RESUMEN
Mesenchymal stem cells (MSCs) are multipotent cells that can differentiate into mature cells of various cell types. Although the differentiation process of MSCs requires lineage-specific transcription factors, the exact molecular mechanism that determines MSCs differentiation is not clearly addressed. Here, we demonstrate a Smad4-Taz axis as a new intrinsic regulator for adipo-osteogenic differentiation of MSCs and show that this function of Smad4 is independent of the transforming growth factor-ß signal. Smad4 directly bound to the Taz protein and facilitated nuclear localization of Taz through its nuclear localization signal. Nuclear retention of Taz by direct binding to Smad4 increased expression of osteogenic genes through enhancing Taz-runt-related transcription factor 2 (Runx2) interactions in the C3H10T1/2 MSC cell line and preosteoblastic MC3T3-E1 cells, whereas it suppressed expression of adipogenic genes through promoting Taz-peroxisome proliferator-activated receptor-γ (PPARγ) interaction in C3H10T1/2 and preadipogenic 3T3-L1 cells. A reciprocal role of the Smad4 in osteogenic and adipogenic differentiation was also observed in human adipose tissue-derived stem cells (hASCs). Consequently, Smad4 depletion in C3H10T1/2 and hASCs reduced nuclear retention of Taz and thus caused the decreased interaction with Runx2 or PPARγ, resulting in delayed osteogenesis or enhanced adipogenesis of the MSC. Therefore, these findings provide insight into a novel function of Smad4 to regulate the balance of MSC lineage commitment through reciprocal targeting of the Taz protein in osteogenic and adipogenic differentiation pathways. Stem Cells 2019;37:368-381.
Asunto(s)
Adipogénesis , Células Madre Mesenquimatosas/metabolismo , Osteogénesis , Transducción de Señal , Proteína Smad4/metabolismo , Transactivadores/metabolismo , Proteínas Adaptadoras Transductoras de Señales , Animales , Diferenciación Celular , Línea Celular , Humanos , Células Madre Mesenquimatosas/citología , Ratones , Proteína Smad4/genética , Transactivadores/genética , Proteínas Coactivadoras Transcripcionales con Motivo de Unión a PDZRESUMEN
Adipocyte differentiation is controlled by multiple signaling pathways. To identify new adipogenic factors, C3H10T1/2 adipocytes were treated with previously known antiadipogenic phytochemicals (resveratrol, butein, sulfuretin, and fisetin) for 24 hours. Commonly regulated genes were then identified by transcriptional profiling analysis. Three genes (chemokine (C-X-C motif) ligand 1 [ Cxcl1], heme oxygenase 1 [ Hmox1], and PHD (plant homeo domain) finger protein 16 [ Phf16]) were upregulated while two genes (G0/G1 switch gene 2 [ G0s2] and patatin-like phospholipase domain containing 3 [ Pnpla3]) were downregulated by these four antiadipogenic compounds. Tissue expression profiles showed that the G0s2 and Pnpla3 expressions were highly specific to adipose depots while the other three induced genes were ubiquitously expressed with significantly higher expression in adipose tissues. While Cxcl1 expression was decreased, expressions of the other four genes were significantly increased during adipogenic differentiation of C3H10T1/2 cells. Small interfering RNA-mediated knockdown including Phf16 and Pnpla3 indicated that these genes might play regulatory roles in lipid accumulation and adipocyte differentiation. Specifically, the silencing of two newly identified adipogenic genes, Phf16 or Pnpla3, suppressed lipid accumulation and expression of adipocyte markers in both 3T3-L1 and C3H10T1/2 cells. Taken together, these data showed previously uncovered roles of Phf16 and Pnpla3 in adipogenesis, highlighting the potential of using phytochemicals for further investigation of adipocyte biology.
Asunto(s)
Adipogénesis/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Proteínas Oncogénicas/metabolismo , Fosfolipasas A2 Calcio-Independiente/metabolismo , Fitoquímicos/farmacología , Células 3T3-L1 , Animales , Quimiocina CXCL1/biosíntesis , Ratones , Proteínas Oncogénicas/genética , Fosfolipasas A2 Calcio-Independiente/genéticaRESUMEN
Activating transcription factor 3 (Atf3) has been previously demonstrated to impact obesity and metabolism. However, a metabolic role of Atf3 in mice remains debatable. We investigated the role of Atf3 in mice and further investigated Atf3 expression as a therapeutic target for obesity and metabolic diseases. Atf3 knockout (KO) mice fed with a high fat diet (HFD) aggravated weight gain and impaired glucose metabolism compared to littermate control wild type (WT) mice. Atf3 KO aged mice fed with a chow diet (CD) for longer than 10 months also displayed increased body weight and fat mass compared to WT aged mice. We also assessed requirements of Atf3 in a phytochemical mediated anti-obese effect. Effect of sulfuretin, a previously known phytochemical Atf3 inducer, in counteracting weight gain and improving glucose tolerance was almost completely abolished in the absence of Atf3, indicating that Atf3 induction can be a molecular target for preventing obesity and metabolic diseases. We further identified other Atf3 small molecule inducers that exhibit inhibitory effects on lipid accumulation in adipocytes. These data highlight the role of Atf3 in obesity and further suggest the use of chemical Atf3 inducers for prevention of obesity and metabolic diseases.
