypertension-associated vascular remodeling. Here, we investigated the role of an X-linked G6pd mutation (N126D polymorphism), which is known to increase the risk of cardiovascular disease in individuals from sub-Saharan Africa and many others with African ancestry, in the pathogenesis of pulmonary hypertension induced by a vascular endothelial cell growth factor receptor blocker used for treating cancer. METHODS AND RESULTS: CRISPR-Cas9 genome editing was used to generate the G6pd variant (N126D; G6pdN126D) in rats. A single dose of the vascular endothelial cell growth factor receptor blocker sugen-5416 (SU; 20 mg/kg in DMSO), which is currently in a Phase 2/3 clinical trial for cancer treatment, was subcutaneously injected into G6pdN126D rats and their wild-type littermates. After 8 weeks of normoxic conditions, right ventricular pressure and hypertrophy, pulmonary artery remodeling, the metabolic profile, and cytokine expression were assessed. Right ventricular pressure and pulmonary arterial wall thickness were increased in G6PDN126D+SU/normoxic rats. Simultaneously, levels of oxidized glutathione, inositol triphosphate, and intracellular Ca2+ were increased in the lungs of G6PDN126D+SU/normoxic rats, whereas nitric oxide was decreased. Also increased in G6PDN126D+SU/normoxic rats were pulmonary levels of plasminogen activator inhibitor-1, thrombin-antithrombin complex, and expression of proinflammatory cytokines CCL3 (chemokine [C-C motif] ligand), CCL5, and CCL7. CONCLUSIONS: Our results suggest G6PDN126D increases inositol triphosphate-Ca2+ signaling, inflammation, thrombosis, and hypertrophic pulmonary artery remodeling in SU-treated rats. This suggests an increased risk of vascular endothelial cell growth factor receptor blocker-induced pulmonary hypertension in those carrying this G6PD variant.

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BACKGROUND & AIMS: The size and function of the enteric nervous system (ENS) can vary substantially between individuals. Since ENS function is involved in the etiology of growing number of common human diseases understanding mechanisms which regulate ENS variation is important. METHODS: We analyzed RNAseq data from 41 normal adult human colon biopsies and single-cell RNA-seq data from human and mouse developing gut. To establish cause-consequence relationship we utilized alleles in mice which allow levels change of the candidate effector molecule in the comparable range to human samples. We employed siRNA and primary neuronal cultures to define downstream molecular events and characterized gut functional changes in mice where molecular phenotypes paralleled findings in humans. RESULTS: We found that GDNF levels in the human colon vary about 5-fold and correlate strongly with nitrergic marker expression. In mice, we defined that GDNF levels are regulated via its 3' untranslated region (3' UTR) in the GI tract and observed similar correlation between GDNF levels and nitrergic lineage development. We identified miR-9 and miR-133 as evolutionarily conserved candidates for negative regulation of GDNF expression in the GI tract. Functionally, an increase in inhibitory nitrergic innervation results in an increase in GI tract transit time, stool size and water content accompanied with modestly reduced epithelial barrier function. Mechanistically, we found that GDNF levels regulate nitrergic lineage development via induction of transcription factor ETV1, corroborated by single-cell gene expression data in human and mouse developing enteric neurons. CONCLUSION: Our results reveal how normal variation in GDNF levels influence ENS size, composition and gut function, suggesting a mechanism for well-known interindividual variation among those parameters.