Asunto(s)
Factor de Transcripción Activador 3/metabolismo , Fármacos Antiobesidad/farmacología , Benzofuranos/farmacología , Enfermedades Metabólicas/metabolismo , Obesidad/metabolismo , Factor de Transcripción Activador 3/genética , Envejecimiento/genética , Animales , Peso Corporal/genética , Dieta Alta en Grasa/efectos adversos , Flavonoides/farmacología , Intolerancia a la Glucosa/genética , Enfermedades Metabólicas/genética , Ratones Noqueados , Terapia Molecular Dirigida/métodos , Obesidad/tratamiento farmacológico , Obesidad/etiología , Obesidad/genéticaRESUMEN
Increasing the thermogenic activity of adipocytes holds promise as an approach to combating human obesity and related metabolic diseases. We identified induction of mouse PR domain containing 4 (Prdm4) by the small molecule butein as a means to induce expression of uncoupling protein 1 (Ucp1), increase energy expenditure, and stimulate the generation of thermogenic adipocytes. This study highlights a Prdm4-dependent pathway, modulated by small molecules, that stimulates browning of white adipose tissue.
Asunto(s)
Tejido Adiposo Pardo/efectos de los fármacos , Tejido Adiposo Blanco/efectos de los fármacos , Chalconas/farmacología , Proteínas de Unión al ADN/antagonistas & inhibidores , Factores de Transcripción/antagonistas & inhibidores , Tejido Adiposo Pardo/metabolismo , Tejido Adiposo Blanco/metabolismo , Animales , Chalconas/química , Proteínas de Unión al ADN/metabolismo , Dieta Alta en Grasa/efectos adversos , Ratones , Ratones Endogámicos C57BL , Ratones Obesos , Factores de Transcripción/metabolismoRESUMEN
The phytochemical oxyresveratrol has been shown to exert diverse biological activities including prevention of obesity. However, the exact reason underlying the anti-obese effects of oxyresveratrol is not fully understood. Here, we investigated the effects and mechanism of oxyresveratrol in adipocytes and high-fat diet (HFD)-fed obese mice. Oxyresveratrol suppressed lipid accumulation and expression of adipocyte markers during the adipocyte differentiation of 3T3-L1 and C3H10T1/2 cells. Administration of oxyresveratrol in HFD-fed obese mice prevented body-weight gains, lowered adipose tissue weights, improved lipid profiles, and increased glucose tolerance. The anti-obese effects were linked to increases in energy expenditure and higher rectal temperatures without affecting food intake, fecal lipid content, and physical activity. The increased energy expenditure by oxyresveratrol was concordant with the induction of thermogenic genes including Ucp1, and the reduction of white adipocyte selective genes in adipose tissue. Furthermore, Foxo3a was identified as an oxyresveratrol-induced gene and it mimicked the effects of oxyresveratrol for induction of thermogenic genes and suppression of white adipocyte selective genes, suggesting the role of Foxo3a in oxyresveratrol-mediated anti-obese effects. Taken together, these data show that oxyresveratrol increases energy expenditure through the induction of thermogenic genes in adipose tissue and further implicates oxyresveratrol as an ingredient and Foxo3a as a molecular target for the development of functional foods in obesity and metabolic diseases.
Asunto(s)
Dieta Alta en Grasa/efectos adversos , Metabolismo Energético/efectos de los fármacos , Proteína Forkhead Box O3/metabolismo , Obesidad/etiología , Obesidad/metabolismo , Extractos Vegetales/farmacología , Estilbenos/farmacología , Proteína Desacopladora 1/genética , Células 3T3-L1 , Adipocitos/efectos de los fármacos , Adipocitos/metabolismo , Tejido Adiposo/efectos de los fármacos , Tejido Adiposo/metabolismo , Animales , Diferenciación Celular/efectos de los fármacos , Línea Celular , Supervivencia Celular/efectos de los fármacos , Regulación de la Expresión Génica , Metabolismo de los Lípidos/efectos de los fármacos , Masculino , Metabolómica/métodos , Ratones , Termogénesis/genética , Proteína Desacopladora 1/metabolismoRESUMEN
MicroRNAs (miRNAs) play essential roles in various cellular processes including proliferation and differentiation. In this study, we identified miRNA-195a (miR-195a) as a regulator of adipocyte differentiation. Differential expression of miR-195a in preadipocytes and adipocytes suggests its role in lipid accumulation and adipocyte differentiation. Forced expression of miR-195a mimics suppressed lipid accumulation and inhibited expression of adipocyte markers such as PPARγ and aP2 in 3T3-L1 and C3H10T1/2 cells. Conversely, downregulation of miR-195a by anti-miR-195a increased lipid accumulation and expression of adipocyte markers. Target prediction analysis suggested zinc finger protein 423 (Zfp423), a preadipogenic determinator, as a potential gene recognized by miR-195a. In line with this, mimicked expression of miR-195a reduced the expression of Zfp423, whereas anti-miR-195a increased its expression. Predicted targeting sequences in Zfp423 3'UTR, but not mutated sequences fused to luciferase, were regulated by miR-195a. Ectopic Zfp423 expression in 3T3-L1 cells increased lipid accumulation and expression of adipocyte markers, consistent with the observation that miR-195a targets Zfp423, resulting in suppressed adipocyte differentiation. In addition, miR-195a and Zfp423 were inversely correlated in obese fat tissues, raising the possibility of miRNA's role in obesity. Together, our data show that miR-195a is an anti-adipogenic regulator, which acts by targeting Zfp423, and further suggest the roles of miR-195a in obesity and metabolic diseases.
Asunto(s)
Adipocitos/citología , Proteínas de Unión al ADN/genética , MicroARNs/genética , MicroARNs/metabolismo , Obesidad/metabolismo , Factores de Transcripción/genética , Regiones no Traducidas 3' , Células 3T3-L1 , Adipocitos/metabolismo , Animales , Diferenciación Celular , Proteínas de Unión al ADN/metabolismo , Dieta Alta en Grasa/efectos adversos , Regulación de la Expresión Génica , Metabolismo de los Lípidos , Ratones , Obesidad/etiología , Obesidad/genética , Regiones Promotoras Genéticas , Factores de Transcripción/metabolismoRESUMEN
The identification and examination of potential determinants controlling the progression of cell fate toward osteoblasts can be intriguing subjects. In this study, the effects of sulfuretin, a major compound isolated from Rhus verniciflua Stokes, on osteoblast differentiation were investigated. Treatments of sulfuretin induced alkaline phosphatase (ALP) activity in mesenchymal C3H10T1/2 cells and mineralization in preosteoblast MC3T3-E1 cells. Pro-osteogenic effects of sulfuretin were consistently observed in freshly isolated primary bone marrow cells. In mechanical studies, sulfuretin specifically induced expression of TGF-ß target genes, such as SMAD7 and PAI-1, but not other signaling pathway-related genes. Similar to the results of gene expression analysis, reporter assays further demonstrated TGF-ß-specific induction by sulfuretin. Furthermore, disruption of TGF-ß signaling using treatment with TGF-ß-specific inhibitor, SB-431542, and introduction of SMAD2/3 small interfering RNA impaired the effects of sulfuretin in inducing ALP activity and expression of ALP mRNA. Together, these data indicate that the pro-osteogenic effects of sulfuretin are mediated through activation of TGF-ß signaling, further supporting the potential of sulfuretin in the prevention of bone-related diseases such as bone fracture and osteoporosis.
Asunto(s)
Benzofuranos/farmacología , Conservadores de la Densidad Ósea/farmacología , Diferenciación Celular/efectos de los fármacos , Osteoblastos/efectos de los fármacos , Osteogénesis/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Factor de Crecimiento Transformador beta/metabolismo , Células 3T3 , Fosfatasa Alcalina/genética , Fosfatasa Alcalina/metabolismo , Animales , Benzamidas/farmacología , Dioxoles/farmacología , Relación Dosis-Respuesta a Droga , Fémur/efectos de los fármacos , Fémur/metabolismo , Flavonoides/farmacología , Masculino , Ratones , Ratones Endogámicos C57BL , Osteoblastos/metabolismo , Inhibidor 1 de Activador Plasminogénico/genética , Inhibidor 1 de Activador Plasminogénico/metabolismo , Interferencia de ARN , Proteína Smad2/genética , Proteína Smad2/metabolismo , Proteína smad3/genética , Proteína smad3/metabolismo , Proteína smad7/genética , Proteína smad7/metabolismo , Factores de Tiempo , Transfección , Factor de Crecimiento Transformador beta/antagonistas & inhibidoresRESUMEN
Advances in proteomics and mass spectrometry enable the study of limited cell populations, where high-mass accuracy instruments are typically required. While triple quadrupoles offer fast and sensitive low-mass accuracy measurements, these instruments are effectively restricted to targeted proteomics. Linear ion traps (LITs) offer a versatile, cost-effective alternative capable of both targeted and global proteomics. Here, we describe a workflow using a new hybrid quadrupole-LIT instrument that rapidly develops targeted proteomics assays from global data-independent acquisition (DIA) measurements without needing high-mass accuracy. Using an automated software approach for scheduling parallel reaction monitoring assays (PRM), we show consistent quantification across three orders of magnitude in a matched-matrix background. We demonstrate measuring low-level proteins such as transcription factors and cytokines with quantitative linearity below two orders of magnitude in a 1 ng background proteome without requiring stable isotope-labeled standards. From a 1 ng sample, we found clear consistency between proteins in subsets of CD4+ and CD8+ T cells measured using high dimensional flow cytometry and LIT-based proteomics. Based on these results, we believe hybrid quadrupole-LIT instruments represent an economical solution to democratizing mass spectrometry in a wide variety of laboratory settings.