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Previously, we showed the antitumor activity of the new NOS/PDK inhibitor T1084 (1-isobutanoyl-2-isopropylisothiourea dichloroacetate). The present study included an assessment of in vitro cytotoxicity against human malignant and normal cells according to the MTT-test and in vivo antitumor effects in solid tumor models in comparison with precursor compounds T1023 (NOS inhibitor; 1-isobutanoyl-2-isopropylisothiourea hydrobromide) and Na-DCA (PDK inhibitor; sodium dichloroacetate), using morphological, histological, and immunohistochemical methods. The effects of T1084 and T1023 on the in vitro survival of normal (MRC-5) and most malignant cells (A375, MFC-7, K562, OAW42, and PC-3) were similar and quantitatively equal. At the same time, melanoma A375 cells showed 2-2.5 times higher sensitivity (IC50: 0.39-0.41 mM) to the cytotoxicity of T1023 and T1084 than other cells. And only HeLa cells showed significantly higher sensitivity to the cytotoxicity of T1084 compared to T1023 (IC50: 0.54 ± 0.03 and 0.81 ± 0.02 mM). Comparative studies of the in vivo antitumor effects of Na-DCA, T1023, and T1084 on CC-5 cervical cancer and B-16 melanoma in mice were conducted with subchronic daily i.p. administration of these agents at an equimolar dose of 0.22 mmol/kg (33.6, 60.0, and 70.7 mg/kg, respectively). Cervical cancer CC-5 fairly quickly evaded the effects of both Na-DCA and T1023. So, from the end of the first week of Na-DCA or T1023 treatment, the tumor growth inhibition (TGI) began to decrease from 40% to an insignificant level by the end of the observation. In contrast, in two independent experiments, CC-5 showed consistently high sensitivity to the action of T1084: a significant antitumor effect with high TGI (43-58%) was registered throughout the observation, without any signs of neoplasia adaptation. The effect of precursor compounds on melanoma B-16 was either minimal (for Na-DCA) or moderate (for T1023) with TGI only 33%, which subsequently decreased by the end of the experiment. In contrast, the effect of T1084 on B-16 was qualitatively more pronounced and steadily increasing; it was accompanied by a 3-fold expansion of necrosis and dystrophy areas, a decrease in proliferation, and increased apoptosis of tumor cells. Morphologically, the T1084 effect was 2-fold superior to the effects of T1023-the TGI index reached 59-62%. This study suggests that the antitumor effects of T1084 develop through the interaction of NOS-dependent and PDK-dependent pathophysiological effects of this NOS/PDK inhibitor. The NOS inhibitory activity of T1084 exerts an anti-angiogenic effect on neoplasia. At the same time, the PDK inhibitory activity of T1084 enhances the cytotoxicity of induced intratumoral hypoxia and suppresses the development of neoplasia adaptation to anti-angiogenic stress. Such properties allow T1084 to overcome tumor resistance and realize a stable synergistic antitumor effect.

Asunto(s)

Antineoplásicos , Humanos , Animales , Ratones , Antineoplásicos/farmacología , Óxido Nítrico Sintasa/antagonistas & inhibidores , Óxido Nítrico Sintasa/metabolismo , Línea Celular Tumoral , Tiourea/análogos & derivados , Tiourea/farmacología , Tiourea/uso terapéutico , Ensayos Antitumor por Modelo de Xenoinjerto , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora/antagonistas & inhibidores , Femenino , Inhibidores Enzimáticos/farmacología , Supervivencia Celular/efectos de los fármacos , Células HeLa

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Caveolin is a structural protein within caveolae that may be involved in transmembrane molecular transport and/or various intercellular interactions within cells. Specific mutations of caveolin-3 in muscle fibers are well known to cause limb-girdle muscular dystrophy. Altered expression of caveolin-3 has also been detected in Duchenne muscular dystrophy, which may be a part of the pathological process leading to muscle weakness. Interestingly, it has been shown that the renovation of nitric oxide synthase (NOS) in sarcolemma with muscular dystrophy could improve muscle health, suggesting that NOS may be involved in the pathology of muscular dystrophy. Here, we summarize the notable function of caveolin and/or NOS in skeletal muscle fibers and discuss their involvement in the pathology as well as possible tactics for the innovative treatment of muscular dystrophies.

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Caveolina 3 , Distrofias Musculares , Óxido Nítrico Sintasa , Animales , Humanos , Caveolas/metabolismo , Caveolina 3/metabolismo , Caveolina 3/genética , Caveolinas/metabolismo , Distrofias Musculares/metabolismo , Distrofias Musculares/genética , Distrofias Musculares/patología , Distrofia Muscular de Duchenne/metabolismo , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/patología , Mutación , Óxido Nítrico Sintasa/metabolismo , Óxido Nítrico Sintasa/genética

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BACKGROUND: Lennert lymphoma (LL) is a variant of peripheral T-cell lymphoma, not otherwise specified (PTCL, NOS), also known as a lymphoepithelioid variant of PTCL. Because of the rarity and lack of clear-cut diagnostic criteria, LL is susceptible tomisdiagnosis. Although previously diagnosed with LL might be reclassified and evaluated with the advent of of molecular and/or genetic findings, cytomorphology and immunohistochemistry are still the key to give rise to correct diagnosis. CASE PRESENTATION: We report a case of a patient who was diagnosed as LL based on cytomorphology and immunohistochemistry. Routine stain (Hematoxlin and Eosin-H&E) revealed tumor cells were mainly small to medium-sized CD4(+) T cells, the CD8 +/TIA-1 + cytotoxic cells were less minority, no expressions of follicle helper T cell markers (CD10, BCL6, PD1, CXCL13, ICOS) or CD21(+) hyperplastic FDC network, or proliferation of high edndothelial venules were noted; however, numerous epithelioid histiocytes are noted in the background and scattered EBV(+) cells were also present. The patient was achieved complete remission after six courses of chemotherapy with cyclophosphamide, epirubicin, vincristine, etoposide, and prednisone regimen. She was followed for 5 years without recurrence or progression. CONCLUSIONS: Classic LL is not difficult to diagnose by cytomorphology and immunohistochemistry, and the mutation profiles can be helpful to distinguish LL from other lymphomas.