RESUMEN
Intratumoral Tregs are key mediators of cancer immunotherapy resistance, including anti-programmed cell death (ligand) 1 [anti-PD-(L)1] immune checkpoint blockade (ICB). The mechanisms driving Treg infiltration into the tumor microenvironment (TME) and the consequence on CD8+ T cell exhaustion remain elusive. Here, we report that heat shock protein gp96 (also known as GRP94) was indispensable for Treg tumor infiltration, primarily through the roles of gp96 in chaperoning integrins. Among various gp96-dependent integrins, we found that only LFA-1 (αL integrin), and not αV, CD103 (αE), or ß7 integrin, was required for Treg tumor homing. Loss of Treg infiltration into the TME by genetic deletion of gp96/LFA-1 potently induced rejection of tumors in multiple ICB-resistant murine cancer models in a CD8+ T cell-dependent manner, without loss of self-tolerance. Moreover, gp96 deletion impeded Treg activation primarily by suppressing IL-2/STAT5 signaling, which also contributed to tumor regression. By competing for intratumoral IL-2, Tregs prevented the activation of CD8+ tumor-infiltrating lymphocytes, drove thymocyte selection-associated high mobility group box protein (TOX) induction, and induced bona fide CD8+ T cell exhaustion. By contrast, Treg ablation led to striking CD8+ T cell activation without TOX induction, demonstrating clear uncoupling of the 2 processes. Our study reveals that the gp96/LFA-1 axis plays a fundamental role in Treg biology and suggests that Treg-specific gp96/LFA-1 targeting represents a valuable strategy for cancer immunotherapy without inflicting autoinflammatory conditions.
Asunto(s)
Linfocitos T CD8-positivos , Inmunoterapia , Linfocitos T Reguladores , Microambiente Tumoral , Animales , Linfocitos T Reguladores/inmunología , Ratones , Linfocitos T CD8-positivos/inmunología , Microambiente Tumoral/inmunología , Glicoproteínas de Membrana/inmunología , Glicoproteínas de Membrana/genética , Ratones Noqueados , Antígeno-1 Asociado a Función de Linfocito/inmunología , Antígeno-1 Asociado a Función de Linfocito/genética , Antígeno-1 Asociado a Función de Linfocito/metabolismo , Humanos , Agotamiento de Células TRESUMEN
Obesity is a global public health problem and is related with fatal diseases such as cancer and cardiovascular and metabolic diseases. Medical and lifestyle-related strategies to combat obesity have their limitations. White adipose tissue (WAT) browning is a promising strategy for increasing energy expenditure in individuals with obesity. Uncoupling protein 1 (UCP1) drives WAT browning. We previously screened natural products that enable induction of Ucp1 and demonstrated that these natural products induced WAT browning and increased energy expenditure in mice with diet-induced obesity. In this study, we aimed to extensively optimise the structure of compound 1, previously shown to promote WAT browning. Compound 3 s exhibited a significantly higher ability to induce Ucp1 in white and brown adipocytes than did compound 1. A daily injection of compound 3 s at 5 mg/kg prevented weight gain by 13.6 % in high-fat diet-fed mice without any toxicological observation. In addition, compound 3 s significantly improved glucose homeostasis, decreased serum triacylglycerol levels, and reduced total cholesterol and LDL cholesterol levels, without altering dietary intake or physical activity. Pharmaceutical properties such as solubility, lipophilicity, and membrane permeability as well as metabolic stability, half-life (T1/2), and blood exposure ratio of i.p to i.v were significantly improved in compound 3 s when compared with those in compound 1. Regarding the mode of action of WAT browning, the induction of Ucp1 and Prdm4 by compounds 1 and 3 s was dependent on Akt1 in mouse embryonic fibroblasts. Therefore, this study suggests the potential of compound 3 s as a therapeutic agent for individuals with obesity and related metabolic diseases, which acts through the induction of WAT browning as well as brown adipose tissue activation.
Asunto(s)
Dieta Alta en Grasa , Metabolismo Energético , Resistencia a la Insulina , Ratones Endogámicos C57BL , Obesidad , Proteína Desacopladora 1 , Animales , Dieta Alta en Grasa/efectos adversos , Obesidad/tratamiento farmacológico , Obesidad/metabolismo , Metabolismo Energético/efectos de los fármacos , Masculino , Ratones , Proteína Desacopladora 1/metabolismo , Tejido Adiposo Blanco/efectos de los fármacos , Tejido Adiposo Blanco/metabolismo , Chalconas/farmacología , Ratones Obesos , Fármacos Antiobesidad/farmacología , Células 3T3-L1RESUMEN
BACKGROUND: The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant is highly transmissible and evades pre-established immunity. Messenger RNA (mRNA) vaccination against ancestral strain spike protein can induce intact T-cell immunity against the Omicron variant, but efficacy of booster vaccination in patients with late-stage lung cancer on immune-modulating agents including anti-programmed cell death protein 1(PD-1)/programmed death-ligand 1 (PD-L1) has not yet been elucidated. METHODS: We assessed T-cell responses using a modified activation-induced marker assay, coupled with high-dimension flow cytometry analyses. Peripheral blood mononuclear cells (PBMCs) were stimulated with various viral peptides and antigen-specific T-cell responses were evaluated using flow cytometry. RESULTS: Booster vaccines induced CD8+ T-cell response against the ancestral SARS-CoV-2 strain and Omicron variant in both non-cancer subjects and patients with lung cancer, but only a marginal induction was detected for CD4+ T cells. Importantly, antigen-specific T cells from patients with lung cancer showed distinct subpopulation dynamics with varying degrees of differentiation compared with non-cancer subjects, with evidence of dysfunction. Notably, female-biased T-cell responses were observed. CONCLUSION: We conclude that patients with lung cancer on immunotherapy show a substantial qualitative deviation from non-cancer subjects in their T-cell response to mRNA vaccines, highlighting the need for heightened protective measures for patients with cancer to minimize the risk of breakthrough infection with the Omicron and other future variants.
Asunto(s)
COVID-19 , Neoplasias Pulmonares , Humanos , Femenino , Vacunas de ARNm , Vacunas contra la COVID-19/uso terapéutico , SARS-CoV-2 , Leucocitos Mononucleares , COVID-19/prevención & controlRESUMEN
ABSTRACT: Chimeric antigen receptor (CAR) T-cell therapy has revolutionized treatment for relapsed/refractory B-cell non-Hodgkin lymphoma (NHL). Robust biomarkers and a complete understanding of CAR T-cell function in the postinfusion phase remain limited. Here, we used a 37-color spectral flow cytometry panel to perform high dimensional single-cell analysis of postinfusion samples in 26 patients treated with CD28 costimulatory domain containing commercial CAR T cells for NHL and focused on computationally gated CD8+ CAR T cells. We found that the presence of postinfusion Programmed cell death protein 1 (PD-1)+ CD8+ CAR T cells at the day 14 time point highly correlated with the ability to achieve complete response (CR) by 6 months. Further analysis identified multiple subtypes of CD8+ PD-1+ CAR T cells, including PD-1+ T cell factor 1 (TCF1)+ stem-like CAR T cells and PD-1+ T-cell immunoglobulin and mucin-domain containing-3 (TIM3)+ effector-like CAR T cells that correlated with improved clinical outcomes such as response and progression-free survival. Additionally, we identified a subset of PD-1+ CD8+ CAR+ T cells with effector-like function that was increased in patients who achieved a CR and was associated with grade 3 or higher immune effector cell-associated neurotoxicity syndrome. Here, we identified robust biomarkers of response to CD28 CAR T cells and highlight the importance of PD-1 positivity in CD8+ CAR T cells after infusion in achieving CR.