Asunto(s)

Protocolos de Quimioterapia Combinada Antineoplásica , Biomarcadores de Tumor , Inmunohistoquímica , Linfoma de Células T Periférico , Humanos , Biomarcadores de Tumor/análisis , Linfoma de Células T Periférico/patología , Linfoma de Células T Periférico/diagnóstico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Femenino , Persona de Mediana Edad , Resultado del Tratamiento , Valor Predictivo de las Pruebas

RESUMEN

Peripheral T-cell lymphomas (PTCLs) are an aggressive form of non-Hodgkin lymphomas. PTCLs have multiple subtypes, with PTCL not otherwise specified (PTCL-NOS) being the most common. This subtype usually has a high rate of relapse. Making an accurate diagnosis requires molecular genetic analyses, histopathological examination, and immunophenotyping. Treatment for PTCL traditionally starts with the CHOP regimen (cyclophosphamide, doxorubicin, vincristine, and prednisone). We present a case of a patient with PTCL-NOS who progressed despite multiple treatment regimens, including both traditional and novel therapeutic agents, and finally achieved good results with azacytidine, selected based on a TET2 mutation. This case proposes future research into Azacytidine's efficacy in this patient population and further exploration of the broader utility of epigenetic therapies in PTCL.

RESUMEN

AIM: Molecular alterations of diabetic gastroenteropathy are poorly identified. This study investigates the effects of prolonged GABA supplementation on key protein expression levels of trypsin-1, PAR-1, PAR-2, PAR-3, PI3K, Akt, COX-2, GABAA, and GABAB receptors in the gastric tissue of type 2 diabetic rats (T2DM). METHOD: -To induce T2DM, a 3-month high-fat diet and 35 mg/kg of streptozotocin was used. Twenty-four male Wistar rats were divided into 4 groups: (1) control, (2) T2DM, (3) insulin-treated (2.5 U/kg), and (4) GABA-treated (1.5 g/kg GABA). Blood glucose was measured weekly. The protein expressions were assessed using western blotting. Histopathological changes were examined by H&E and Masson's staining. RESULTS: -Diabetic rats show reduced NOS1 and elevated COX-2 and trypsin-1 protein expression levels in gastric tissue. Insulin and GABA therapy restored the NOS1 and COX-2 levels to control values. Insulin treatment increased PI3K, Akt, and p-Akt and, decreased trypsin-1, PAR-1, PAR-2, and PAR-3 levels in the diabetic rats. Levels of GABAA and GABAB receptors normalized following insulin and GABA therapy. H&E staining indicated an increase in mucin secretion following GABA treatment. CONCLUSION: -These results suggest that GABA by acting on GABA receptors may regulate the trypsin-1/PARs/Akt/COX-2 pathway and thereby improve complications of diabetic gastroenteropathy.

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Objective: Mitochondrial dysfunction is closely linked to chronic disorders. This study aims to explore the correlation between pediatric anemia and mitochondrial markers, specifically fibroblast growth factor 21 (FGF21), growth/differentiation factor 15 (GDF-15), and nitric oxide synthase (eNOS). Method: This study included 66 children, with 34 diagnosed with anemia and 32 in the healthy control group. Statistically significant biomarkers were determined through cutoff levels. Results: Among the participants, 34 children were classified as anemic, while 32 were categorized as healthy. The study revealed that FGF21 levels ≥ 0.745 pg/mL and eNOS levels ≥ 1.265 µg/mL predicted anemia. Hemoglobin levels exhibited a negative correlation with FGF21 (r = -0.381; p = 0.002) and eNOS levels (r = -0.462; p < 0.001). Furthermore, a significant negative correlation was observed between GDF-15 and ferritin (r = -0.311; p = 0.019), while eNOS levels correlated positively with folate (r = 0.313; p = 0.019). Conclusions: Anemia induced elevated mitochondrial biomarkers; FGF21 and eNOS levels. The findings suggest that the long-term ramifications of anemia in childhood may be associated with mitochondrial dysfunction.