Asunto(s)
Linfocitos T CD8-positivos , Inmunoterapia Adoptiva , Linfoma no Hodgkin , Receptor de Muerte Celular Programada 1 , Humanos , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Linfoma no Hodgkin/terapia , Linfoma no Hodgkin/inmunología , Receptor de Muerte Celular Programada 1/metabolismo , Inmunoterapia Adoptiva/métodos , Masculino , Femenino , Persona de Mediana Edad , Anciano , Adulto , Antígenos CD19/inmunología , Receptores Quiméricos de Antígenos/metabolismo , Receptores Quiméricos de Antígenos/inmunología , Resultado del TratamientoRESUMEN
Rhus verniciflua Stokes (RVS) has been used as a traditional herbal medicine for its various biological activities including anti-adipogenic effects. Activity-guided separation led to the identification of the anti-adipogenic functions of butein. Butein, a novel anti-adipogenic compound, robustly suppressed lipid accumulation and inhibited expression of adipogenic markers. Molecular studies showed that activated transforming growth factor-ß (TGF-ß) and suppressed signal transducer and activator of transcription 3 (STAT3) signaling pathways were mediated by butein. Analysis of the temporal expression profiles suggests that TGF-ß signaling precedes the STAT3 in the butein-mediated anti-adipogenic cascade. Small interfering RNA-mediated silencing of STAT3 or SMAD2/3 blunted the inhibitory effects of butein on adipogenesis indicating that an interaction between two signaling pathways is required for the action of butein. Upon butein treatments, stimulation of TGF-ß signaling was still preserved in STAT3 silenced cells, whereas regulation of STAT3 signaling by butein was significantly impaired in SMAD2/3 silenced cells, further showing that TGF-ß acts upstream of STAT3 in the butein-mediated anti-adipogenesis. Taken together, the present study shows that butein, a novel anti-adipogenic compound from RVS, inhibits adipocyte differentiation through the TGF-ß pathway followed by STAT3 and peroxisome proliferator-activated receptor γ signaling, further implicating potential roles of butein in TGF-ß- and STAT3-dysregulated diseases.
Asunto(s)
Tejido Adiposo , Chalconas/administración & dosificación , Obesidad/metabolismo , Células 3T3-L1 , Tejido Adiposo/efectos de los fármacos , Tejido Adiposo/crecimiento & desarrollo , Animales , Diferenciación Celular/efectos de los fármacos , Línea Celular , Chalconas/química , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Ratones , Ratones Noqueados , Obesidad/patología , Fosforilación/efectos de los fármacos , Rhus/química , Factor de Transcripción STAT3/genética , Factor de Transcripción STAT3/metabolismo , Transducción de Señal/efectos de los fármacos , Proteína Smad2/genética , Proteína Smad2/metabolismo , Proteína smad3/genética , Proteína smad3/metabolismo , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
We assessed vaccine-induced antibody responses to the SARS-CoV-2 ancestral virus and Omicron variant before and after booster immunization in 57 patients with B cell malignancies. Over one-third of vaccinated patients at the pre-booster time point were seronegative, and these patients were predominantly on active cancer therapies such as anti-CD20 monoclonal antibody. While booster immunization was able to induce detectable antibodies in a small fraction of seronegative patients, the overall booster benefit was disproportionately evident in patients already seropositive and not receiving active therapy. While ancestral virus- and Omicron variant-reactive antibody levels among individual patients were largely concordant, neutralizing antibodies against Omicron tended to be reduced. Interestingly, in all patients, including those unable to generate detectable antibodies against SARS-CoV-2 spike, we observed comparable levels of EBV- and influenza-reactive antibodies, demonstrating that B cell-targeting therapies primarily impair de novo but not preexisting antibody levels. These findings support rationale for vaccination before cancer treatment.
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COVID-19 , Neoplasias , Humanos , Vacunas contra la COVID-19 , Formación de Anticuerpos , SARS-CoV-2 , Neoplasias/terapia , Anticuerpos Monoclonales , Anticuerpos AntiviralesRESUMEN
Although there are a number of discoveries from genome-wide association studies (GWAS) for obesity, it has not been successful in linking GWAS results to biology. We sought to discover causal genes for obesity by conducting functional studies on genes detected from genetic association analysis. Gene-based association analysis of 917 individual exome sequences showed that HOGA1 attains exome-wide significance (p-value < 2.7 × 10-6) for body mass index (BMI). The mRNA expression of HOGA1 is significantly increased in human adipose tissues from obese individuals in the Genotype-Tissue Expression (GTEx) dataset, which supports the genetic association of HOGA1 with BMI. Functional analyses employing cell- and animal model-based approaches were performed to gain insights into the functional relevance of Hoga1 in obesity. Adipogenesis was retarded when Hoga1 was knocked down by siRNA treatment in a mouse 3T3-L1 cell line and a similar inhibitory effect was confirmed in mice with down-regulated Hoga1. Hoga1 antisense oligonucleotide (ASO) treatment reduced body weight, blood lipid level, blood glucose, and adipocyte size in high-fat diet-induced mice. In addition, several lipogenic genes including Srebf1, Scd1, Lp1, and Acaca were down-regulated, while lipolytic genes Cpt1l, Ppara, and Ucp1 were up-regulated. Taken together, HOGA1 is a potential causal gene for obesity as it plays a role in excess body fat development.