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Protein phosphatase 1D (PPM1D, Wip1) is induced by the tumor suppressor p53 during DNA damage response signaling and acts as an oncoprotein in several human cancers. Although PPM1D is a potential therapeutic target, insights into its atomic structure were challenging due to flexible regions unique to this family member. Here, we report the first crystal structure of the PPM1D catalytic domain to 1.8 Å resolution. The structure reveals the active site with two Mg2+ ions bound, similar to other structures. The flap subdomain and B-loop, which are crucial for substrate recognition and catalysis, were also resolved, with the flap forming two short helices and three short ß-strands that are followed by an irregular loop. Unexpectedly, a nitrogen-oxygen-sulfur bridge was identified in the catalytic domain. Molecular dynamics simulations and kinetic studies provided further mechanistic insights into the regulation of PPM1D catalytic activity. In particular, the kinetic experiments demonstrated a magnesium concentration-dependent lag in PPM1D attaining steady-state velocity, a feature of hysteretic enzymes that show slow transitions compared with catalytic turnover. All combined, these results advance the understanding of PPM1D function and will support the development of PPM1D-targeted therapeutics.

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Dominio Catalítico , Proteína Fosfatasa 2C , Proteína Fosfatasa 2C/metabolismo , Proteína Fosfatasa 2C/química , Proteína Fosfatasa 2C/genética , Humanos , Cristalografía por Rayos X , Magnesio/metabolismo , Magnesio/química , Simulación de Dinámica Molecular , Cinética , Fosfoproteínas Fosfatasas/química , Fosfoproteínas Fosfatasas/metabolismo , Fosfoproteínas Fosfatasas/genética

RESUMEN

Our prior investigations have established that Inonotus obliquus (Chaga) possesses hypoglycemic effects. Persistent hyperglycemia is known to precipitate renal function abnormalities. The functionality of the kidneys is intricately linked to the levels of cyclic guanosine-3',5'-monophosphate (cGMP), which are influenced by the activities of nitric oxide synthase (NOS) and phosphodiesterase (PDE). Enhanced cGMP levels can be achieved either through the upregulation of NOS activity or the downregulation of PDE activity. The objective of the current study is to elucidate the effects of Chaga on disorders of glucolipid metabolism and renal abnormalities in rats with type 2 diabetes mellitus (T2DM), while concurrently examining the NOS-cGMP-PDE5 signaling pathway. A model of T2DM was developed in rats using a high-fat diet (HFD) combined with streptozotocin (STZ) administration, followed by treatment with Chaga extracts at doses of 50 and 100 mg·kg-1 for eight weeks. The findings revealed that Chaga not only mitigated metabolic dysfunctions, evidenced by improvements in fasting blood glucose, total cholesterol, triglycerides, and insulin resistance, but also ameliorated renal function markers, including serum creatinine, urine creatinine (UCr), blood urea nitrogen, 24-h urinary protein, and estimated creatinine clearance. Additionally, enhancements in glomerular volume, GBM thickness, podocyte foot process width (FPW), and the mRNA and protein expressions of podocyte markers, such as nephrin and wilms tumor-1, were observed. Chaga was found to elevate cGMP levels in both serum and kidney tissues by increasing mRNA and protein expressions of renal endothelial NOS and neural NOS, while simultaneously reducing the expressions of renal inducible NOS and PDE5. In summary, Chaga counteracts HFD/STZ-induced glucolipid metabolism and renal function disturbances by modulating the NOS-cGMP-PDE5 signaling pathway. This research supports the potential application of Chaga in the clinical prevention and treatment of T2DM and diabetic nephropathy (DN), with cGMP serving as a potential therapeutic target.

Asunto(s)

GMP Cíclico , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 5 , Dieta Alta en Grasa , Inonotus , Riñón , Óxido Nítrico Sintasa , Transducción de Señal , Animales , GMP Cíclico/metabolismo , Masculino , Transducción de Señal/efectos de los fármacos , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 5/metabolismo , Riñón/efectos de los fármacos , Riñón/metabolismo , Dieta Alta en Grasa/efectos adversos , Ratas , Óxido Nítrico Sintasa/metabolismo , Óxido Nítrico Sintasa/genética , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Ratas Sprague-Dawley , Estreptozocina , Humanos , Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Experimental/metabolismo , Glucemia/metabolismo , Glucemia/efectos de los fármacos , Hipoglucemiantes/farmacología