RESUMEN
The leptin receptor (LepR) acts as a signaling nexus for the regulation of glucose uptake and obesity, among other metabolic responses. The functional role of LepR under leptin-deficient conditions remains unclear. This study reports that epiregulin (EREG) governed glucose uptake in vitro and in vivo in Lepob mice by activating LepR under leptin-deficient conditions. Single and long-term treatment with EREG effectively rescued glucose intolerance in comparative insulin and EREG tolerance tests in Lepob mice. The immunoprecipitation study revealed binding between EREG and LepR in adipose tissue of Lepob mice. EREG/LepR regulated glucose uptake without changes in obesity in Lepob mice via mechanisms, including ERK activation and translocation of GLUT4 to the cell surface. EREG-dependent glucose uptake was abolished in Leprdb mice which supports a key role of LepR in this process. In contrast, inhibition of the canonical epidermal growth factor receptor (EGFR) pathway implicated in other EREG responses, increased glucose uptake. Our data provide a basis for understanding glycemic responses of EREG that are dependent on LepR unlike functions mediated by EGFR, including leptin secretion, thermogenesis, pain, growth, and other responses. The computational analysis identified a conserved amino acid sequence, supporting an evolutionary role of EREG as an alternative LepR ligand.
Asunto(s)
Intolerancia a la Glucosa , Receptores de Leptina , Animales , Glucemia/metabolismo , Epirregulina , Receptores ErbB , Leptina/metabolismo , Ligandos , Ratones , Obesidad/metabolismo , Receptores de Leptina/genética , Receptores de Leptina/metabolismoRESUMEN
Sex bias exists in the development and progression of nonreproductive organ cancers, but the underlying mechanisms are enigmatic. Studies so far have focused largely on sexual dimorphisms in cancer biology and socioeconomic factors. Here, we establish a role for CD8+ T cell-dependent antitumor immunity in mediating sex differences in tumor aggressiveness, which is driven by the gonadal androgen but not sex chromosomes. A male bias exists in the frequency of intratumoral antigen-experienced Tcf7/TCF1+ progenitor exhausted CD8+ T cells that are devoid of effector activity as a consequence of intrinsic androgen receptor (AR) function. Mechanistically, we identify a novel sex-specific regulon in progenitor exhausted CD8+ T cells and a pertinent contribution from AR as a direct transcriptional transactivator of Tcf7/TCF1. The T cell-intrinsic function of AR in promoting CD8+ T cell exhaustion in vivo was established using multiple approaches including loss-of-function studies with CD8-specific Ar knockout mice. Moreover, ablation of the androgen-AR axis rewires the tumor microenvironment to favor effector T cell differentiation and potentiates the efficacy of anti-PD-1 immune checkpoint blockade. Collectively, our findings highlight androgen-mediated promotion of CD8+ T cell dysfunction in cancer and imply broader opportunities for therapeutic development from understanding sex disparities in health and disease.
Asunto(s)
Linfocitos T CD8-positivos , Neoplasias , Andrógenos , Animales , Diferenciación Celular , Femenino , Masculino , Ratones , Sexismo , Microambiente TumoralRESUMEN
BACKGROUND: Immune checkpoint blockade (ICB) has revolutionized cancer immunotherapy. However, most patients with cancer fail to respond clinically. One potential reason is the accumulation of immunosuppressive transforming growth factor ß (TGFß) in the tumor microenvironment (TME). TGFß drives cancer immune evasion in part by inducing regulatory T cells (Tregs) and limiting CD8+ T cell function. Glycoprotein-A repetitions predominant (GARP) is a cell surface docking receptor for activating latent TGFß1, TGFß2 and TGFß3, with its expression restricted predominantly to effector Tregs, cancer cells, and platelets. METHODS: We investigated the role of GARP in human patients with cancer by analyzing existing large databases. In addition, we generated and humanized an anti-GARP monoclonal antibody and evaluated its antitumor efficacy and underlying mechanisms of action in murine models of cancer. RESULTS: We demonstrate that GARP overexpression in human cancers correlates with a tolerogenic TME and poor clinical response to ICB, suggesting GARP blockade may improve cancer immunotherapy. We report on a unique anti-human GARP antibody (named PIIO-1) that specifically binds the ligand-interacting domain of all latent TGFß isoforms. PIIO-1 lacks recognition of GARP-TGFß complex on platelets. Using human LRRC32 (encoding GARP) knock-in mice, we find that PIIO-1 does not cause thrombocytopenia; is preferentially distributed in the TME; and exhibits therapeutic efficacy against GARP+ and GARP- cancers, alone or in combination with anti-PD-1 antibody. Mechanistically, PIIO-1 treatment reduces canonical TGFß signaling in tumor-infiltrating immune cells, prevents T cell exhaustion, and enhances CD8+ T cell migration into the TME in a C-X-C motif chemokine receptor 3 (CXCR3)-dependent manner. CONCLUSION: GARP contributes to multiple aspects of immune resistance in cancer. Anti-human GARP antibody PIIO-1 is an efficacious and safe strategy to block GARP-mediated LTGFß activation, enhance CD8+ T cell trafficking and functionality in the tumor, and overcome primary resistance to anti-PD-1 ICB. PIIO-1 therefore warrants clinical development as a novel cancer immunotherapeutic.