RESUMEN

In response to mechanical loading of bone, osteocytes produce nitric oxide (NO•) and decrease sclerostin protein expression, leading to an increase in bone mass. However, it is unclear whether NO• production and sclerostin protein loss are mechanistically linked, and, if so, the nature of their hierarchical relationship within an established mechano-transduction pathway. Prior work showed that following fluid-shear stress (FSS), osteocytes produce NOX2-derived reactive oxygen species, inducing calcium (Ca2+) influx. Increased intracellular Ca2+ results in calcium-calmodulin dependent protein kinase II (CaMKII) activation, which regulates the lysosomal degradation of sclerostin protein. Here, we extend our discoveries, identifying NO• as a regulator of sclerostin degradation downstream of mechano-activated CaMKII. Pharmacological inhibition of nitric oxide synthase (NOS) activity in Ocy454 osteocyte-like cells prevented FSS-induced sclerostin protein loss. Conversely, short-term treatment with a NO• donor in Ocy454 cells or isolated murine long bones was sufficient to induce the rapid decrease in sclerostin protein abundance, independent of changes in Sost gene expression. Ocy454 cells express all three NOS genes, and transfection with siRNAs targeting eNOS/Nos3 was sufficient to prevent FSS-induced loss of sclerostin protein, while siRNAs targeting iNOS/Nos2 mildly blunted the loss of sclerostin but did not reach statistical significance. Similarly, siRNAs targeting both eNOS/Nos3 and iNOS/Nos2 prevented FSS-induced NO• production. Together, these data show iNOS/Nos2 and eNOS/Nos3 are the primary producers of FSS-dependent NO•, and that NO• is necessary and sufficient for sclerostin protein control. Further, selective inhibition of elements within this sclerostin-controlling mechano-transduction pathway indicated that NO• production occurs downstream of CaMKII activation. Targeting Camk2d and Camk2g with siRNA in Ocy454 cells prevented NO• production following FSS, indicating that CaMKII is needed for NO• production. However, NO• donation (1min) resulted in a significant increase in CaMKII activation, suggesting that NO• may have the ability to tune CaMKII response. Together, these data support that CaMKII is necessary for, and may be modulated by NO•, and that the interaction of these two signals is involved in the control of sclerostin protein abundance, consistent with a role in bone anabolic responses.

Asunto(s)

Proteínas Adaptadoras Transductoras de Señales , Óxido Nítrico , Osteocitos , Óxido Nítrico/metabolismo , Animales , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Osteocitos/metabolismo , Ratones , Estrés Mecánico , Ratones Endogámicos C57BL , Mecanotransducción Celular , Línea Celular , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/metabolismo

RESUMEN

BACKGROUND: The age at onset (AO) of Machado-Joseph disease (SCA3/MJD), a disorder due to an expanded CAG repeat (CAGexp) in ATXN3, is quite variable and the role of environmental factors is still unknown. Caffeine was associated with protective effects against other neurodegenerative diseases, and against SCA3/MJD in transgenic mouse models. We aimed to evaluate whether caffeine consumption and its interaction with variants of caffeine signaling/metabolization genes impact the AO of this disease. METHODS: a questionnaire on caffeine consumption was applied to adult patients and unrelated controls living in Rio Grande do Sul, Brazil. AO and CAGexp were previously determined. SNPs rs5751876 (ADORA2A), rs2298383 (ADORA2A), rs762551 (CYP1A2) and rs478597 (NOS1) were genotyped. AO of subgroups were compared, adjusting the CAGexp to 75 repeats (p < 0.05). RESULTS: 171/179 cases and 98/100 controls consumed caffeine. Cases with high and low caffeine consumption (more or less than 314.5 mg of caffeine/day) had mean (SD) AO of 35.05 (11.44) and 35.43 (10.08) years (p = 0.40). The mean (SD) AO of the subgroups produced by the presence or absence of caffeine-enhancing alleles in ADORA2A (T allele at rs5751876 and rs2298383), CYP1A2 (C allele) and NOS1 (C allele) were all similar (p between 0.069 and 0.516). DISCUSSION: Caffeine consumption was not related to changes in the AO of SCA3/MJD, either alone or in interaction with protective genotypes at ADORA2A, CYP1A2 and NOS1.

RESUMEN

Background Despite advances in chronic myeloid leukemia (CML) genetics, the role of nitric oxide (NO) and hydrogen sulfide (H2S) gene mutations and their relationship to apoptotic genes is unclear. Therefore, this study investigated NO- and H2S-producing genes' mutations and their interactions with apoptotic genes using Sanger sequencing and next-generation sequencing (NGS). Methodology A complete blood count (CBC) was carried out to measure the total number of white blood cells, while IL-6 levels were assessed in both control and CML patients using an ELISA technique. Sanger sequencing was used to analyze mutations in the CTH and NOS3 genes, whereas NGS was applied to examine mutations on all chromosomes. Results White blood cell (WBC) and granulocyte counts were significantly higher in CML patients compared to controls (p<0.0001), and monocyte counts were similarly higher (p<0.05). Interleukin-6 (IL-6) levels were significantly elevated in CML patients than controls (p<0.0001), indicating a possible link to CML etiology or progression. Multiple mutations have been identified in both genes, notably in CTH exon 12 and the NOS3 genes VNTR, T786C, and G894T. This study also measured IL-6 concentrations using IL-6 assays, identifying its potential as a CML prognostic diagnostic. WBC counts, granulocyte counts, and mid-range absolute counts, or MID counts, were significantly higher in CML patients than in normal control individuals. NGS identified 1643 somatic and sex chromosomal abnormalities and 439 actively expressed genes in CML patients. The findings imply a genomic landscape beyond the BCR-ABL1 mutation in CML development compared to other databases. Conclusion In conclusion, this study advances the understanding of the genetic characteristics of CML by identifying mutations in the NO- and H2S-producing genes and their complex connections with genes involved in apoptosis. The comprehensive genetic profile obtained by Sanger sequencing and NGS provides possibilities for identifying novel targets for therapy and personalized treatments for CML, therefore contributing to developments in hematological diseases.

RESUMEN

BACKGROUND: Nitric oxide synthase (NOS) is an enzyme that catalyzes the formation of nitric oxide (NO), the altered production of which is characteristic of diabetic nephropathy. NOS exists in three isoforms: NOS1, NOS2, and NOS3. Moreover, there are reports about the potential role of NOS3 polymorphisms in the development of diabetes complications. The aim of this study was to assess the role of selected NOS polymorphisms-rs3782218 (NOS1), rs1137933 (NOS2), rs1799983, rs2070744, and rs61722009 (NOS3)-in the risk of developing diabetic nephropathy and in the likelihood of renal replacement therapy. METHODS: The studied polymorphisms were analyzed in a group of 232 patients divided into three groups. Four polymorphisms (rs3782218, rs1137933, rs1799983, rs2070744) were genotyped using the PCR-RFLP, while the rs61722009 polymorphism was genotyped using the PCR. RESULTS: The C/C genotype and the C allele of the rs3782218 polymorphism (NOS1) were associated with an increased risk of developing diabetic nephropathy and an increased likelihood of renal replacement therapy. In turn, the G allele of the rs1137933 polymorphism (NOS2) reduces the likelihood of renal replacement therapy. CONCLUSIONS: The specific genotypes or alleles of the rs3782218 (NOS1) and rs1137933 (NOS2) polymorphisms seem to be potential risk factors for diabetic nephropathy and renal replacement therapy.

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Odontogenic tumors (OGTs), which originate from cells of odontogenic apparatus and their remnants, are rare entities. Primary intraosseous carcinoma NOS (PIOC), is one of the OGTs, but it is even rarer and has a worse prognosis. The precise characteristics of PIOC, especially in immunohistochemical features and its pathogenesis, remain unclear. We characterized a case of PIOC arising from the left mandible, in which histopathological findings showed a transition from the odontogenic keratocyst to the carcinoma. Remarkably, the tumor lesion of this PIOC prominently exhibits malignant attributes, including invasive growth of carcinoma cell infiltration into the bone tissue, an elevated Ki-67 index, and lower signal for CK13 and higher signal for CK17 compared with the non-tumor region, histopathologically and immunohistopathologically. Further immunohistochemical analyses demonstrated increased expression of ADP-ribosylation factor (ARF)-like 4c (ARL4C) (accompanying expression of ß-catenin in the nucleus) and yes-associated protein (YAP) in the tumor lesion. On the other hand, YAP was expressed and the expression of ARL4C was hardly detected in the non-tumor region. In addition, quantitative RT-PCR analysis using RNAs and dot blot analysis using genomic DNA showed the activation of Wnt/ß-catenin signaling and epigenetic alterations, such as an increase of 5mC levels and a decrease of 5hmC levels, in the tumor lesion. A DNA microarray and a gene set enrichment analysis demonstrated that various types of intracellular signaling would be activated and several kinds of cellular functions would be altered in the pathogenesis of PIOC. Experiments with the GSK-3 inhibitor revealed that ß-catenin pathway increased not only mRNA levels of ankyrin repeat domain1 (ANKRD1) but also protein levels of YAP and transcriptional co-activator with PDZ-binding motif (TAZ) in oral squamous cell carcinoma cell lines. These results suggested that further activation of YAP signaling by Wnt/ß-catenin signaling may be associated with the pathogenesis of PIOC deriving from odontogenic keratocyst in which YAP signaling is activated.