Asunto(s)
Neoplasias , Microambiente Tumoral , Animales , Linfocitos T CD8-positivos/metabolismo , Glicoproteínas , Humanos , Inhibidores de Puntos de Control Inmunológico , Proteínas de la Membrana , Ratones , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
BACKGROUND: Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19) through direct lysis of infected lung epithelial cells, which releases damage-associated molecular patterns and induces a pro-inflammatory cytokine milieu causing systemic inflammation. Anti-viral and anti-inflammatory agents have shown limited therapeutic efficacy. Soluble CD24 (CD24Fc) blunts the broad inflammatory response induced by damage-associated molecular patterns via binding to extracellular high mobility group box 1 and heat shock proteins, as well as regulating the downstream Siglec10-Src homology 2 domain-containing phosphatase 1 pathway. A recent randomized phase III trial evaluating CD24Fc for patients with severe COVID-19 (SAC-COVID; NCT04317040) demonstrated encouraging clinical efficacy. METHODS: Using a systems analytical approach, we studied peripheral blood samples obtained from patients enrolled at a single institution in the SAC-COVID trial to discern the impact of CD24Fc treatment on immune homeostasis. We performed high dimensional spectral flow cytometry and measured the levels of a broad array of cytokines and chemokines to discern the impact of CD24Fc treatment on immune homeostasis in patients with COVID-19. RESULTS: Twenty-two patients were enrolled, and the clinical characteristics from the CD24Fc vs. placebo groups were matched. Using high-content spectral flow cytometry and network-level analysis, we found that patients with severe COVID-19 had systemic hyper-activation of multiple cellular compartments, including CD8+ T cells, CD4+ T cells, and CD56+ natural killer cells. Treatment with CD24Fc blunted this systemic inflammation, inducing a return to homeostasis in NK and T cells without compromising the anti-Spike protein antibody response. CD24Fc significantly attenuated the systemic cytokine response and diminished the cytokine coexpression and network connectivity linked with COVID-19 severity and pathogenesis. CONCLUSIONS: Our data demonstrate that CD24Fc rapidly down-modulates systemic inflammation and restores immune homeostasis in SARS-CoV-2-infected individuals, supporting further development of CD24Fc as a novel therapeutic against severe COVID-19.
Asunto(s)
Antígeno CD24/uso terapéutico , COVID-19/prevención & control , Síndrome de Liberación de Citoquinas/prevención & control , Inflamación/prevención & control , SARS-CoV-2/efectos de los fármacos , Anciano , Alarminas/inmunología , Alarminas/metabolismo , Antígeno CD24/química , COVID-19/inmunología , COVID-19/virología , Síndrome de Liberación de Citoquinas/inmunología , Síndrome de Liberación de Citoquinas/metabolismo , Método Doble Ciego , Femenino , Proteína HMGB1/inmunología , Proteína HMGB1/metabolismo , Proteínas de Choque Térmico/inmunología , Proteínas de Choque Térmico/metabolismo , Homeostasis/efectos de los fármacos , Homeostasis/inmunología , Humanos , Inflamación/inmunología , Inflamación/metabolismo , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/metabolismo , Células Asesinas Naturales/virología , Masculino , Persona de Mediana Edad , SARS-CoV-2/inmunología , SARS-CoV-2/fisiología , Solubilidad , Linfocitos T/inmunología , Linfocitos T/metabolismo , Linfocitos T/virología , Resultado del TratamientoRESUMEN
Butea monosperma (Lam.) Taub. has been applied to treat inflammatory, metabolic, and infectious diseases. However, the antiobesity effects of B. monosperma (Lam.) Taub. flower (BMF) and the underlying mechanisms have not been determined. In this study, we analyzed the various extraction procedures, investigated the antiobesity effects, and identified the main chemical constituents of BMF. The BMF was subjected to acid hydrolysis in 5% H2SO4 in methanol at 50°C for 48 h and partitioned with ethyl acetate. The acid-hydrolyzed BMF ethyl acetate extracts (BMFE) strongly induced the expression of uncoupling protein 1 (Ucp1) and other thermogenic genes in C3H10T1/2 adipocytes. Daily oral administration of 70 mg/kg BMFE (BMFE70) to mice with diet-induced obesity resulted in less body weight gain, increased glucose tolerance, higher rectal temperature, and increased oxygen consumption. Qualitative and quantitative analyses along with treatments in Akt1 knockout mouse embryonic fibroblasts indicate that butein is a major active ingredient of BMFE, which stimulates Ucp1 gene expression. These data show the effects of butein-containing B. monosperma flower extract on thermogenesis and energy expenditure, further suggesting the potential role of BMFE as a functional ingredient in obesity and related metabolic diseases.