Asunto(s)

Proteínas Adaptadoras Transductoras de Señales , Quistes Odontogénicos , Tumores Odontogénicos , Factores de Transcripción , Vía de Señalización Wnt , Humanos , Quistes Odontogénicos/patología , Quistes Odontogénicos/metabolismo , Tumores Odontogénicos/patología , Tumores Odontogénicos/metabolismo , Vía de Señalización Wnt/fisiología , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Factores de Transcripción/metabolismo , beta Catenina/metabolismo , beta Catenina/genética , Proteínas Señalizadoras YAP , Neoplasias Mandibulares/patología , Neoplasias Mandibulares/metabolismo , Masculino , Femenino

RESUMEN

AIMS: Common genetic variations in the nitric oxide synthase-1 adaptor protein (NOS1AP) gene are associated with QT-interval prolongation. In a previous study, we observed an association between the rs10494366 variant of this gene and an increased QT-interval shortening in digoxin users. As QT-interval shortening is a risk factor for sudden cardiac death (SCD), in this study, we investigated whether the association between digoxin use and risk of SCD differs in participants with different NOS1AP rs10494366 genotypes. METHODS: We included 11 377 individuals from the prospective population-based cohort of the Rotterdam Study. We used Cox proportional hazard regression analysis with digoxin as time-dependent exposure to estimate the associations between current digoxin use and the risk of SCD among different rs10494366 genotype groups in the adjusted models. We also studied whether such an association was dose-dependent, comparing high dosage (≥ 0.250 mg), moderate dosage (0.125 mg ≤ dose< 0.250 mg) and low dosage (< 0.125 mg) digoxin users with non-users. RESULTS: The median baseline age of the total study population was 62 (interquartile range [IQR] 58-71) years. The cumulative incidence of SCD was 4.1% (469 cases), and among them, 74 (15.7%) individuals were current digoxin users at the time of death, during a median follow-up of 11.5 (IQR 6.5-17) years. Current digoxin users had an increased risk of SCD (multivariable adjusted model hazard ratio [HR]: 3.07; 95% confidence interval [CI]: 2.38-3.98), with no significant differences between the three genotype groups. The adjusted HRs were 4.03 [95% CI: 1.98-8.21] in the minor homozygous GG, 3.46 [95% CI: 2.37-5.04] in the heterozygous TG and 2.56 [95%CI: 1.70-3.86] in the homozygous TT genotype groups. Compared to low- and moderate-dose, high-dose digoxin users with GG genotype had the highest risk of SCD (HR: 5.61 [95% CI: 1.34-23.47]). CONCLUSIONS: Current use of digoxin is associated with a significantly increased risk of SCD. The NOS1AP gene rs10494366 variant did not modify the digoxin-associated risk of SCD in a population of European ancestry.

Asunto(s)

Proteínas Adaptadoras Transductoras de Señales , Muerte Súbita Cardíaca , Digoxina , Genotipo , Humanos , Digoxina/efectos adversos , Digoxina/administración & dosificación , Femenino , Masculino , Persona de Mediana Edad , Muerte Súbita Cardíaca/etiología , Muerte Súbita Cardíaca/epidemiología , Anciano , Estudios Prospectivos , Proteínas Adaptadoras Transductoras de Señales/genética , Factores de Riesgo , Relación Dosis-Respuesta a Droga , Polimorfismo de Nucleótido Simple , Países Bajos/epidemiología , Cardiotónicos/efectos adversos , Cardiotónicos/administración & dosificación , Cardiotónicos/uso terapéutico , Síndrome de QT Prolongado/genética , Síndrome de QT Prolongado/inducido químicamente

RESUMEN

Idiopathic multicentric Castleman disease (iMCD) and TAFRO syndrome present a variety of symptoms thought to be caused by excessive inflammatory cytokines and chemokines, but the underlying mechanisms are unknown. iMCD is broadly classified into two types: iMCD-NOS and iMCD-TAFRO, which have distinct laboratory findings, pathological features, and responses to treatments. It is thought that iMCD-NOS, particularly the IPL type, responds favorably to IL-6 inhibitors due to its IL-6-centric profile. iMCD-TAFRO frequently progresses acutely and seriously, similar to TAFRO syndrome. Elevated levels of cytokines, including IL-1ß, TNF-α, IL-10, and IL-23, as well as chemokines like CXCL13 and CXCL-10 (especially in iMCD-TAFRO), SAA, and VEGF, have been linked to the disease's pathology. Recent research has identified key signaling pathways including PI3K/Akt/mTOR and JAK-STAT3, as well as those regulated by type I IFN, as crucial in iMCD-TAFRO. These results suggest that dominant pathways may vary between subtypes. Further research into the peripheral blood and lymph nodes is required to determine the disease spectrum of iMCD-NOS/iMCD-TAFRO/TAFRO syndrome.

RESUMEN

Glioblastoma (GBM) is the most common malignant primary brain tumor, resulting in poor survival despite aggressive therapies. GBM is characterized by a highly heterogeneous and immunosuppressive tumor microenvironment (TME) made up predominantly of infiltrating peripheral immune cells. One significant immune cell type that contributes to glioma immune evasion is a population of immunosuppressive cells, termed myeloid-derived suppressor cells (MDSCs). Previous studies suggest that a subset of myeloid cells, expressing monocytic (M)-MDSC markers and dual expression of chemokine receptors CCR2 and CX3CR1, utilize CCR2 to infiltrate the TME. This study evaluated the mechanism of CCR2+/CX3CR1+ M-MDSC differentiation and T cell suppressive function in murine glioma models. We determined that bone marrow-derived CCR2+/CX3CR1+ cells adopt an immune suppressive cell phenotype when cultured with glioma-derived factors. Glioma secreted CSF1R ligands M-CSF and IL-34 were identified as key drivers of M-MDSC differentiation while adenosine and iNOS pathways were implicated in M-MDSC suppression of T cells. Mining a human GBM spatial RNAseq database revealed a variety of different pathways that M-MDSCs utilize to exert their suppressive function that are driven by complex niches within the microenvironment. These data provide a more comprehensive understanding of the mechanism of M-MDSCs in glioblastoma.

RESUMEN

Sulfated marine polysaccharides, so-called fucoidans, have been shown to exhibit anti-inflammatory and immunomodulatory activities in retinal pigment epithelium (RPE). In this study, we tested the effects of different fucoidans (and of fucoidan-treated RPE cells) on retinal microglia to investigate whether its anti-inflammatory effect can be extrapolated to the innate immune cells of the retina. In addition, we tested whether fucoidan treatment influenced the anti-inflammatory effect of RPE cells on retinal microglia. Three fucoidans were tested (FVs from Fucus vesiculosus, Fuc1 and FucBB04 from Laminaria hyperborea) as well as the supernatant of primary porcine RPE treated with fucoidans for their effects on inflammatory activated (using lipopolysaccharide, LPS) microglia cell line SIM-A9 and primary porcine retinal microglia. Cell viability was detected with a tetrazolium assay (MTT), and morphology by Coomassie staining. Secretion of tumor necrosis factor alpha (TNFα), interleukin 1 beta (IL1ß) and interleukin 8 (IL8) was detected with ELISA, gene expression (NOS2 (Nitric oxide synthase 2), and CXCL8 (IL8)) with qPCR. Phagocytosis was detected with a fluorescence assay. FucBB04 and FVs slightly reduced the viability of SIM-A9 and primary microglia, respectively. Treatment with RPE supernatants increased the viability of LPS-treated primary microglia. FVs and FucBB04 reduced the size of LPS-activated primary microglia, indicating an anti-inflammatory phenotype. RPE supernatant reduced the size of LPS-activated SIM-A9 cells. Proinflammatory cytokine secretion and gene expression in SIM-A9, as well as primary microglia, were not significantly affected by fucoidans, but RPE supernatants reduced the secretion of LPS-induced proinflammatory cytokine secretion in SIM-A9 and primary microglia. The phagocytosis ability of primary microglia was reduced by FucBB04. In conclusion, fucoidans exhibited only modest effects on inflammatorily activated microglia by maintaining their cell size under stimulation, while the anti-inflammatory effect of RPE cells on microglia irrespective of fucoidan treatment could be confirmed, stressing the role of RPE in regulating innate immunity in the retina.

Asunto(s)

Supervivencia Celular , Microglía , Polisacáridos , Epitelio Pigmentado de la Retina , Microglía/efectos de los fármacos , Microglía/metabolismo , Animales , Polisacáridos/farmacología , Porcinos , Epitelio Pigmentado de la Retina/efectos de los fármacos , Epitelio Pigmentado de la Retina/metabolismo , Epitelio Pigmentado de la Retina/citología , Supervivencia Celular/efectos de los fármacos , Fagocitosis/efectos de los fármacos , Retina/efectos de los fármacos , Retina/metabolismo , Retina/citología , Línea Celular , Lipopolisacáridos/farmacología , Antiinflamatorios/farmacología , Citocinas/metabolismo